HEMOLYTIC TRANSFUSION REACTIONS
I V . DLFFERENTIAL DLAGNOSIS: "DANGEROUS UNIVERSAL DONOR" OR
INTRAGROUP INCOMPATIBILITY?
ALEXANDER S. WIENER AND WILLIAM C. MOLONEY
From the Transfusion Division of the Jewish Hospital of Brooklyn and the St. Elizabeth's
Hospital of Brighton, Mass.
Three types of intergroup incompatibility can be distinguished : 1) The
patient's sérum agglutinâtes (or hemolyzes) the donor's cells, but the donor's
sérum does not react with the patient's cells. 2) The donor's sérum contains
isoantibodies for the patient's cells, but the patient's sérum is compatible with
the donor's cells. 3) The patient's sérum and donor's cells, and the donor's
sérum and the patient's cells are mutually incompatible. With regard to the
first two types of incompatibility, Ottenberg 1 was the first to point out that
while donors whose cells are incompatible with the patient's sérum must be
rejected, blood merely containing incompatible isoagglutinins can usually be
transfused with benefit to the patient. This led to the use of group 0 blood
("universal donor" blood) for patients of other groups, and of blood of groups
0, A and B for group AB patients ("universal récipients").
For a long time, a controversy existed with regard to the safety of universal
donor blood. Some workers insisted that only blood of the homologous group
be used for transfusions, while others claimed that universal donors were as
safe as donors belonging to the patient's own group. On the basis of "in vitro"
tests, Levine and Mabee2, and Freeman and Whitehouse 3 adopted the view that
while in most cases group 0 donors could probably be used without danger for
patients not belonging to group O, blood from group O donors with exceptionally
potent isoagglutinins might cause hemolytic reactions. Ordinarily, the dilution
of the donor's sérum in the patient's blood should be sufficient to protect the
patient's cells from the action of incompatible isoantibodies, but when the
latter are very potent, this protective mechanism might prove inadéquate,
particularly in patients with severe anémias.
The view of Levine and Mabee is supported by sporadic reports of hemolytic
reactions that hâve been attributed to the use of "dangerous universal"
donors 4,5 - 6 ' 7. On the other hand, universal donors were used almost exclusively for transfusions in Belgium and France before the war. Such donors
were preferred because they were more convenient to obtain and time-saving,
the grouping and matching tests being omitted. According the Riddell8, the
Transfusion Sanguine D'Urgence in France supplied universal donors for 6,000
transfusions a year, yet claimed there were no hemolytic reactions ascribable to
this source. Rosenthal and Vogel9 analyzed 819 transfusions where universal
donors had been used for patients not belonging to group O and reported that
such transfusions were not followed by any greater number of reactions than
transfusions of homologous blood. Such reports hâve led some authors to question the view that the indiscriminate use of universal donors can ever cause
74
HEMOLYTIC TRANSFUSION REACTIONS
75
severe or fatal hemolytic reactions. A further complication arose when it was
shown that in patient receiving repeated transfusions or those who hâve been
pregnant, hemolytic reactions can occur based on intragroup incompatibility,
in particular with regard to the Rh factor11. Obviously, this mechanism could
also operate when universal donors are used, so that hemolytic reactions following
such transfusions are not necessarily due to incompatibility between donor's
sérum and patient's cells. Inasmuch as this possibility has not been taken into
account except in the single récent report by Klendshoj and McNeil7, one may
well question the validity of the conclusions in some of the older reports.
Accordingly, the purpose of this paper is to présent the differential diagnosis
of hemolytic reactions caused by intragroup incompatibility and those due to
dangerous universal donors. That the two sorts of incompatibility be distinguished is obviously important in the event it becomes essential to transfuse
the patient again.
DIFFERENTIAL DIAGNOSIS
As was first shown by Ashby10, following a transfusion of group 0 blood to a
patient not belonging to group 0, the donor's blood cells can, as a rule, be demonstrated in the patient's circulation for periods up to four months. For example,
the blood of a group B patient who has received a transfusion of group 0 blood
should give only partial agglutination in anti-B serums (method of differential
agglutination). However, in hemolytic reactions caused by intragroup incompatibility (between the patient's sérum and donor's cells) the donor's cells
are rapidly eliminated from the patient's circulation so that the appearance of
the agglutination reaction does not change after the transfusion11'12. On the
other hand, in hemolytic reactions caused by the use of a "dangerous universal
donor," the incompatibility involves the donor's sérum and the patient's cells
so that the donor's cells should survive normally. Accordingly, by the method
of differential agglutination the two types of hemolytic transfusion reaction can
readily be distinguished. If in the test the patient's cells show complète agglutination, then one is dealing with a case of intragroup incompatibility; this
conclusion will be further supported if it is found that the patient is Rh-negative
and the donor Rh-positive, especially if the patient's sérum contains anti-Rh
isoantibodies. On the other hand, if the test shows partial agglutination, then
the hemolytic reaction must hâve resulted from the hemolysis of the patient's
own cells by the donor's sérum. This conclusion will be further supported if it
is shown that the donor's sérum contains exceptionally potent isoagglutinins,
and clinically the possibility of intra-group incompatibility can practically be
excluded in patients who hâve never received a previous transfusion or been
pregnant.
The following case illustrâtes the differential diagnostic points discussed
above.
CASE REPORT
The case will be described in the order in which the facts became known to us, since in
this way it is most interesting and instructive.
The senior author's first contact with the case occurred when he received a sample of the
76
ALEXANDEE S. WIENER AND WILLIAM C. MOLONEY
patient's blood for examination together with a brief note containing the following data.
The patient, group A, Rh-negative, was transfused with Rh-positive blood. The patient
had had a toxemia of pregnancy and a stillbirth. The intern had advised the use of an
Rh-negative professional donor but the family insisted that the blood be taken from a
friend. Since the direct matching test at 38°C. by the centrifuge technic showed no agglutination the family donor was used. When 400 ce. of blood from this donor had been
transfused, the patient had a violent chill with a rise in température to 105°F., and at the
same time complained of intractable lumbar pain. Within thirty-six hours after the transfusion the patient became noticeably jaundiced, she developed anuria and the blood NPN
rose to 160 mg./lOO ce.
With the above history it seemed almost certain that the patient's hemolytic
reaction was due to isoimmunization in pregnancy, the Rh factor being the antigen involved. The patient's blood was retested and its classification as group A
confirmed. However, only partial agglutination was obtained with ail the
high-titered anti-A serums tried, about one-tenth of the cells remaining unagglutinated, even though tests with absorbed B serums showed that the patient
belonged to subgroup Ai. This indicated that the patient's circulation contained a mixture of group A and group 0 blood, which came as a surprise since
no mention had been made in the case history that a universal donor had been
used. On the basis of thèse results, hemolysis of the donor's cells due to intragroup incompatibility appeared to be ruled out, and the possibility remained that
the reaction had been caused instead by hemolysis of the patient's own cells by
the donor's sérum. The M-N tests threw no light on the question, because the
patient and donor both belonged to the same type (MN), but the results of the
Rh tests were interesting. With the aid of a potent human anti-Rh sérum, the
patient's blood was found to contain a mixture of about 9 parts of Rh-negative
and 1 part of Rh-positive blood. This confirmed the report that the patient
was Rh-negative and the donor Rh positive, and at the same time demonstrated
again that the donor's blood had nevertheless not been hemolyzed.
Based on thèse results it was predicted that the donor who had been used belonged to group 0 (type MN) and that his sérum contained isoagglutinins of
high titer for group A blood. A sample of the donor's blood was then procured
and the expectations were found to be correct in every respect. The results of
the titration tests on the donor's sérum are given in table 1.
The exceptional nature of the donor's sérum is emphasized when the titers of
the agglutinins in his sérum are compared with those of a random séries of normal
individuals. In table 2 are summarized the results of such a séries of titrations
on blood donors who were tested in order to ascertain whether or not their serums
were of high enough titer for the production of typing sérum. Those with a
titer higher than 60 were considered to hâve désirable serums for this purpose.
I t will be seen that among more than 100 donors there were only 5 per cent with
titers higher than 60, and none in this particular séries with agglutinin titers as
high as that of the donor used for transfusing our patient. It is clear, therefore,
that donors with titers higher than 100 are rare, and this probably explains why
as a rule transfusions of universal donor blood to patients not belonging to
group O does not produce noticeable harmful effects.
77
HEMOLYTIC TRANSFUSION REACTIONS
Many investigators hâve reported higher isoagglutinins titers for normal individuals
than those given in table 2. Therefore, we should like to emphasize that the titer reported
for a given sérum will vary with the technic used. For example, if instead of mixing one
drop each of sérum (or sérum dilution) and 2 per cent cell suspension, one mixes 0.5 ce. of
TABLE 1
TlTRATION OP DONOK'S SERUM
SERUM DILUTION
TESTED
WITH
CELLS Undil. 1:2
DONOR'S
Untreated
Ai
A2
B
1:4
1:8
1:16
1:32
TITER
1:64 1:128 1:256 1:512 1:1024
hem. hem. + ± + + ± + + ± + + ± + + ± + + + ± +
+± + + + + ± + + ± + + ± + + ± + + + + +± +
+
hem. + • + ± + + + + + ±
Mixed with A! + + + + + +
Ï Ï volume A2 tr.
of Witeb- B + + ± + + + +
sky's
group
substance
+ +
+ ±
+
+ +
+±
+
Mixed with Ai
TV volume A2
of Witeb- B
sky's
group
substance
tr.
+±
+±
+
— 512
512
64
32
<1
32
tr.
-
4
0
8
+ +
+
±
-
For the titrations, fresh washed 2 per cent (in terras of blood sédiment) suspensions were
used. The tests were set up on Boerner well slides by mixing one drop (about .05 ce.)
each of blood suspension, sérum or sérum dilution, and saline, and the readings were taken
after the mixtures had been shaken for 10 minutes on the Boerner shaking machine.
TABLE 2
TITERS OF ISOAGGLUTININS IN A RANDOM SÉRIES OF NORMAL INDIVIDUALS
SERUM TITER
1
2
4
8
16
32
64
128
Totals..
NUMBER OF GROUP 0 AND GROUP BNUMBER OF GROUP 0 AND GROUP A
SERUMS WITH
SERUMS WITH
INDICATED TITER FOR M CELLS
INDICATED TITER FOR B CELLS
5
11
11
12
16
12
4
1
4
13
28
39
25
22
4
1
72
136
In the présent séries, the average titer of the anti-A agglutinins is not higher than that
of the anti-B agglutinins, as usually reported, beeause blood of subgroup A2 was selected
for the tests. When the group A test cells are chosen at random, they are apt to belong to
the more sensitive subgroup Ai, beeause Ai is about 4 times as fréquent as A2.
78
ALEXANDEK S. WIENER AND WILLIAM C. MOLONEY
sérum (or sérum dilution) with a drop of 1 per cent cell suspension, ail the titer values
become 8 times as high (Candela13). While almost any titer value can be obtained for a
given sérum by varying the technic, the relative titers of two différent serums should remain
constant unless technical errors occur. When a single pipet is used for preparing the séries
of dilutions, if sérum is carried over because of inadéquate rinsing, erroneous high titers
may be obtained. Obviously, such mistakes can readily be avoided by changing pipets
between dilutions or instituting a rigorous technic of rinsing. If mouth pipets are used,
care must be taken to prevent any saliva slipping in, which may neutralize the isoagglutinins and thus give rise to false low titers.
When titrating serums which are to be used as testing serums, a practical considération
is that the method of titration should be comparable to the technic used in the actual tests.
If the method of titration is more sensitive than the tests themselves, a poor sérum might
be passed. Moreover, the highest dilution of sérum used in the tests giving a distinct
reaction should be taken as the titer rather than the dilution of the sérum after it is mixed
with the test cell suspension, because the former measures the number of units actually
available, the dilution of the sérum in the tests being unavoidable.
Following the hemolytic transfusion reaction, our patient was treated
heroically, but after a rather protracted course extending over a period of about
a month, she finally died. An attempt was made during the patient's life to obtain a sample of saliva in order to détermine whether or not she was a secretor,
but this was not possible because of the patient's dehydrated and comatose condition. Postmortem, a sample of gastric juice was obtained, and then it was
found that the patient was a secretor.* I t is évident, therefore, that the présence
of group-substances in the patient's sécrétions was not sufficient to protect her
from the harmful action of the isoantibodies in the donor's sérum, and this is
not surprising considering the unusual potency of this donor's isoantibodies.
A sample of the patient's blood procured a week after the transfusion reaction
occurred was even more deeply icteric than the sample obtained the day after
the transfusion, indicating that hemolysis was still going on. It is difficult to
décide whether the deepening of the icterus was due to the continued action of
the donor's isoantibodies on the patient's cells, which is entirely pbssible considering their extremely high titer, or whether the intercurrent infection which
the patient later developed was responsible, or the sulfonamide therapy, Itis
of interest to note that the sample taken a week following the transfusion showed
that about half the donor's blood cells had already been eliminated from the
patient's circulation; evidently, therefore, even though there was no outspoken
incompatibility between the patient's sérum and the donor's cells sufficient to
cause a reaction, there was a latent incompatibility of the donor's cells for the
patient. The patient was transfused again, but in the subséquent transfusions
only group A, Rh-negative blood was used. There was no reaction of any sort
following thèse transfusions.
It occurred to us that the reaction might hâve been prevented if the purified
group substance of Witebsky had been added to the donor's blood 15 ' l6 ' n . Accordingly, an attempt was made to answer this question by titrating the donor's
sérum after it was mixed with ^ its volume of the group-substance (Witebsky
* For technique see Wiener.14
HEMOLYTIC TRANSFUSION REACTIONS
79
recommends the use of 10 ce. for 500 ce. of whole blood or 250 ce. of plasma).
It will be seen that while the addition of the group substance did not completely
neutralize the isoagglutinins, it brought their titers down to a safer level. The
addition of xV volume of group substance naturally had a more pronounced effect
and practically eliminated the isoagglutinins.
COMMENT
The présent case is probably the first completely proven instance of a fatal
hemolytic reaction caused by the use of a universal donor. It was demonstrated
that the hemolysis was caused by the présence in the donor's sérum of exceptionally potent isoantibodies. The fact that serums with isoagglutinins of such
high titer are very rare explains why the indiscriminate use of universal donors
hardly ever gives rise to dangerous reactions. At any rate, the indiscriminate
use of universal donors for patients of other groups is justified only in emergencies; but if group substances are added to the blood in order to neutralize the
isoantibodies, the transfusion of such blood would be safe and bénéficiai enough
even for routine use.
If the group substances are not available, universal donor blood with potent
isoagglutinins can probably be transfused safely by injecting it slowly and well
diluted18. In this connection the experiments of Aubert, Boorman, Dodd and
Loutit19 are of interest. Thèse investigators transfused 12 persons not belonging
to group 0 with group 0 sérum (volume 30 to 500 ce.) containing high-titered
isoagglutinins. Following thèse transfusions, there was definite évidence of
destruction of the recipient's blood cells (increased urobilin excrétion, fallin
red cell count and hemoglobin). A few récipients had signs and symptoms of
acute hemolysis (pain in the back, hemoglobinemia and hemoglobinuria). Fortunately, in no case were there any serious sequelae, possibly because ail the
récipients were fully alkalinized, and conscious, and the transfusions were discontinued as soon as untoward symptoms of any sort appeared. On the other
hand, transfusions of group O sérum of low isoantibody content produced no
untoward effects of any sort.
In our case, the most obvious and apparently most logical diagnosis was not
the correct one. From the clinical data it seemed fairly certain that we were
dealing with an example of isoimmunization in pregnancy and that the hemolytic
reaction was caused by the Rh factor. It developed instead that the reaction
was due to the use of a dangerous universal donor.
SUMMARY
Methods of differentiating hemolytic reactions due to isoimmunization with
intragroup incompatibility from those caused by universal donor blood are
described.
An illustrative case is presented in which the clinical data indicated that the
hemolytic reaction was due to intragroup incompatibility resulting from isoimmunization in pregnancy, but which in fact turned out to hâve been caused
by the use of a universal donor with exceedingly high-titered isoantibodies.
80
ALEXANDER S. WIENER AND WILLIAM C. MOLONEY
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