Mollison’s Blood Transfusion in Clinical Medicine - part 8 pptx

Transfusion of haematopoietic cells
The object of transfusing allogeneic haematopoietic
progenitor cells is to establish a permanent graft of
transfused progenitor cells in the recipient. The fate
of allogeneic progenitor cells infused into the venous
circulation depends on their ability to traffic to sites of
haematopoietic tolerance (‘microenvironment’) and on
managing two immunological phenomena: (1) the rejec-
tion of donor progenitor cells by the host immuno-
logical response and (2) an immunological reaction
of grafted immunologically competent cells against
the host: GvHD. Both of these reactions depend on the
degree of histocompatibility between donor and recipi-
ent and also on the immunological competence of the
recipient. Engraftment and kinetics also depend on
patient age, disease status, the preparative regimen,
GvHD prophylaxis and the cellular content of the graft.
Bone marrow was the original source of progenitor
cells for haematopoietic grafting, but mobilized
peripheral blood and cord blood have gradually sup-
planted marrow as a source of PC. The engraftment
potential of the component is commonly designated
in terms of mononuclear cells that express the CD34
antigen, the cluster designation of a transmembrane
glycoprotein present on haematopoietic progenitor cells
(Krause et al. 1996), although accessory cells in the
graft clearly play an important role (Ash et al. 1991).
Cells that express CD34 include lineage-committed
haematopoietic progenitors, multipotent progenitors
and possibly pluripotent stem cells as well. Flow
cytometric assays are used to quantify CD34

+
cells
in both the donor and the component. However,
problems with interlaboratory accuracy and repro-
ducibility, especially of different PC sources, have been
notorious, even with the adoption of a standardized
technique (Sutherland et al. 1996; Keeney et al. 1998).
Peripheral blood-derived progenitor cells
Peripheral blood-derived progenitor cells (PBPCs)
were reported to circulate in mammalian blood as
early as 1909 (Maximow 1909), and the ability of cir-
culating cells to repopulate a lethally irradiated animal
was demonstrated in a parabiotic rat model in 1951
(Brecher and Cronkite 1951). However, circulating
haematopoietic progenitor cells were not confirmed in
human blood until the 1970s (McCredie et al. 1971).
Collection of PBPCs obtained from peripheral blood
by leucapheresis (see Chapter 17) has now all but
replaced infusion of bone marrow. PBPCs have the
advantages of engrafting more rapidly and sparing the
donor a general anaesthetic, which result in lower
morbidity and cost (Kessinger et al. 1989; Azevedo
et al. 1995; Bensinger et al. 1995; Korbling et al. 1995;
Schmitz et al. 1995). Allogeneic PBPCs have a higher
CD34
+
cell content than does marrow, which, inde-
pendent of stem cell source, increases patient survival
while reducing transplant-related mortality and relapse
(Mavroudis et al. 1996; Bahceci et al. 2000; Zaucha

et al. 2001). A theoretical argument against the use
of PBPCs is the greater number of ‘T’ lymphocytes
that contaminate these collections, compared with the
number of T cells in bone marrow, suggesting the
possibility of an increased risk of severe acute GvHD.
Indeed, although the risk of acute GVHD after PBPCs
is similar to that observed among historic bone mar-
row transplant (BMT) controls (Pavletic et al. 1997;
Przepiorka et al. 1997), the probability and severity
of chronic GVHD (cGVHD) appear to be increased
(Bacigalupo et al. 1996; Flowers et al. 2002).
Liquid storage and cryopreservation
Collection of allogeneic PBPCs is ordinarily scheduled
to coincide with the conclusion of the patient’s
preparatory regimen, so that the graft can be infused
while fresh. Most centres opt to transfuse the cells as
soon as possible. Refrigerated storage of unmanip-
ulated mobilized collections at 2–6°C for 24 h and as
long as 72 h results in little detectable loss of the in
vitro functional properties (Beaujean et al. 1996;
Moroff et al. 2004). PBPCs, like bone marrow, can
be cryopreserved by slow cooling (1–3°C/min) in the
presence of the cryprotectant dimethylsulphoxide
(DMSO), variable amounts of plasma, with or without
hydroxyethyl starch (Hubel 1997). Grafts can be stored
at –80°C, but are usually placed in liquid nitrogen at
–140°C or colder, at which engraftment potential is
preserved for years.
Cryopreserved grafts are thawed in a waterbath at
37–40°C and infused through a 170-µ filter. Prolonged

post-thaw storage is inadvisable, as prolonged expos-
ure to 10% DMSO may harm the cells. Storage up
to 1 h does not reduce viability or colony-forming
activity (Rowley and Anderson 1993). Rapid infusion
of DMSO has been associated with flushing, nausea,
vomiting, diarrhoea and hypotension, probably the
CHAPTER 14
628
result of histamine release. Reversible encephalopathy
has been reported when doses have approached 2 g/kg,
so that caution is advisable when large volumes of
PBSCs are thawed (Dhodapkar et al. 1994). The graft
can be washed free of cryoprotectant, but progenitor
cells may be lost in the process.
Cord blood progenitor cells
Umbilical cord blood is a rich source of progenitor
cells (Knudtzon 1974; Broxmeyer et al. 1989a). The
use of cord blood progenitor cells (CBPCs) has import-
ant real and potential advantages. The number of
donors is unlimited, procurement is easy and inexpens-
ive and the cells can be HLA typed and preserved in
liquid nitrogen. Human CBPCs with high proliferative
capacity and NOD/SCID mouse engrafting ability can
be stored frozen for > 15 years, and probably remain
effective for clinical transplantation (Broxmeyer et al.
2003). In addition, because many of the functions of
the immunologically competent cells in cord blood are
not fully developed, the chance of their inducing
GvHD appears to be diminished (Szabolcs et al. 2003).
Even after the transplantation of CBPCs from unrelated

donors, mismatched for two, or as many as three, HLA
antigens, the risk of severe GvHD seems to be low
(Wagner 1995).
Umbilical cord blood banking
Umbilical cord blood is collected by either the obstetri-
cian or the midwife in utero during the third stage of
delivery or ex utero after delivery of the placenta by
trained nurses or technologists (Wall et al. 1997;
Fraser et al. 1998). Collection volume and cell yield
appear to be similar with both methods (Lasky et al.
2002). A maternal blood specimen is screened for
markers of transmissible disease, and a sample from
the unit is cultured, HLA typed, analysed for cell
count, viability and in many instances CD34
+
cell
number and colony count by culture. Suitable units
are processed to remove red cells and plasma and are
frozen at a controlled rate and stored in liquid nitrogen
(Armitage et al. 1999).
Related donor transplants
The first successful transplantation of CBPCs from
an HLA-identical sibling was given to a patient with
Fanconi’s anaemia (Broxmeyer et al. 1989a). In 44
paediatric transplantations of CBPCs from sibling
donors, patients receiving HLA-identical or 1-antigen
mismatched grafts showed an actuarial probability of
engraftment of 85% at 50 days after transplantation;
there were no instances of late graft failure (Wagner
et al. 1995). The median total nucleated cells per

kilogram (TNC/kg) was 5.2 × 10
7
. The probability
of GvHD at 100 days post transplant was 3% and of
chronic GvHD at 1 year was 6%. The probability of
survival with a median follow-up of 1.6 years was
72%. Among 102 children with acute leukaemia
transplanted by the Eurocord collaborative investiga-
tors, 42 received a graft from a related cord blood
donor; 12 of these were HLA mismatched (Locatelli
et al. 1999). Nucleated cell dose (> 3.7 × 10
7
/kg) cor-
related with engraftment; two-year survival was 41%.
Rocha and co-workers (2000) compared 113 recipi-
ents of HLA-identical sibling CBPC transplants for
malignant disease with records of 2052 siblings trans-
planted with bone marrow between 1990 and 1997.
Although the umbilical cord blood (UCB) had a
significantly longer delay in recovery of neutrophil
and platelet reconstitution, no significant difference in
survival and a significantly lower risk of GvHD and
chronic GvHD was reported in the CBPC group. Bone
marrow recipients received nearly 10-fold the total
nucleated cells per kilogram body weight (TNC/kg).
Of 44 children with non-malignant conditions (thalas-
saemia, sickle cell disease), two-year survivals were
79% and 90% respectively (Locatelli et al. 2003). One
child with sickle cell disease and seven with thalas-
saemia failed to sustain engraftment. Four children

suffered acute grade II GvHD.
Unrelated donor transplants
Several thousand CBPC transplants have been per-
formed and more than 100 000 umbilical cord com-
ponents are available worldwide. With the growth of
public (‘unrelated’) CBPC banks, the number of CBPC
transplants from unrelated donors now exceeds that
from related donors. In one early series, a high rate
of engraftment (23 out of 25 cases) was observed in
children infused with allogeneic CBPC despite the
donor–recipient pair discordance of 1–3 HLA antigens
(Kurtzberg et al. 1996). A retrospective analysis of 562
unrelated CBPC transplants found that engraftment
exceeded 80% and survival rate was 61%; pre-freeze
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629
cell count of the graft ranged from 0.7 to 10 TNC/kg
(Rubinstein et al. 1998). The number of nucleatedcord
blood cells that were transfused per kilogram of the
recipient’s weight emerged as the main influence on
engraftment. A retrospective analysis of 537 paediatric
CBPC transplants from the Eurocord Registry, includ-
ing 138 related transplants and 291 unrelated donors,
reported similar results (Gluckman and Locatelli
2000). Laughlin and co-workers (2001) reported
CBPC transplants in 68 adult recipients who received
a median of 2.1 × 10
7
TNC/kg. TNC number per
kilogram correlated with rapidity of engraftment and

high CD34
+
number was associated with event-free
survival. Overall survival (22 months) was 28%. As
expected from the experience with bone marrow
transplantation, GvHD is significantly higher in the
setting of grafts from unrelated donors and depends as
well upon the age of the recipient, the degree of histo-
compatibility between donor and recipient, the nature
of the preparatory regimen and a variety of other fac-
tors. In this series, grades III and IV GvHD occurred in
20% and chronic GvHD in 36%.
Reconstitution of adult recipients of cord blood
CBPCs have so far been used primarily for children
and doubt has been expressed as to whether the num-
ber of progenitor cells in cord blood from a single
donor will be sufficient to repopulate the majority of
large adults who require a transplant. Most analyses
indicate that the key clinical outcomes (days to
neutrophil engraftment, platelet engraftment, severe
GvHD and disease-free survival) are all superior in
younger patients; age-related outcomes are widely
attributed to the number of nucleated cells in a single
unit of cord blood (Laughlin et al. 2001). There is,
as yet, no quantitative assay for the progenitor cell
subset that has the capacity for long-term bone mar-
row repopulation. On the other hand, the number of
progenitor cells that can be assayed (CFU-GM, CFU-
GEMM, etc.) is large enough, suggesting that the
number of the more primitive progenitor cells may be

sufficient (Broxmeyer 1995). There is evidence that the
total cellular content of placental cord blood (PCB)
grafts is related to the speed of engraftment, though
the total nucleated cell (TNC) dose is not a precise pre-
dictor of the time of neutrophil or platelet engraft-
ment. It is important to understand the reasons for the
quantitative association and to improve the criteria for
selecting PCB grafts by using indices that are more
precisely predictive of engraftment (Rubinstein et al.
1998). The post-transplant course of 204 patients who
received grafts evaluated for haematopoietic colony-
forming cell (CFC) content among 562 patients
reported previously were analysed using univariate
and multivariate life-table techniques to determine
whether CFC doses predicted haematopoietic engraft-
ment speed and risk for transplant-related events more
accurately than the TNC dose. Actuarial times to
neutrophil and platelet engraftment were shown to
correlate with the cell dose, whether estimated as TNC
or CFC per kilogram of recipient’s weight. CFC associ-
ation with the day of recovery of 500 neutrophils/µl
was stronger than that of the TNC. In multivariate
tests of speed of platelet and neutrophil engraftment
and of probability of post-transplantation events,
the inclusion of CFCs in the model displaced the
significance of the high relative risks associated with
TNC. The CFC content of PCB units is associated
more rigorously with the major covariates of post-
transplantation survival than is the TNC and is,
therefore, a better index of the haematopoietic content

of PCB grafts (Migliaccio et al. 2000). A positive cor-
relation between CD34
+
cells and circulating day-14
colony counts (CFU-GM) has been reported suggesting
that with umbilical cord progenitor cells (UCPCs), as
with PBPC, CD34 is a reliable measure of haematopoi-
etic potential (Payne et al. 1995; Siena et al. 2000).
Data from 102 patients identified CD34
+
cell dose as
the only factor that correlated with rate of engraftment
(Wagner et al. 2002). Studies from Spain and Japan
of small numbers of adults with haematological malig-
nancies report promising rates of engraftment and
disease-free survival (Sanz et al. 2001; Ooi et al. 2004).
Progenitor cell expansion
If the number of progenitor cells in cord blood proves
to be scarcely sufficient for repopulation in many
adults, the possibility of expanding the number by
culture in vitro has been proposed (Apperley 1994;
Broxmeyer et al. 1995) and several groups are devel-
oping methods to do so (Kogler et al. 1999; Pecora
et al. 2000; Jaroscak et al. 2003). Whether the most
important primitive progenitor cells are expanded by
culture cannot be established in vitro. As yet, no
evidence has confirmed that increase in engraftment
kinetics or expansion of stem cells has been achieved,
CHAPTER 14
630

and the possibility of increased frequency of GvHD
with some expansion methods has been raised (Shpall
et al. 2002; Jaroscak et al. 2003).
Plasma from cord blood has been found to increase
the self-renewal capacity of stem cells in vitro (Carow
et al. 1993). Cord blood plasma, but not plasma from
adults or fetal calf serum, had this effect and cord
blood plasma also increased the expansion in vitro of
the number of progenitor cells induced by growth fac-
tors (Bertolini et al. 1994). Furthermore, CBPCs fully
retain this expansion potential after cryopreservation
(Bertolini et al. 1994).
Use of multiple cord blood collections
Because limited cell dose compromises may comprom-
ise the outcome of adult UCB transplants, multiple
cord blood units have been combined to augment
the dose. Zanjani and co-workers (2000) have trans-
planted human UCB from multiple donors in a fetal
sheep model. Short-term donor engraftment derived
from both donors,but for long-term haematopoiesis, a
single donor predominated.
Multidonor human UCB transplants using up to
12 units have been published (Ende and Ende 1972;
Shen et al. 1994). Weinreb and co-workers (1998)
reported that a unit that was partially HLA matched
predominated in a recipient who received a combina-
tion of 12 units. Another patient with advanced acute
lymphocytic leukaemia received a mismatched, unre-
lated UCB transplant using units from two donors and
achieved a complete remission with double chimerism,

which persisted until relapse (De Lima et al. 2002).
Barker and co-workers (2005) have augmented graft
cell dose by combining two partially HLA-matched
units. Twenty-three patients with high-risk haemato-
logical malignancy received 2 UCB units (median
infused dose, 3.5 × 10
7
NC/kg) and 21 evaluable
patients engrafted at a median of 23 days. At day 21,
engraftment was derived from both donors in 24%
of patients and a single donor in 76% of patients.
One unit predominated in all patients by day 100.
Although neither nucleated or CD34
+
cell doses nor
HLA match predicted which unit would predominate,
the predominating unit had a significantly higher
CD34
+
dose. The result is similar to the predominant
lymphocyte chimerism that persists in trauma patients
who receive multiple blood transfusions (Lee et al.
1999).
Law, ethics, related banks and genetic selection
Controversy continues regarding the propriety of
related (‘private’) CBPCs for which the family pays to
have the infant’s cells cryopreserved for future use, as
contrasted with unrelated (‘public’) banks, in which
donated cords are stored for general use (Burgio et al.
2003). Both systems have their adherents and they

should not be mutually exclusive. Although most
related banks with commercial origins have sought
participation from expectant mothers who agree to
pay for storage of a cord from their newborn infant,
others have been supported by federal grants (Reed
et al. 2001). The infrequent utilization of a related
cord blood unit does minimize its utility. The probabil-
ity that the cord blood will be of use in a family with
no history of blood or genetic disease is low (estimated
at 1/200 000); moreover, one’s own stem cells may be
immunologically less potent than those of an unrelated
donor for treating neoplastic diseases. However,
several such transplants have been performed success-
fully and prohibiting such storage despite appropriate
informed consent seems curiously patronizing. The
legal issues regarding property rights have been dis-
cussed (Munzer 1999). In vitro fertilization and pre-
natal genetic diagnosis to select an embryo donor on
the basis of specific, desirable disease and HLA charac-
teristics have been used successfully to treat a child
with Fanconi’s anaemia (Grewal et al. 2004).
Effect of ABO incompatibility of grafted cells
As ABO and HLA antigens are inherited independ-
ently, ABO incompatibility may occur in 20–40% of
HLA-matched allogeneic haematopoietic stem cell
transplants. ABO incompatibility between donor pro-
genitor cells and the recipient’s plasma is not a barrier
to successful transplantation (Storb et al. 1977;
Buckner et al. 1978). In a series of 12 subjects who
received major ABO-incompatible marrow, not one

rejected the graft and the incidence of GvHD was no
higher than in subjects who received ABO-compatible
marrow (Hershko et al. 1980).
With major ABO-incompatible marrow grafts,
defined as incompatibility of donor ABO antigens with
the recipient’s immune system, steps must be taken to
prevent an acute haemolytic reaction due to lysis of
incompatible red cells contained in the progenitor cell
graft. To avoid haemolysis, grafts are purged of red
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
631
cells. A satisfactory method has been described
(Warkentin et al. 1985). An alternative method of
removing red cells from marrow uses a cell separator
(Blacklock et al. 1982). When PBPC or CBPC are used,
the number of contaminating red cells is small.
Delayed donor red cell engraftment and pure red
cell aplasia are well-recognized complications of major
ABO-incompatible haematopoietic stem cell trans-
plantation (Hows et al. 1983; Sniecinski et al. 1988;
Stussi et al. 2002; Griffith et al. 2005). Donor red
blood cell chimerism is delayed as long as three-fold
(median 114 days) following reduced-intensity non-
myeloablative compared with myeloablative condi-
tioning for transplant and the delay correlates with the
recipient anti-donor isohaemagglutinin titre (Bolan
et al. 2001a). Late-onset red cell aplasia, most likely
related to delayed lymphoid engraftment, may occur
(Au et al. 2004). In some patients, thrombopoiesis may
be delayed as well (Sniecinski et al. 1988).

After transplants of major ABO-incompatible grafts,
the direct antiglobulin test (DAT) may turn positive
after about 3 weeks. If substantial numbers of donor red
cells enter the circulation, transient immune-mediated
haemolysis may result (Sniecinski et al. 1987). Anti-A
and anti-B may remain demonstrable in the recipient’s
plasma for some months and the DAT may remain
positive during this time. In patients with minor
ABO-incompatible transplants, defined as those in
which the recipient antigens are incompatible with the
donor’s immune system, haemolysis may develop 1–
2 weeks after transplantation owing to lysis of ABO-
incompatible recipient cells as the donor immune
lymphocytes engraft (Hows et al. 1986). This type of
haemolysis has been seen only in patients receiving
ciclosporin and prednisone GvHD prophylaxis, and
may not develop in patients receiving methotrexate
(Gajewski et al. 1992). Massive immune haemolysis
may occur, and fatalities can be avoided by early,
vigorous donor-compatible red cell transfusion until
haemolysis subsides (Bolan et al. 2001a). Reactions
are most common and severe when the donor is group
O and the recipient group A, but neither blood group
nor agglutinin titre reliably predict clinical severity. In
some of the patients, haemolysis caused by anti-A or
anti-B (or both) destroys transfused group O cells,
probably as a result of activated complement compon-
ents affixing the group O cells (bystander haemolysis).
Haemolysis has also been observed when the donor
lymphocytes produce anti-D, etc. (see Chapter 11).

Special consideration of the ABO group of compon-
ents transfused to patients receiving ABO-incompatible
grafts should begin with the initiation of the prepar-
atory regimen to ensure that blood is compatible
with both donor and recipient (Table 14.1). With bi-
directional (major–minor) incompatibility, red cell
transfusions should be limited to group O. Platelet
concentrates administered to adults may be of any
blood group, although plasma reduction may be pru-
dent, especially for large-volume group O platelets.
Plasma-compatible platelets should be used for infants
and children. Some centres use the soluble antigens
contained in plasma to neutralize isohaemagglutinins.
As intravenous immunoglobulin contains variable
titres of red cell antibodies, especially of anti-A, some
centres screen for alloantibodies, whereas others avoid
high-dose IVIG for group A recipients during the post-
transplant period.
Donor lymphocyte infusion
Lymphocytes have been studied more often as blood
component contaminants responsible for adverse
effects than as therapeutic cells. However, some ostens-
ibly adverse effects of mononuclear cell infusions can
be exploited for therapeutic benefit. The mechanisms
involved in TA-GvHD (see Chapters 13 and 15) are
probably responsible for the graft-versus-malignancy
effect in allogeneic stem cell transplantation. Studies in
animal models are consistent with the observation by
Barnes and Loutit (1957) that transplanted bone mar-
row has immune activity against residual leukaemia

(Kloosterman et al. 1995). Clinical experience with
haematopoietic transplantation has been consistent with
the presence of antileukaemic activity also in humans,
now commonly referred to as the graft-versus-leukaemia
effect (GvL). The term ‘adoptive immunotherapy’
was coined by Mathé (1965) who used both marrow
transplants and leucocyte infusions to treat acute
leukaemia. Kolb and co-workers (1990) provided
direct clinical evidence for GvL: the transfusion of
donor lymphocytes in conjunction with the adminis-
tration of alpha-interferon (IFN-α) induced cytoge-
netic remission in three patients with CML in relapse
following allogeneic bone marrow transplantation.
Numerous independent studies confirm the GvL effect
in CML (Slavin et al. 1992; Bar et al. 1993; Porter et al.
1994). In both European and North American reg-
istries, more than 90% of the patients received original
CHAPTER 14
632
grafts and subsequent donor lymphocyte infusion
(DLI) from related donors, typically from an HLA-
identical sibling. The results reported by the European
Group for Blood and Marrow Transplantation
(27 centres, 135 patients, 75 evaluable with CML) are
similar to those reported by the North American
Multicenter Bone Marrow Transplantation Registry
(25 centres, 140 patients, 55 evaluable with CML):
DLI-induced clinical remission at a rate approaching
80%, and molecular remission (inability to detect bcr-
abl mRNA transcript using polymerase chain reaction)

in nearly all patients entering clinical remission (Kolb
et al. 1995; Collins et al. 1997).
Infusion of small numbers of lymphocytes (10
7
)
(‘bulk dose’) usually suffices, and excess cells collected
by leucapheresis are often aliquoted and stored for
repeated treatment if needed. The host’s circulation,
which often contains a mixture of both donor and host
cells during chronic phase relapse, typically converts
to cells of only donor origin. The time to remission
ranges from 1 to 9 months, with a mean of about
3 months (Kolb et al. 1995; Collins et al. 1997). Nearly
all responses are seen within 8 months after DLI, and
the probability of remaining in remission at 2 and
3 years is 90% and 87% respectively (Kolb et al. 1995;
Collins et al. 1997). Although late relapses occur and
toxicity may be significant, DLI efficacy is durable in
surviving patients with CML: 26 out of 39 (67%) pati-
ents were alive at follow-up with 25 (96% of survivors)
remaining in complete remission (Porter et al. 1999).
Separating graft-versus-leukaemia from
graft-versus-host-disease
GvHD occurring after DLI correlates strongly with
antileukaemic response. However, the GvL effect and
GvHD may be separable, and GvHD may not be
required for durable disease remission (Weiss et al.
1994; Rocha et al. 1997; Slavin et al. 2002). Murine
studies suggest that the rate of GvHD is inversely
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS

633
Blood group Blood products
Recipient Donor Red cells* Platelets
†
FFP*
A B O Any AB
‡
A O O Any A
§
A AB O,A Any AB
‡
B A O Any AB
‡
B O O Any B
§
B AB O,B Any AB
‡
O A O Any A
‡
O B O Any B
‡
O AB O Any AB
‡
AB A O,A Any AB
§
AB B O,B Any AB
§
AB O O Any AB
§
Rh pos Rh neg Rh neg

††
Rh neg** N/A
Rh neg Rh pos Rh pos
¶
Rh pos N/A
* Restrictions for ABO- and/or Rh-incompatible transplant recipients supported
with blood components during pre-transplant conditioning and during the
post-transplant period.
†
Use any ABO group for platelet support for adults. Use plasma-compatible
components for children.
‡
Plasma may be transfused to neutralize isohaemagglutinin(s).
§
Graft plasma depleted, no plasma neutralization required.
¶
Rh-positive components initiated on the day of transplant.
** Rh-negative platelets preferred.
††
Rh-negative red cells preferred during the pre-transplant conditioning regimen
and post transplant.
Table 14.1 ABO and/or Rh-
incompatible progenitor cell:
transplant transfusion restrictions.
proportional to the interval between transplant and
DLI (Johnson et al. 1999). These considerations have
popularized escalating dose DLI regimens (Dazzi et al.
2000). Although probability of achieving remission
in relapsed CML does not differ, the escalating dose
regimen is associated with a lower incidence of GvHD.

DLI is typically initiated early in disease as soon as
disease recurrence is anticipated, and a starting dose
of 10
5
T cells/kg is escalated 10-fold at 2- to 4-week
intervals (Weiss et al. 1994). Efforts to modify the
composition of donor-derived lymphocytes (DDLs)
have focused on selective CD8-positive T-cell depletion,
which appears to be more effective than non-selective
T-cell depletion in reducing GvHD while preserving
GvL (Soiffer et al. 2002).
Target antigen as the primary determinant of
efficacy and toxicity
Falkenburg and colleagues (1999) reported the first
successful treatment of relapsed accelerated CML using
in vitro-expanded leukaemia-specific lymphocytes.
Presumably, cell selection and culture restored the
anti-tumour activity and specificity against leukaemic
cells that weakened with disease progression (Smit
et al. 1998). Successful salvage therapy of a child with
previously DLI-resistant CML by using DDL pulsed in
vitro with a mixture of normal irradiated lymphocytes
obtained from the child’s parents has been reported.
Efficacy and toxicity in viral diseases
Walter and co-workers (1995) have used in vitro-
stimulated, culture-expanded, CMV-specific donor-
derived cytotoxic T cells to successfully reconstitute
cellular immunity against CMV in 11 out of 14 allo-
geneic marrow transplant patients. The DLI therapy
consisted of four escalating cell doses (0.33, 1.0, 3.3

and 10.0 × 10
8
cells) administered at weekly intervals
beginning at days 30–40 after transplantation. DLI-
associated toxicity, CMV disease and CMV viraemia
were not observed. The results have been confirmed in
similar studies (Einsele et al. 2002; Roback et al. 2003).
Epstein–Barr virus-related lymphoproliferative
disorders
The incidence of Epstein–Barr virus-related lympho-
proliferative disorders (EBV-LPDs) occurring in T cell-
depleted transplants has been estimated at 6–12%,
and secondary lymphomas occurring in this clinical
setting respond readily to DLI at a dose approximately
10-fold smaller than that typically used for activity
against the primary leukaemia. Sustained clinical
remissions have been achieved with only mild GvHD,
and patients have often required no additional mainten-
ance therapy (Papadopoulos et al. 1994; Wagner et al.
2004). EBV-specific lymphocyte infusions have suc-
cessfully treated EBV-LPD and EBV-positive Hodgkin
disease (Rooney et al. 1998; Bollard et al. 2004).
The transfusion of plasma components
Fresh-frozen plasma
Fresh-frozen plasma (FFP) is plasma obtained from a
single donor by normal donation or plasmapheresis
and frozen within 6 h of collection to a temperature
of –30°C or below. FFP contains all circulating coagu-
lation factors in the concentration present in fresh
plasma, and haemostatic activity is maintained for a

year or longer, depending upon the storage temper-
ature. Once thawed, FFP must be stored at 4 ± 2°C for
no longer than 24 h before infusion. FFP must not be
refrozen, but once thawed (or after 1 year of storage
and thaw), it can be used as single-donor plasma, i.e.
not to replace labile coagulation factors, for as long
as 5 weeks. The concentration of coagulation factor,
the citrate concentration and the volume of each unit
may vary depending on the characteristics of the donor
and of the collection. In 51 units collected by apheresis
from plasma donors, factor concentrations at the fifth
and ninety-fifth percentile measured: V (690–1270 units/
l); VII (830–1690 units/l); fibrinogen 1800–3700 µg/l;
antithrombin (920–1290 units/l) (Beeck et al. 1999).
Risks of fresh-frozen plasma
Allergic reactions may occur after transfusion of FFP,
of which the most serious is severe anaphylaxis, which
may develop in IgA-deficient patients with class-
specific anti-IgA (Chapter 15). Such reactions are rare.
Transfusion-related acute lung injury (TRALI) may
occur when the FFP contains strong leucocyte antibod-
ies (see Chapter 15). The other main risk of treatment
with FFP is the transmission of infectious agents, par-
ticularly viruses such as hepatitis B and C viruses, HIV,
parvovirus and West Nile virus. Owing to donor selec-
CHAPTER 14
634
tion and the availability of methods of inactivating
viruses that are used to treat whole plasma in some
countries, the risk of transmitting viruses has greatly

decreased. However, the problem of inactivating
non-lipid-enveloped viruses and the transmission of
non-viral agents remains (Chapter 16). Therefore, FFP
should still be used only when no safer alternative
exists (Shimizu and Robinson 1996).
Precautions to be taken before infusion
FFP containing potent anti-A or anti-B agglutinins or
haemolysins, or FFP that has not been tested for their
presence, should not be given to recipients with corres-
ponding red cell antigens. Fresh plasma, which is now
rarely used, may contain red cells, so that appropriate
measures should be taken to prevent immunization of
D-negative women of childbearing age. There is no
credible evidence that FFP presents such a risk.
Indications for fresh-frozen plasma:
overused and abused
There is no justification for the use of FFP as a volume
expander because safer alternatives (colloids and
crystalloids) are available.
Factor V deficiency. No concentrate of factor V is
available and FFP can be used as a source of factor V.
Cryoprecipitate-poor plasma contains 80% of the
amount of factor V in FFP and can be used as an
alternative for FFP (Hellings 1981).
Severe liver disease. The liver is the major site of syn-
thesis of coagulation factors II, V, VII, IX, X, XI, XII
and fibrinogen as well as of factors with potential
antithrombotic activity such as proteins C, S and
antithrombin. Patients with severe liver disease may
experience defects in factor synthesis and increased

factor degradation that can result in generalized bleed-
ing. Unfortunately, studies regarding the predictability
of bleeding and its most effective management by
transfusion in the presence of different degrees of hep-
atic impairment are old, poorly documented or both.
Most recommendations rely upon expert opinion.
What seems clear is that no single coagulation assay
predicts bleeding (Spector and Corn 1967). Prolonga-
tions of the prothrombin time (PT) and activated
partial thromboplastin time (aPTT) are the most fre-
quent abnormalities among the commonly performed
clotting tests in patients with liver disease, and may
reflect impaired protein synthesis, vitamin K deficiency
or even disseminated intravascular coagulation (DIC).
The presence of an abnormal test does not necessitate
intervention, especially in the non-bleeding patient.
Furthermore, in 30 patients with chronic liver disease,
a moderate-dose plasma infusion (12 ml/kg or about
4 units) did not return the PT and aPTT to normal
(Mannucci et al. 1982). In the bleeding patient, doses
calculated to bring coagulation factor levels to the
20–30% range (20 ml/kg or 6–7 units) may be required
as frequently as every 4–6 h to correct the abnormal
coagulation tests (Spector et al. 1966). The routine use
of FFP as prophylaxis for excessive surgical bleeding in
patients with severe liver disease finds few supporters
and less evidence of benefit (Oberman 1990).
Treatment of acquired deficiencies of factors II,
VII, IX and X due to treatment with
anticoagulants: warfarin reversal

The major risk of anticoagulant therapy is haemor-
rhage. For patients treated with the oral vitamin K
antagonists, the annual risk of severe haemorrhage
ranges from 1–5% (Levine et al. 2001). The intensity
of anticoagulation (including poor control), its dura-
tion and, in some studies, advanced age and cerebrov-
ascular disease all increase the bleeding risk (Landefeld
and Goldman 1989). For the bleeding patient or the
patient at extreme risk, urgent reversal of vitamin K
antagonists can be achieved with plasma infusion to
bring factor levels to 30–40%. The volume of plasma
can be calculated easily based on the patient’s body
weight but as 6 units or more (1500 ml) may be required
to reverse anticoagulation in an adult, volume consid-
erations may make a prothrombin complex concentrate
(PCC) the preferred infusion (Schulman 2003). Recom-
binant VIIa has also been used in this situation (Deveras
and Kessler 2002) (see Chapter 18). Intravenous vita-
min K
1
, the specific warfarin antagonist, may require
12 h or more to be fully effective (Nee et al. 1999).
Disseminated intravascular coagulation:
a vehicle on the road to multi-organ
dysfunction syndrome
Disseminated intravascular coagulation is a condition
in which the intravascular activation of the clotting
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
635
cascade leads to the final common pathway of sus-

tained and excessive thrombin generation. Liberated
thrombin and proteolytic enzymes bring about the
intravascular production of fibrin and deposition of
platelets, with activation of the fibrinolytic system and
an increased level of fibrin degradation products
(FDPs) (Levi et al. 2001). In mild DIC the platelet
count and the levels of clotting factors may be normal
due to compensatory increases in production. As DIC
becomes more severe, the levels of clotting factors
and platelets fall, and a state that may be described as
decompensated DIC may lead to multi-organ dysfunc-
tion syndrome (MODS).
DIC may be precipitated by a wide variety of stimuli,
most related to the entry of tissue thromboplastins into
the circulation, for example after abruptio placentae,
crush injury, head trauma and snake envenomation.
Other conditions associated with DIC include infec-
tions, malignancies, amniotic fluid embolism, giant
haemangioma and intravascular lysis of incompatible
red cells (Levi et al. 2004).
The cardinal principle of treatment of DIC remains
elimination of the underlying cause as, once this
has been accomplished, haemostasis usually returns to
normal. When the underlying cause cannot be treated
effectively, uncontrollable bleeding may result. The
transfusion of blood may be essential and the replace-
ment of clotting factors has to be considered. This
replacement should be guided by coagulation assays
and fibrinogen levels. If levels of clotting factors are
severely reduced (< 25%), FFP may be given and if the

fibrinogen concentration falls below 60 mg/dl, cryo-
precipitate may be helpful. An initial dose of 10 bags,
to provide 4–6 g of fibrinogen, has been suggested
(Prentice 1985). Despite the theoretical objection of
adding ‘fuel to the fire’, the administration of fibrino-
gen does not seem to be particularly dangerous.
Thrombotic thrombocytopenia purpura
Before the mechanisms involved in thrombotic throm-
bocytopenia purpura (TTP) were suspected, a plasma
factor was postulated to correct the syndrome charac-
terized by microangiopathic haemolysis and throm-
bocytopenia (Upshaw 1978). Relapses in chronic TTP
were reversed or prevented by infusions of small
volumes of FFP or cryoprecipitate-depleted FFP or by
plasma infusion combined with plasmapheresis (Byrnes
and Khurana 1977; Bukowski et al. 1981). The plasma
factor is not destroyed by the solvent detergent treat-
ment of FFP used to inactivate lipid-encapsulated
viruses (Moake et al. 1994). In the majority of cases,
the plasma factor relates to the activity of a metallo-
proteinase that cleaves unusually large multimers of
vWF that are associated with the TTP thrombi (Asada
et al. 1985; Tsai 1996) (see Chapter 17).
Cryoprecipitate-depleted fresh-frozen plasma
(cryosupernatant)
Cryosupernatant is plasma from which about one-half
of the fibrinogen, factor VIII and fibronectin has
been removed as cryoprecipitate. The product is also
depleted of the largest multimers of vWF, which
sediment in the cryoprecipitate fraction and which

may be partly responsible for platelet aggregation in
TTP (Moake 2004). In some circumstances cryosuper-
natant may be more effective than FFP in the treatment
of TTP. Seven patients with TTP who failed to respond
to intensive plasma exchange with whole plasma
responded to plasma exchange with cryosupernatant
(Byrnes et al. 1990).
Cryoprecipitate
When plasma is fast frozen and then thawed slowly at
4–6°C, the small amount of protein precipitated is
rich in fibrinogen, factor VIII, vWF and factor XIII.
After decanting almost all of the supernatant plasma,
the precipitated protein can be dissolved by warming
to yield a small volume of solution. The introduction
of cryoprecipitate revolutionized the treatment of
haemophilia by providing a highly effective, conveni-
ent, readily available source of factor VIII. Modern
treatment has moved away from cryoprecipitate to
pathogen-inactivated factor VIII concentrate and to
recombinant factor VIII. Cryoprecipitate is used now
as a source of factor VIII and vWF only if safer concen-
trates are not available.
Cryoprecipitate, containing approximately 200–
250 mg of fibrinogen in a volume of 10–15 ml, pre-
pared from a single donor, is used primarily as a source
of fibrinogen. The most common indication remains
as a replacement for fibrinogen consumed in DIC,
although it has been used as a topical haemostatic
agent as well (fibrin glue) (Reiss and Oz 1996).
Commercial fibrin sealants are safer, better standard-

ized and more effective, and avoid the potential risk of
CHAPTER 14
636
immunization to contaminant factor V that has been
reported when bovine thrombin is used to activate
topical cryoprecipitate (Rousou et al. 1989; Rapaport
et al. 1992; Atrah 1994) (see also Chapter 18).
Thawing by microwave is rapid and preserves fibrino-
gen concentration (Bass et al. 1985).
Cryoprecipitate also contains about 60% of the
vWF and 20–30% of the factor XIII of the original unit
of FFP, but the component in not often used as a source
of these proteins.
‘Contaminants’ in cryoprecipitate. Cryoprecipitates
contain about 30–50% of the original fibrinogen and
have about the same titre of anti-A and anti-B as that
of the original plasma unit (Rizza and Biggs 1969; Pool
1970). Because of the risk of haemolysis, neonates
should receive only ABO-compatible cryoprecipitate.
Plasma fractionation
The transfusion of whole plasma is unnecessary and
usually inefficient if recipients require only a single
protein, for example factor VIII. Plasma contains hun-
dreds of different proteins, many of which are obvious
candidates for replacement therapy, whereas others
are well characterized physicochemically, but of un-
known function. Commercial plasma fractionation uses
dilution, pasteurization and nanofiltration to remove
and inactivate most viruses, although no product can
be guaranteed ‘pathogen free’. The immunoglobulin

(Ig) fraction (predominantly IgG) separated from
whole plasma by alcohol fractionation was at first
considered virtually free of the risk of transmitting
viral hepatitis. However, HCV has been transmitted
by both IVIG and anti-D Ig (Bjoro et al. 1994; Meisel
et al. 1995; Power et al. 1995).
The most widely used method of fractionating
plasma is still the cold alcohol precipitation technique
described by Cohn and colleagues (1944) or modifica-
tions thereof (Kistler and Nitschmann 1962). Cohn
fractionation relies on changes in ethanol concentra-
tion and pH for bulk precipitation of different protein
fractions. An example of a fractionation scheme is
shown in Fig. 14.6. Ethanol is removed by lyophiliza-
tion or by ultrafiltration. Alcohol fractionation is now
combined with glycine precipitation or polyethylene
glycol, and with other separation methods such as
chromatography to isolate specific proteins, such as
coagulation factors and protease inhibitors.
Albumin
Albumin is available for clinical use either as human
albumin in saline containing 4%, 4.5%, 5%, 20% or
25% protein, of which not less than 95% is albumin,
or as plasma protein fraction (PPF), available only as
a 5% solution, of which at least 83% is albumin.
Compared with albumin, most preparations of PPF
contain larger amounts of contaminating proteins.
Hypotensive reactions attributed to pre-kallikrein
activator and acetate have been observed with PPF,
but not with albumin (Alving et al. 1978; Ng et al.

1981). For these reasons, most clinicians find little
reason to select PPF when an albumin solution is
indicated.
Albumin preparations are pasteurized by heat
treatment at 60°C for 10 h and filtered. When pre-
pared in this way, the fraction has proved free of
transfusion-transmitted agents such as hepatitis viruses
and HIV.
Although albumin contributes 75–80% of the col-
loid osmotic pressure of the plasma, subjects with a
genetically determined total absence of plasma albu-
min, in whom the colloid osmotic pressure of plasma is
between one-third and one-half of normal, may be
completely asymptomatic (Bearn 1978). Such subjects
show an increase in various plasma globulins and a
slight decrease in blood pressure, changes that are
regarded as compensatory. The indications for infu-
sions of albumin in hypovolaemic patients are discussed
in Chapter 2.
Recombinant albumin
Recombinant albumin has been synthesized in yeast,
in Saccharomyces cerevisiae or Pichia pastoris, and
appears to be similar to the plasma-derived protein
(Dodsworth et al. 1996). A 20% solution (Recombumin
20%, Aventis Behring) prepared as a pharmaceutical
excipient has been tested for safety in doses up to
65 mg in some 500 subjects. It is uncertain when, if
ever, recombinant albumin might be commercially
available as a product for transfusion.
Fibrinogen

The rate of disappearance of injected fibrinogen has
been studied by giving infusions to patients with the
very rare condition, hereditary afibrinogenaemia: in
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
637
two cases, one-half of the injected fibrinogen dis-
appeared during the first 24 h, presumably due to mix-
ing with the protein ‘pool’; thereafter the fibrinogen
disappeared with a T
1/2
of 4 days (Gitlin and Borges
1953).
In clinical practice, hypofibrinogenaemia is most
often encountered as one feature of the syndrome of
DIC; in this condition the transfusion of fibrinogen is
seldom indicated. Purified fibrinogen prepared by frac-
tionation of pooled plasma, unless virally inactivated,
carries a high risk of the transmission of viral diseases
such as hepatitis and no commercial fractionation con-
centrate is licensed.
Factor VIII (anti-haemophilic factor)
Factor VIII levels in haemophiliacs
Severely affected patients with haemophilia A have no
detectable factor VIII activity in their plasma and suf-
fer from repeated episodes of spontaneous bleeding,
particularly in the large joints and muscles. Patients
whose factor VIII activity is 1–5% of normal, about
10% of affected individuals, are defined as ‘moderate
haemophilia’ and have infrequent attacks of bleeding.
Those with levels exceeding 5%, some 30–40% of

patients, are mildly affected and seldom if ever suffer
CHAPTER 14
638
5000 or more litres of
pooled frozen plasma
Cryoprecipitation
Cryosupernatant
Factor VIII
Factor XIII
Factor XI
Factor IX Factor VII
Cryoprecipitate
Supernatant
DEAE-sepharoseHeparin-sepharose
Albumin
Immunoglobulin
Cold ethanol fractionation
Supernatant
Supernatant
Supernatant
Alcohol
DEAE-cellulose
Cold ethanol precipitation
Fig 14.6 The various blood products
(in squares) obtained by stepwise
fractionation of large pools of
fresh-frozen plasma using different
cryoprecipitation, ethanol
precipitation and adsorption
procedures.

spontaneous bleeding. Intracranial haemorrhage is the
most common cause of death from bleeding, is spontan-
eous in about 50% of cases and should always be con-
sidered in patients with haemophilia who complain of
headache.
Treatment with factor VIII
In severe haemophilia A, treatment with factor VIII
must be provided as soon as possible after bleeding has
occurred. The dose depends on the kind of haemor-
rhage. The aim of initial ‘episodic’ therapy in the case
of haemarthrosis or serious bruising is to raise the fac-
tor VIII level to 30–50% of normal; if a haematoma
has developed, the level should be raised to 50%, and
in case of gastrointestinal bleeding, to > 50%. The
level should be raised to 100% if there is spontaneous
intracerebral haemorrhage or head trauma (Furie et al.
1994). One unit of factor VIII is the amount of factor
VIII activity in 1 ml of normal plasma. For example, as
the plasma volume is about 50 ml/kg, 3500 units must
be given to a recipient of 70 kg to increase the level to
100%. Regardless of the source of factor VIII, the
plasma level reached after administration is only about
70% of the expected level and this must be taken into
account in calculating the dose required. The half-life
of factor VIII is 8–12 h. Thus if one-half of a dose is
given at 12-h intervals after the initial dose, the level is
kept relatively constant (Furie et al. 1994).
Prophylactic administration of factor VIII
The incidence of bleeding can be all but abolished and
even arthropathy can be prevented if factor VIII is

administered prophylactically from a very early age so
as to maintain the factor VIII concentration above 1%
of normal (Nilsson et al. 1994). Once joint damage
has occurred, it cannot be reversed by prophylactic
treatment (Manco-Johnson et al. 1994). The amount
of factor VIII needed for universal application of pro-
phylactic treatment is impractically large. Short-term
prophylaxis (3 months of bi-weekly infusions calcu-
lated to keep trough values at 1–3%) should be con-
sidered for patients with frequent haemorrhages or
with chronic synovitis, especially if active rehabilitation
is considered (Kasper et al. 1989). With preoperative
prophylaxis, surgical mortality from haemorrhage
approaches zero for most procedures (Kitchens 1986).
The initial infusion is routinely given several hours
prior to surgery and the factor VIII level confirmed
before induction of anaesthesia. The duration of post-
operative infusions depends on both the nature of the
procedure and the clinical course (Kasper et al. 1985).
Continuous postoperative factor infusion has become
an increasingly popular strategy to maintain constant
factor levels (> 50%) and consume less factor concen-
trate, although experience with this technique is still
limited.
Factor VIII levels of healthy donors: maximizing
collection potential
The factor VIII activity in group A donors is on
average 8% higher than in group O donors, and the
level in males is about 6% higher than in females
(Preston and Barr 1964). Strenuous exercise produces

an almost immediate increase in factor VIII levels, last-
ing for at least 6 h (Rizza 1961). These observations
carried greater importance when cryoprecipitate was
used as a source of factor VIII.
DDAVP (1-deamino-8-D arginine vasopressin, also
known as desmopressin acetate), a synthetic derivative
of vasopressin, injected intravenously, produces a rapid
release of vWF into the circulation. Although vWF is a
carrier protein for factor VIII, factor VIII and vWF
may not increase concurrently (Cattaneo et al. 1994;
Castaman et al. 1995). DDAVP is the treatment of
choice in patients with mild haemophilia with factor
VIII levels of > 10%, but should not be used in children
under 1 year of age because of the risk of hypona-
traemia (Weinstein et al. 1989; see also Chapter 18).
DDAVP is used primarily for therapeutic purposes.
However, if DDAVP (0.2 µg/kg) is injected into blood
donors 15 min before venepuncture, the yield of factor
VIII in fractions prepared from the resulting plasma
is increased two-fold (Nilsson et al. 1979; Mannucci
1986). DDAVP can also be administered intranasally
and is effective within 1 h with minimal side-effects
(Mikaelson et al. 1982).
Clotting factor concentrates: reducing the risk
and allaying the fear
Treatment with clotting concentrates was revolution-
ized by methods to inactivate viruses in plasma frac-
tions and by recombinant technology. The former
advance all but eliminated the risks of the major
transfusion-transmitted infections, whereas the latter

THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
639
added assurance that emerging agents would not
evade inactivation technology.
Choice of factor VIII concentrate
Factor VIII can be administered as cryoprecipitate, as
plasma-derived factor VIII concentrate or as recombi-
nant factor VIII. Factor VIII activity is well maintained
at –30°C to –40°C for cryoprecipitate or at 4°C
for lyophilized products. Commercial plasma-derived
factor VIII is now treated with at least two methods
for inactivating transfusion-transmitted viruses, and
no documented transmissions of the lipid-encapsulated
viruses HIV, HBV or HCV have been reported since
1985. Non-enveloped viruses such as hepatitis A and
parvovirus B19 may still be transmitted by plasma
fractions (Mannucci 1992; Santagostino et al. 1997).
Both plasma-derived and recombinant products are
labelled for factor VIII potency and all preparations
appear to be equally effective when assessed by post-
administration factor levels. Patient age, susceptibility
and product cost still largely determine the choice of
treatment.
Purified factor VIII concentrates
Intermediate-/high-purity concentrate. Factor purity
is commonly defined as specific activity (International
Units of clotting activity per milligram of protein).
Most fractionation centres use large pools of plasma
(5000–30 000 donations) to prepare products of inter-
mediate purity (< 50 units/mg) and high purity (> 50

units/mg). The primary procedure is cryoprecipitation,
but additional fractionation steps are undertaken to
give a higher potency, stability and solubility than are
obtained with the freeze-dried cryoprecipitate.
Ultrapure concentrate. Concentrates purified by
using affinity chromatography with monoclonal anti-
bodies against factor VIII have a specific activity of
factor VIIIC of > 3000 iu/mg protein. In a group of
patients treated with this product for more than
24 months, clinical efficacy, T
1/2
and recovery were
excellent (Brettler et al. 1989). One oft-stated advant-
age of purified high-potency concentrates is a less pro-
nounced effect on the immune system of the patients,
documented primarily in HIV-positive patients in
whom the CD4
+
cell count declines less rapidly after
treatment with high- than after intermediate-purity
concentrates (de Biasi et al. 1991; Hilgartner et al.
1993; Seremetis et al. 1993). No increase in AIDS-
associated infections or decrease in survival has been
documented (Goedert et al. 1994). No difference in
effect on the immune system in HIV-negative patients
has been established. Suspicion that ultra high-purity
concentrates (and recombinant concentrates) may
more easily induce factor VIII inhibitors, particularly
in children, have not been confirmed by prospective
studies (Bray 1994; Peerlinck 1994). The relationship

between mutation type and inhibitor development is
probably far more important. As many as 35% of
patients with ‘severe molecular defects’, intron 22
inversions, large deletions or stop mutations develop
an inhibitor, whereas few with small insertions or dele-
tions do so (Oldenburg et al. 2002).
Recombinant factor VIII. Recombinant clones encod-
ing the complete 2351-amino-acid sequence for human
factor VIII have been isolated and used to produce fac-
tor VIII in cultured mammalian cells. The recombinant
protein corrects the clotting time of plasma from
haemophiliacs and is virtually indistinguishable from
plasma-derived factor VIII (Wood et al. 1984).
Clinical trials have shown that in vivo recovery and
T
1/2
of recombinant factor VIII (r-factor VIII) are not
significantly different from those of plasma-derived
factor VIII. The r-factor VIII Recombinate (Baxter
Bioscience) and Koginate (Cutter Biological/Miles)
have now been used successfully in the treatment of
bleeding episodes and for prophylaxis. The observed
incidence of inhibitor formation is similar to studies of
previously untreated patients (PUPs) receiving plasma-
derived FVIII. Long-term trials demonstrate the safety
and efficacy of r-FVIII in chronic treatment of
haemophilia A (Lusher 1994; White et al. 1997).
Preparations of the early recombinant proteins used
plasma-derived reagents during manufacture. How-
ever the most recent generation is processed and for-

mulated without the addition of human or animal
plasma additives.
Animal factor VIII concentrates. Concentrates pre-
pared from bovine or porcine plasma have 100 times
more factor VIII activity per milligram of protein than
normal human plasma. The original preparations
were immunogenic and could as a rule be used effect-
ively for only 7–10 days, following which antibodies
against the animal protein developed. Polyelectrolyte-
CHAPTER 14
640
fractionated porcine factor VIII concentrate (PE porcine
VIII) appears to be considerably less antigenic and
contains negligible amounts of platelet-aggregating
factor (Kernoff et al. 1984).
Factor VIII inhibitors (antibodies)
Factor VIII inhibitors are found either as isoantibodies
in 15–35% of haemophilia A patients following treat-
ment with factor VIII-containing materials or, more
rarely, as autoantibodies in non-haemophiliacs. The
titre of the antibody measured in Bethesda units (Bu)
determines both risk and management. About one-half
of the antibodies in haemophiliacs are of low titre
and transient. Patients with factor VIII inhibitors are
relatively refractory to treatment with factor VIII and
must be given very high doses to secure a response.
Concentrates of factor VIII are effective if the inhibitor
titre is less than 20 Bu/ml; with higher titres, factor VIII
concentrates alone are ineffective. Haemophiliacs with
factor VIII inhibitors may show a rise in inhibitor titre

following the infusion of factor VIII; in such patients
factor VIII concentrates are not effective after 5–7 days
of treatment (Blatt 1982).
In treating major haemorrhage in haemophiliacs
with inhibitors, provided that the inhibitor titre does
not exceed 20 Bu/ml, human factor VIII concentrate
can be used initially. In average-sized adults (70 kg),
5000 units of factor VIII are given initially, followed
by 500–1000 units per hour. Thereafter, the dose is
adjusted according to the factor VIII level (Blatt 1982).
In patients with inhibitor titres exceeding 20 Bu/ml,
activated prothrombin complex concentrates appear
to be effective, but have an increased risk of throm-
boembolic complications (Hilgartner et al. 1990;
Tjonnfjord et al. 2004). These concentrates bypass
the need for factor VIII in a manner not thoroughly
understood. Recombinant activated factor VII (rFVIIa)
has been found effective in patients with antibodies
against factor VIII (see also Chapter 18) (Hedner and
Kisiel 1983; Abshire and Kenet 2004). Standard dos-
ing of rFVIIa (90 µg/kg) allows binding of FVIIa to the
surface of activated platelets and can directly activate
factor X in the absence of tissue factor. Experience with
bolus dosing suggests that higher dosing (> 200 µg/kg)
may be more efficacious in treating haemophilia patients.
In patients who require very large amounts of fac-
tor VIII, animal factor concentrates have been used
successfully (Kernoff et al. 1984). Four haemophiliacs
with factor VIII inhibitors received repeated infusions
of porcine VIII for periods up to 27 days with satisfac-

tory results. An antibody response was detected in
only one out of the four patients. Equally good results
were obtained in a later trial (Hay et al. 1990). The
newer treatments should render animal factor concen-
trates a historical curiosity.
In about 50% of patients with high-titre inhibitors
and up to 90% of those with titre < 5, immune toler-
ance can be induced by desensitization regimens that
involve daily factor VIII infusions (Ewing et al. 1988).
Treatment may be required for weeks or months and
infusions of factor VIII are sometimes combined with
short courses of immunosuppressive agents such as
corticosteroids, cyclophosphamide and IVIG (Kasper
et al. 1989; Mariani et al. 1994).
Treatment with IVIG is beneficial in some patients
with haemophilia A and inhibitors and particularly in
patients with factor VIII autoantibodies. The effect
may be due to anti-idiotype antibodies (Sultan et al.
1994; Schwartz et al. 1995).
Transfusion in patients with von Willebrand
disease
Von Willebrand disease is a common inherited auto-
somal dominant bleeding disorder characterized by
easy bruising, epistaxis, bleeding with dental proced-
ures and gastrointestinal haemorrhage. In the majority
of patients, bleeding results from decreased von
Willebrand factor (vWF), a protein carrier of factor
VIII that mediates platelet–platelet interaction and
platelet binding to vascular subendothelium (Ruggeri
and Ware 1993). The vWF gene is located on chromo-

some 12, and numerous polymorphisms and mutations
have been reported; the von Willebrand syndromes are
now classified by molecular, functional and clinical
criteria (Sadler 1998).
Appropriate treatment depends on the specific type
of vWD. Type 1 vWD, the most common form, usually
presents with mild or moderate bleeding. Laboratory
diagnosis shows low vWF antigen, activity (ristocetin
cofactor) and factor VIII levels, as well as a prolonged
bleeding time. Such patients usually respond well to
an infusion of DDAVP (0.3 µg/kg) with release of
sufficient vWF and factor VIII from tissue stores
within 30 min to elevate these levels several fold for
about 4 h (Scott and Montgomery 1993). Some of the
less common vWD variants do not respond well to
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
641
DDAVP (Ruggeri et al. 1980; Sutor 2000). For the
10–20% of patients with vWD who do not respond
to DDAVP and for patients who become refractory
(‘tachyphylactic’) with repeated treatment given over a
long period of time (Rodeghiero et al. 1992), replace-
ment therapy is available in several forms. Patients
who require treatment with vWF should be given high-
purity, solvent detergent-treated vWF concentrate
(Pasi et al. 1990; Burnouf-Radosevich and Burnouf
1992). If this is not available, factor VIII concentrate
which contains vWF should be used instead (Cohen
and Kernoff 1990). Cryoprecipitate, although rich in
vWF, has generally not been treated to reduce the risk

of viral transmission, and is therefore the least desir-
able of the available components. For planned surgery,
the products should be given a few hours before opera-
tion and, if factor VIII or ristocetin cofactor is used to
estimate effectiveness, the level should be measured
immediately before the operation is started; if the
level is not high enough (50–100%), more concentrate
should be given. Repeated infusions every 12 h may be
necessary for a week or more.
Treatment of factor IX deficiency (haemophilia B
or Christmas disease)
Patients with haemophilia B are treated with factor
IX concentrate. The administration of crude factor IX
concentrates derived from cryoprecipitate supernatant
(‘prothrombin complex concentrate’) was associated
with thrombotic complications (Kohler 1999), but
highly purified, virus-inactivated factor IX concen-
trates, based either on a combination of three conven-
tional chromatographic steps (Burnouf et al. 1989) or
on immune-affinity chromatography with monoclonal
antibodies against factor IX, are now available (Kim
et al. 1990; Tharakan et al. 1990). Recombinant factor
IX is also available. Some patients achieve only 80% of
the predicted plasma level and interpatient variability
is wide when this product is used, so that baseline
recovery measurements are important to ensure ade-
quate treatment. Treatment with highly purified or
recombinant product avoids the side-effects men-
tioned above (Kim et al. 1990). Each unit of factor IX
administered raises the plasma concentration by 1%.

The T
1/2
of plasma factor IX is 20 h. As in haemophilia
A, the dose needed is determined by the kind of bleed-
ing (see above) (Furie et al. 1994). If no factor IX
concentrate is available, PCC (see below) can be used
instead. Treatment with PCC may lead to thrombotic
disorders and DIC (Aronson and Menache 1987).
Treatment with factor IX concentrate may induce
inhibitor formation, although this is less frequent than
the formation of factor VIII inhibitors in patients with
haemophilia A (Knobel et al. 2002). The treatment of
patients with factor IX inhibitors is essentially the
same as that of patients with factor VIII inhibitors (see
below).
Prothrombin complex concentrates (concentrates
containing factors II, VII, IX and X)
Concentrates containing these vitamin K-dependent
factors were originally produced for the treatment of
inherited factor IX deficiency but are also used for
acquired deficiencies of factors II, VII, IX and X, for
example in patients with liver disease or warfarin over-
dose. As described above, PCCs have also been used
for treating minor bleeding episodes in haemophiliacs
with inhibitors. The concentrates should be adminis-
tered rapidly, and immediately after reconstitution.
PCCs are thrombogenic and their use remains con-
troversial except in the treatment of haemophilia B,
although, even here, purified factor IX concentrate has
largely replaced them (see above).

Use of some other coagulation factors
Factor VII
Fresh-frozen plasma can be used to treat factor VII
deficiency; however, the need for frequent infusions
and the risk of viral transmission have all but elimin-
ated its use for this indication. Plasma-derived factor
VII concentrate has been used successfully to treat
patients with hereditary factor VII deficiency (Dike
et al. 1980). Long-term prophylaxis in children has
also been successful (Cohen et al. 1995). Because of
its short half-life (3–4 h), frequent infusions are
necessary, especially for surgery, although levels of
15–25% of normal are sufficient for this indication.
Recombinant factor VIIa has been found to be effect-
ive in haemophiliacs with antibodies to factor VIII
and in some subjects with poorly controlled massive or
life-threatening haemorrhage (see above and Chapter
18) (Hedner et al. 1988; Schmidt et al. 1994). rFactor
VIIa is the treatment of choice when no pathogen-
reduced VII concentrate is available (Lusher et al. 1998).
CHAPTER 14
642
Factor XIII
Pasteurized factor XIII concentrates prepared from
either plasma or placenta are used to treat patients
with factor XIII deficiency, a condition that can be as
dangerous as haemophilia A or B (Smith 1990). Levels
as low as 5% are sufficient to control bleeding. Because
the half-life of factor XIII is measured in weeks,
infusions may be given every 14–21 days. In the USA,

where no factor XIII concentrate is licensed, FFP and
cryoprecipitate are the components of choice.
Protein C
Protein C is a serine protease zymogen, which is activ-
ated by thrombin. Activated protein C interferes with
the activated forms of factors V and VIII; it requires a
cofactor, protein S. Activated protein C also stimulates
fibrinolysis by neutralizing the inhibitor of tissue
plasminogen activator. Protein C deficiency, whether
hereditary or acquired as, for example, in severe liver
disease, leads to venous thrombosis. Protein C con-
centrates are now available: a vapour-treated protein
C concentrate that has been shown to be effective
for long-term therapy in an infant with severe
protein C deficiency (Dreyfus et al. 1995) and an
immunoaffinity-purified, activated protein C con-
centrate, virus inactivated by chemical treatment
(Orthner et al. 1995). Recombinant human activated
protein C has anti-inflammatory and profibrinolytic
properties in addition to its antithrombotic activity.
One recombinant formulation, drotrecogin alpha,
produced dose-dependent reductions in the levels of
markers of coagulation and inflammation in patients
with severe sepsis. In a randomized trial, treatment
with drotrecogin alfa activated significantly reduced
mortality in patients with severe sepsis, but seemed to
be associated with an increased risk of bleeding
(Bernard et al. 2001). The mechanism of the anti-
inflammatory effect is poorly understood, but may
involve inhibition of tumour necrosis factor produc-

tion by blockade of leucocyte adhesion or interference
with thrombin-induced inflammation.
C1 esterase inhibitor (C1 inh.)
Hereditary functional deficiency of C1 inh. is due to
either a deficiency or a dysfunction of the protein.
Acquired deficiencies of C1 inh. also occur. This pro-
tease inhibitor is involved in the regulation of several
proteolytic systems in plasma, including the comple-
ment system, the contact system of intrinsic coagula-
tion and kinin release and the fibrinolytic system
(Cugno et al. 1990). Functional deficiency of C1 inh.
permits production of vasoactive peptides that alter
vascular permeability and cause angioneurotic oedema,
a serious, potentially fatal syndrome characterized by
attacks of swelling of the subcutaneous tissues and
mucous membranes of the face, bowel and upper air-
way. Mortality rate in affected kindred approaches
30%. The pathogenesis of angioedema is not com-
pletely understood (Baldwin et al. 1991).
Pasteurized C1 inh. concentrates are now available
and acute attacks of angioedema respond within hours
of treatment (Brummelhuis 1980; Gadek et al. 1980;
Bork and Barnstedt 2001). Long-term prophylaxis with
C1 inh. concentrate in hereditary as well as acquired
C1 inh. deficiency has also been successful (Bork and
Witzke 1989; Waytes et al. 1996). Activation of the
complement and contact systems occurs in septic
shock, together with a decrease of plasma C1 inh. lev-
els. Preliminary results show that complement and
contact activation can be diminished by treatment

with high-dose C1 inh. concentrate (Hack et al. 1992).
Immunoglobulins
IgG
Following i.v. injection, IgG distributes between the
intravascular and extravascular compartments; equi-
librium is reached in about 5 days (Cohen and
Freeman 1960). The daily movement of IgG from the
intravascular to the extravascular compartment is
equivalent to about 25–30% of the plasma IgG and is
balanced by a similar transfer in the opposite direction.
When equilibrium has been attained, the plasma
level declines with a T
1/2
of 21 days. The fractional rate
of catabolism is largely independent of plasma IgG
concentration so that the total IgG turnover varies
directly with the plasma IgG level. The rate of IgG syn-
thesis is the primary factor determining the serum IgG
level (Schultze 1966). The catabolism of IgG has also
been studied by injecting HBs antibody in high titre,
with disappearance of antibody followed with a very
sensitive radio-immunoassay. The T
1/2
of these IgG
antibodies was calculated to be 19.7 days (Shibata
et al. 1983). Catabolism may be enhanced by fever,
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
643
burns and infection. For further information, includ-
ing the turnover rates of IgG subclasses, see Chapter 3.

Intravenous infusion of the ‘standard’ 16% Ig
preparations prepared for intramuscular injection may
produce severe reactions (see Chapter 15). This pre-
paration should not be administered intravenously.
Following i.m. injection of IgG, the protein passes
via the lymphatics into the bloodstream. Analysis of
plasma concentration curves is relatively complex as
the influx into the plasma from the site of injection is
offset by efflux from the plasma into the extravascular
space and also by catabolism. In a study in which
125
I-
labelled IgG was used, the average fraction of the dose
cleared per day from the site of i.m. injection, after
injecting 2 ml of solution into the deltoid muscle, was
estimated to be about 0.37. Plasma levels were almost
maximal at 2 days, and corresponded to about 40% of
the values that would have been attained immediately
after i.v. injection of the same doses (Smith et al. 1972;
see also Table 10.1). In a single case, surface counting
over the site of injection showed that approximately
45% of the total injected dose was cleared per day
(Jouvenceaux 1971). In two normal subjects following
the injection of 10 ml of 16% Ig into the gluteal region,
plasma levels corresponded to 32% of the injected
dose on day 5 in one case and 20% on day 7 in the
other (Morell et al. 1980). In retrospect, uptake may
have been poor in these two cases because the injec-
tions were made into fatty tissue rather than into
muscle. One survey indicated that when injections are

given into the gluteal region, few female patients and
fewer than 15% of male patients receive an i.m. injec-
tion (Cockshott et al. 1982).
Following subcutaneous injection of IgG into the
buttock, the rate of uptake is distinctly slower than
after i.m. injection and maximum plasma levels have
still not been attained 5 days after injection (Smith
et al. 1972).
Composition of IVIG preparations: not all are
created equal
Several Ig preparations containing fully functional
immunoglobulins are now available for i.v. use. These
are generally 5% or 10% solutions but concentrations
range from 3% to 12%. One product is prepared by
DEAE fractionation, followed by treatment of the
IgG-containing fraction at pH 4 with a low concentra-
tion of pepsin; the IgG molecule is not cleaved, but
high-molecular-weight aggregates responsible for ana-
phylaxis and some of the other adverse reactions are
dispersed (Jungi and Barandun 1985). If Ig for i.v. use
is to be stored in the liquid state, pH must be kept low
to maintain purity and stability. The low pH of the
product may be responsible for pain, erythema and
even phlebitis sometimes experienced at the injection
site. Freeze-dried preparations can be reconstituted
immediately before use at a pH of 6.6, thus avoiding
the above side-effects.
Stringent requirements for IVIG have been set by the
Committee for Proprietary and Medicinal Products
(). Human Ig preparations

contain very little IgM, but variable amounts of IgA.
Some two dozen commercial preparations are available
and vary in physicochemical characteristics including
concentration, volume, osmolality, sodium and sugar
content (Lemm 2002).
Changes in immunoglobulin preparations
on storage
Although most IgG antibodies show no obvious
change in potency over a period of several years in Ig
preparations kept at 4°C, the concentration of anti-D
diminished at about 8% a year over 2–4.5 years in
28 preparations tested (Hughes-Jones et al. 1978).
There was no evidence of appreciable breakdown
of IgG molecules in the preparations in the same
period. Low levels of immune complexes and variable
amounts of IgG dimer are found in commercial pre-
parations derived from large numbers of donor plasma
collections. The proportion of dimer increases over
months of storage and is enhanced by low temperature
(Tankersley et al. 1988).
Use of human immunoglobulin
Prophylaxis of infectious disease. Standard human Ig,
prepared for i.m. injection from unselected plasma, is
used in protection against hepatitis A, rubella, measles
and other disorders. The immunity conferred by an
injection of Ig is, of course, temporary and depends
upon the amount of antibody injected. Ordman and
co-workers (1944) found that a dose of Ig that would
protect children from measles for periods up to 14
days would not protect them for as long as 7–10 weeks

(Ordman et al. 1944). A relatively large dose (450 mg)
of Ig provides some protection for periods up to 6 weeks
CHAPTER 14
644
(Kekwick and Mackay 1954, p. 58). In the prophylaxis
of hepatitis A, a single dose of Ig (750 mg) may protect
for about 5 months (Pollock and Reid 1969).
Hyperimmune Igs prepared from selected donors
with high titres of the relevant antibodies are used in
the prophylaxis of hepatitis B, diphtheria, tetanus,
rubella, herpes zoster, rabies, measles and infection
with CMV, pseudomonas and other agents. Anti-D Ig
is used in the prevention of primary Rh D immuniza-
tion (see Chapter 10).
Anti-HBs Ig in very high doses can prevent reactiva-
tion of hepatitis in 65–80% of HbsAg-positive patients
receiving liver transplant grafts (Terrault and Vyas
2003). Monoclonal anti-HBs is available. Specific
anti-pertussis toxoid Ig in high titre has been shown in
a randomized trial to reduce significantly the number
of ‘whoops’ in 47 children with less than 14 days of
disease before therapy. Duration of whoops post treat-
ment was 8.7 days for patients with whooping cough
(Granstrom et al. 1991).
AIDS. A decreased incidence of bacterial infections
and sepsis and an improved survival rate have been
observed in infants with congenital AIDS receiving
monthly IVIG (Calvelli and Rubinstein 1986). The
beneficial effect of IVIG has been confirmed in children
with advanced AIDS receiving zidovudine. The bene-

fit, however, was only apparent in children who
were not receiving trimethoprim-sulphamethoxazole
as prophylaxis (Spector et al. 1994). In one study, pro-
phylactic administration of IVIG to premature infants
significantly reduced the risk of infections (Baker et al.
1992), but in another similar study, in which a differ-
ent preparation was used, IVIG had no effect (Fanaroff
et al. 1992). No recent study has evaluated IVIG
therapy in children with AIDS receiving highly active
antiretroviral agents (HAARTs), and the use of IVIG
should probably be restricted to children who develop
recurrent infections despite the administration of
HAARTs and prophylactic cotrimoxazole.
Replacement therapy for immunodeficiency syn-
dromes. Patients with either congenital or acquired
hypogammaglobulinaemia with IgG levels of less than
2 g/l are candidates for treatment. Intravenous infu-
sion is usually preferred, because large amounts of Ig
have to be given that are poorly tolerated when given
by repeated i.m. injection. Although subcutaneous
injection of Ig preparations for i.m. use has also been
satisfactory (Roord et al. 1982), the ease and ready
acceptance of intravenous administration has largely
replaced all other regimens. Patients with X-linked
agammaglobulinaemia, severe combined immuno-
deficiency (SCID), Wiskott–Aldrich syndrome among
others have benefited. Dosage has been determined
empirically, although prospective unblinded studies
confirm their effectiveness (Buckley and Schiff 1991).
The recommended intravenous dose is 300–400 mg/kg

body weight once per month. If the response is unsatis-
factory, the dose can be increased to attain a trough
level of 400–500 mg/dl.
Neonatal sepsis. Promising results in treating neo-
natal sepsis, particularly in premature infants, led to
the recommendation that infants weighing less than
1500 g at birth should be given 0.5 g of Ig daily for
6 days, as a routine measure (Sidiropoulos et al. 1981).
In a randomized study, 133 newborn infants were
divided into two groups based on whether the dura-
tion of gestation was shorter or longer than 34 weeks.
The infants were assigned to receive either 500 mg/kg
IVIG weekly for 4 weeks or no therapy. Septicaemia
and infection-related deaths were significantly less fre-
quent in the group of infants born before 34 weeks
who had received IVIG (Chiroco et al. 1987). In
another prospective randomized study, 753 premature
infants with early onset sepsis were randomly assigned
to receive a single injection of IVIG (500 mg/kg) or
albumin (5 mg/kg). At 7 days, none of the infants who
received IVIG but five of the control subjects had died
(P < 0.05). However, at 56 days, there was no differ-
ence in death rate. A single infusion may be insufficient
to reduce infection-related mortality for more than a
few weeks. No serious side-effects occurred in any of
the treated infants (Weisman et al. 1992).
Infection in adults. Selected Ig preparations contain-
ing antibodies in high titre may have a role in severe
viral and bacterial infection (Sawyer 2000). Trials
with anti-Pseudomonas Ig prepared from plasma from

vaccinated volunteers reduced the number of Pseudo-
monas infections and mortality in children and adults
with severe burns (Jones et al. 1980). The protective
effect of human IVIG preparations in bacterial infec-
tion has been shown clearly in mice (Imaizumi et al.
1985). However, results of other trials in patients with
severe burns or multiple injuries have been less con-
vincing (Berkman et al. 1990).
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
645
Chronic lymphocytic leukaemia
Hypogammaglobulinaemia is common in patients
with chronic lymphocytic leukaemia (CLL) and
response to immunization is impaired. The incidence
of infections was reduced by 50% in 42 patients with
CLL who received IVIG (400 mg/kg every 3 weeks)
compared with 42 patients receiving a placebo
(Cooperative Group for the Study of Immunoglobulin
in Chronic Lymphocytic Leukemia 1988). This result
has been confirmed (Griffiths et al. 1989). A dose of
250 mg per kilogram appears to be equally effective
(Gamm et al. 1994).
Multiple myeloma. In two prospective studies, a
substantial reduction in bacterial infections has been
observed in patients treated with IVIG (Schedel 1986;
Lee et al. 1994).
Kawasaki syndrome. Kawasaki syndrome, a child-
hood vasculitis thought by some to be associated with
infection by a retrovirus, responds to treatment with
high-dose IVIG (400 mg/kg per day for 3 days) com-

bined with aspirin (Nagashima et al. 1987). Fever
resolves in more than 85% of patients within 48 h and
coronary aneurysm formation is prevented (Burns
et al. 1998). Although the mechanism of action is
unknown, IVIG reduces nitric oxide production and
the expression of inducible nitric oxide synthase,
factors associated both with vascular smooth muscle
relaxation and with aneurysm formation (Fukunishi
et al. 2000).
Immune thrombocytopenic purpura. The chance
observation that IVIG raised the platelet count in
immunodeficient children with severe thrombocytope-
nia has led to its widespread use in the treatment of
immune thrombocytopenic purpura (ITP), first in chil-
dren and subsequently in adults (Imbach et al. 1981;
Fehr et al. 1982). The mechanism of action in this and
in other autoimmune disorders is unknown, but may
involve anti-idiotype antibodies, regulatory effects on
lymphocytes and macrophages or interference with the
effects of complement activation (Tankersley et al.
1988; Mollnes et al. 1997; Hansen and Balthasar
2004). Because children with acute thrombocytopenic
purpura are at risk for life-threatening haemorrhages,
treatment with IVIG has been advised when the
platelet count falls below 10 × 10
9
/l (Blanchette and
Turner 1986). In adults, treatment with IVIG has been
recommended for patients who are bleeding or as pro-
phylaxis for surgery, because infusions usually induce

a significant, albeit short-term rise in the platelet count
(Oral et al. 1984). The recommended dose is the origi-
nal empirical schedule of 400 mg/kg per day for 4 days
or 1 g per kilogram per day. The injection of relatively
small amounts of anti-Rh D Ig has also been successful
in inducing remissions in ITP. In D-positive adults,
750–4500 µg of anti-D was given in one series (Salama
et al. 1986); in another, all of 13 D-positive patients
given 2500 µg responded, and a single D-negative
patient failed to respond (Boughton et al. 1988).
Success has also been achieved by giving a single dose
of 4 ml of D-positive red cells coated in vitro with
100 µg of anti-D (Ambriz et al. 1987). These observa-
tions suggest that anti-D produces its beneficial effects
by causing D-positive red cells to bind to, and thus
block, Fc receptors on macrophages. Remission has
been reported in one D-negative pregnant woman who
had failed to respond to steroids and who was given
120 µg of anti-D intravenously (Moise et al. 1990).
The effective agent in anti-D Ig may be HMW IgG
polymers rather than anti-D. Preparations of anti-D Ig
contain a substantially higher proportion of HMW
IgG polymers than non-specific Ig preparations and
these polymers are more effective in blocking Fc
receptors (Boughton et al. 1990). In a randomized
study, the effect of intravenous anti-D on the platelet
count in childhood acute ITP has been found to be
inferior to that of IVIG (McMillan et al. 1994);
however, one treatment occasionally proves effective
after the other has failed. IVIG and anti-D may work

through different mechanisms (Cooper et al. 2004).
Other autoimmune diseases
IVIG has been used successfully in treating patients
with autoimmune neutropenia (Pollack et al. 1982;
Bussel and Lalezari 1983). For the effect of IVIG in
autoimmune haemolytic anaemia, see Chapter 7.
IVIG has also been used in treating patients with
other autoimmune diseases, among them myasthenia
gravis, the Guillain–Barré syndrome, multiple sclerosis
and chronic demyelinating polyneuropathy (Knezevic-
Maramica and Kruskall 2003). In patients with factor
VIII or factor IX inhibitors, the antibody titre decreases
following the administration of IVIG (Nilsson and
Sundqvist 1984; Sultan et al. 1994).
CHAPTER 14
646
Alloimmune diseases. High-dose IVIG has been used
in Rh D haemolytic disease (see Chapter 12), in neona-
tal alloimmune thrombocytopenia (see Chapter 13)
and in post-transfusion purpura (see Chapter 15).
Bone marrow transplantation
IVIG has been shown to reduce the incidence of septi-
caemia, interstitial pneumonia, fatal CMV disease,
acute GvHD and transplant-related mortality in adult
recipients of related marrow transplants (Siadak et al.
1994). The mechanism responsible for this effect is not
known (Gale and Winston 1991; Sullivan et al. 1991).
A meta-analysis of 12 randomized, controlled trials of
prophylaxis in bone marrow transplantation revealed
a significant reduction in fatal CMV infection; CMV-

interstitial pneumonia, non-CMV pneumonia and
transplant-related mortality (Bass et al. 1993). Con-
tinued administration after day 90 does not appear
to reduce late-occurring infections or chronic GvHD
(Sullivan et al. 1996).
Adverse reactions
Adverse reactions occur in as many as 15% of admin-
istrations (Boshkov and Kelton 1989). A commonly
recognized complex of symptoms including flushing,
headache, nausea and fever has been attributed to the
presence of IgG aggregates and complement fixation
by IgG dimers. The syndrome was first reported with
i.m. administration, but clearly occurs with IVIG,
especially when infusion is rapid (Nydegger and
Sturzenegger 1999). An anaphylaxis-like syndrome
that includes chills, arthralgias, flank pain, urticaria
and circulatory collapse was reported with the first
IVIG preparations, but occurs less commonly with the
low pH pepsin treatment described above (Barandun
and Isliker 1986; Tankersley 1994).
Passive antibody transfer is the intended con-
sequence of treatment with immunoglobulin pre-
parations; however, some passive antibodies cause
diagnostic uncertainty, whereas others such as red cell
alloantibodies may cause haemolysis. Persistent meas-
urable titres of anti-HBs and anti-CMV in particular
may persist for months, although the titre will fall
over time and evidence of virus by other assays will be
lacking (Lichtiger and Rogge 1991). Antibodies to
numerous red cell antigens have been reported in up

to one-half of commercial Ig preparations in the past,
although this is a less common fining in the newer
preparations (Niosi et al. 1971; Nydegger and
Sturzenegger 1999). Both passively acquired DAT and
haemolysis occur after infusion of passive antibody
(Copelan et al. 1986; Moscow et al. 1987). In a series
of 47 patients who received high-dose IVIG as prophy-
laxis for cytomegalovirus infection following bone
marrow transplantation, almost one-half were found
to have an acquired DAT, and a quarter a positive
indirect antiglobulin test (IAT) caused by passive
anti-A, -B, -D and -K (Robertson et al. 1987). Most
findings appeared within 1 week of initiation of IVIG
therapy. Transient neutropenia, presumably as a result
of passive neutrophil antibody, has been reported as
well (Tam et al. 1996).
Thromboembolism, including venous thrombosis,
pulmonary embolism, myocardial infarction, stroke
and hepatic veno-occlusive disease, has been associated
with IVIG infusion (Go and Call 2000). The mechan-
ism is unclear; however, changes in viscosity, proco-
agulant contaminants and platelet activation have all
been implicated. Manufacturers and regulatory agen-
cies have called specific attention to this complication
(Dalakas and Clark 2003).
Renal dysfunction, including acute renal failure and
17 fatalities, has been reported to the FDA (Epstein
and Zoon 2000). The pathological appearance of the
proximal renal tubules of the kidney has suggested
‘sucrose nephropathy’, so that sucrose added to some

IVIG preparations has been implicated as the cause
(Cayco et al. 1997). Patients with underlying renal
disease, especially the elderly and those with diabetes
mellitus sepsis, monoclonal gammopathies and volume
depletion seem particularly susceptible to renal failure.
Aseptic meningitis has been reported as a dose-
related complication of IVIG infusion, particularly in
patients with pre-existing migraine (Sekul et al. 1994).
Case reports of recurrent migraine and seizures have
been published and may be causally related, but the
most intriguing is that of a patient who suffered recur-
rent episodes of hypothermia with IVIG infusion
(Duhem et al. 1996).
Transmission of HCV through IVIG has been
referred to above (Bresee et al. 1996).
Novel intravenous immunoglobulins
By hybridoma technology, genetic engineering and
chemical methods, novel specific monoclonal antibody
THE TRANSFUSION OF PLATELETS, LEUCOCYTES, HAEMATOPOIETIC CELLS AND PLASMA COMPONENTS
647
preparations now constitute a significant proportion
of biopharmaceutical products in development. Several
chimeric and humanized monoclonal antibodies are
now licenced therapeutics (Roque et al. 2004).
Antithrombin
Hereditary AT deficiency occurs in at least two forms:
in one, the level of antithrombin is low (about 50% of
normal), and in the second, antithrombin is function-
ally deficient. In both cases the deficiency of this
natural anticoagulant is associated with a high risk of

venous thrombosis. The first event often presents in
adolescence or young adulthood (Demers et al. 1992).
Antithrombin inactivates five of the activated coagula-
tion factors. This function of antithrombin at several
levels of the coagulation pathway probably explains
why an apparently modest decrease in antithrombin
activity, as in patients with familial low antithrombin
levels, leads to a thrombotic tendency (Abilgaard 1984).
Heat-treated antithrombin concentrates are avail-
able with an initial 50% disappearance time of 22 h
and a biological half-life of 3.8 days, and are indicated
for the prevention or treatment of thromboembolic
disorders in patients with hereditary antithrombin
deficiency (Lechner et al. 1983; Menache et al. 1990;
Lebing et al. 1994). A recombinant human antithrom-
bin has been used in congenitally deficient patients
who require surgery; the optimal dosing regimen
remains to be defined (Konkle et al. 2003).
Acquired AT deficiency has several causes. Admin-
istration of antithrombin may benefit patients with
cirrhosis who are to undergo surgery and patients in
hepatic coma or pre-coma (Lechner et al. 1983).
In DIC, in which antithrombin levels are often low,
treatment with antithrombin concentrate may help,
particularly when treatment is started early and enough
concentrate is given to maintain a plasma level of at
least 100% of normal (Lammle et al. 1984; Gabriel
1994; Schwartz 1994).
Fibronectin
Plasma fibronectin, an opsonic glycoprotein that may

play a role in wound healing, infection and vascular
integrity, enjoyed a short but enthusiastic vogue as a
therapeutic agent when administered in the form of
cryoprecipitate (Saba et al. 1978; Saba and Jaffe 1980).
Treatment of trauma and burn patients deficient in
plasma fibronectin with cryoprecipitate purportedly
resulted in clinical improvement (Saba et al. 1986).
However, a controlled trial of fibronectin found no
benefit for patients with severe abdominal infections
(Lundsgaard-Hansen et al. 1985). Similarly, patients
with septic shock or severe injury showed no evidence
of improvement after treatment with fibronectin
(Rubli et al. 1983; Hesselvik et al. 1987; Mansberger
et al. 1989).
α
1
-Antitrypsin
α
1
-Antitrypsin (α
1
-AT) is a major serine endopeptidase
inhibitor in human plasma, which inhibits neutrophil
elastase, an enzyme involved in the proteolysis of
connective tissue, especially in the lung. Hereditary
deficiency of α
1
-AT may lead to progressive emphy-
sema. Clinical trials have suggested that replacement
therapy in deficient patients may restore the con-

centration of α
1
-AT in plasma and thereby limit the
development of emphysema (Gadek et al. 1981).
Concentrates of α
1
-AT, which can be treated at 60°C
for 10 h, are available. Weekly injections of 4 g for
6 months were given to 21 patients homozygous for
the deficiency allele P1Z (Wewers et al. 1987). Peak
levels in plasma were above the normal upper range.
After a rapid decline during the first 2 days after infu-
sion, corresponding to redistribution of α
1
-AT into the
intravascular space, there was a slower rate of decline
consistent with the normal 4- to 5-day half-life of
plasma α
1
-AT. The lowest levels before the next injec-
tion were always above the threshold level. Diffusion
of the infused material across the alveolus and a
significant increase in elastase activity in epithelial lin-
ing fluid could be demonstrated. Similar results were
obtained in another study (Konietzko et al. 1988).
There are, however, still unanswered questions with
regard to replacement therapy with α
1
-AT. Whether
such therapy actually prevents the development or the

further progress of emphysema remains unknown and
would require a large randomized trial (Dirksen et al.
1999). Neither has the question as to which deficient
patients should be treated been answered. Considering
the number of deficient patients (estimated at 70 000
in the USA), long-term demand cannot be met by α
1
-
AT produced from plasma. Recombinant α
1
-AT would
surely be needed and such products are in develop-
ment. The American Thoracic Society and European
Respiratory Society have reviewed this subject (2003).
CHAPTER 14
648
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