A Group B Analogue of Subgroup A3
ALEXANDER S. WIENER, M.D.,
AND ALBERT F. CIOFFI,
M.D.
Serological Laboratory of the Office of the Chief Medical Examiner of New York City,
and Department of Forensic Medicine, New York University School of Medicine
and Eastern Biologicals, Jersey City, New Jersey
ABSTRACT
Wiener, Alexander S., and Cioffi, Albert F.: A group B analogue of subgroup
A3. Am. J. Clin. Pathol. 58: 693-697, 1972. A rare agglutinogen characterized
by only partial agglutination of the erythrocytes with anti-B (or group O)
sera, failure to agglutinate with anti-A, and strong agglutination with anti-H
(Ulex europeus) has been found in a male Negro donor and his mother.
Their sera contained anti-A but not anti-B, and the anti-A titers could be
increased by injections of blood group substance A. The donor and his
mother were both nonsecretors. As this very rare subgroup appears to be the
analogue of subgroup A3, it is designated B 3 and its corresponding allelic
gene as Bs.
A 3 erythrocytes, like other group
A cells, have the property of being agglutinated by anti-A but not by anti-B reagents.* However, only a fraction of the
erythrocytes take part in the agglutination
reaction, giving rise to a characteristic picture of clumps of erythrocytes on a background of unagglutinated erythrocytes.
Friedenreich, 2 who first described the subgroup A3, reported its incidence to be
about 1 in 1,000 in Denmark, and described families which indicated that the
subgroup A 3 was due to a special allelic
gene A3. Wiener and Silverman 5 described
the agglutinogen A 3 in a Negro woman and
her son, and found that the sera of mother
and child contained, in addition to anti-B,
SUBGROUP
Received November 24, 1971; received revised
manuscript January 17, 1972; accepted for publication February 2, 1972.
Supported in part by U. S. Public Health Service
N.I.H. grant GM-09237.
• T o avoid ambiguity, symbols for blood factors
and their corresponding antibodies are printed in
boldface type, symbols for genes and genotypes are
printed in italics, while symbols for agglutinogens,
phenotypes, and blood group systems are printed in
regular type.
693
a weak anti-Ai isoagglutinin which was
lacking in the cases described by Friedenreich.2 An extensive study of A 3 and other
variants of the agglutinogen A was presented in a thesis by Gammelgaard. 3
Rare variants also have been described
of the agglutinogen B. 4 - 7 The purpose of
the present article is to describe one such
very rare variant which behaves like an
analogue of A3.
The proband was a male Negro blood
donor, 34 years old, whose blood had originally been classified as group O. His blood
was drawn and stored in the usual manner
and later released for hospital use for transfusion. At the hospital, in crossmatching
tests with serum from a group O patient, an
incompatibility was detected by the technician, even though retests of the donor's
blood appeared to confirm its classification
as group O. The unit of blood therefore
was returned for further study.
When the blood grouping tests were repeated, the donor's erythrocytes again gave
no discernible reaction with anti-A or anti-B
694
WIENER AND CIOFFI
serum, but the donor's serum agglutinated
Aj and A 2 erythrocytes and failed to agglutinate group B or group O cells. On
prolonged observation, moreover, especially
when anti-B blood grouping serum of very
high titer and avidity was used, small
clumps of erythrocytes were observed, although the bulk remained unagglutinated.
The clumps were barely visible to the
naked eye, and on microscopic examination it was found that only about a tenth
of the erythrocytes took part in the agglutination (Table 1). In fact, on casual examination with the naked eye, the clumps
could readily be overlooked, and the reaction pronounced as negative. The erythrocytes were strongly agglutinated by anti-H
lectin (Ulex europeus). In order to throw
further light on the problem, saliva was obtained from the blood donor, but unfortunately he proved to be a nonsecretor.
Blood and saliva samples were then obtained from other members of the blood
donor's family, and the results of those tests
are shown in Table 2.
As can be seen, the Rh-Hr tests indicate
that the paterfamilias actually is not the
father of the proband. However, this proved
to be irrelevant to the deductions regarding the proband's A-B-O group possible
from the family study, since the significant
findings proved to involve the mother. As
shown in Table 1, in the A-B-O tests the
mother's blood reacted exactly like the
blood of the proband, namely, in tests with
anti-B serum (either group A serum or
group O serum), agglutination occurred but
only about one tenth of the erythrocytes participated in the reaction. Moreover, the mother's serum, like that of the
son, contained natural anti-A isoagglutinins
but no anti-B. Her erythrocytes were
strongly agglutiated by anti-H lectin, and
saliva tests proved her to be a nonsecretor,
like her son. The erythrocytes of both
mother and son, though giving atypical reactions in the A-B-O tests, gave perfectly
A.J.CP.—Vol.
58
normal agglutination reactions in tests for
blood factors of other blood group systems
such as M-N, Kell, and Rh-Hr.
Titration experiments on the sera of the
proband and his mother further confirmed
the presence of anti-A and the absence of
anti-B and showed the anti-A isoagglutinins
to be of low titers. The proband and his
mother were both given intramuscular injections of 0.5 ml. of a preparation of blood
group substance A (Armour) and the titrations were repeated 10 days later, with the
results shown in Table 3. As is to be expected, the anti-A titers showed a marked
rise, and it is noteworthy that there was
also a qualitative change in that after the
injections the sera agglutinated Ax and A 2
erythrocytes about equally, whereas before
the reactivity had been much greater for Ax
than for A2 erythrocytes. At no time was
any reactivity for group B cells detected.
The proband was given a second injection
of group A substance, but there was no further significant change in titers.
Discussion
The incomplete agglutination of the
erythrocytes of the donor and his mother
by anti-B reagents is entirely comparable to
the reactions of A 3 erythrocytes with anti-A
reagents, although the proportion of erythrocytes agglutinated, one tenth for the proband and his mother, is lower than in our
former case of A3, in which a third to half
of the erythrocytes were clumped. Therefore, the designation of "B 3 " for this rare
type of blood appears appropriate. Unfortunately, the symbol "B 3 " has already been
used in previous reports 4 - 7 to describe rare
variants of B with quite different properties. The authors trust, therefore, that the
designations for the other B variants will
be changed and the designation "B 3 " retained only for the presently described
variant, which evidently is analogous to A3.
December 1972
695
A GROUP B ANALOGUE OF SUBGROUP A,
Table I. Composite Table of Titrations Comparing the Reactions of the Erythrocytes of the
Subgroup Bj Proband and His Mother with Those of an Artificial Mixture of Group O
and Group B Erythrocytes
Reactions* with Anti-B Serum Diluted
Erythrocytes
lludil.
Blood donor
(l.M.)
uner
Donor's mother
(A.B.M.)
Controls
Croup O
Group B
1 part group B
and 9 parts
group O
1 part group B
and 4 parts
group O
I
1:2
1:16
1:4
1:32
l/5(++)
l/10(+±) l/10(+±)
l/5(+±)
0(+)
1/10(+)
1/10(+)
1/10(+)
1/10(++)
l/10(+±) l/10(+)
|/10(+)
1/10(+) 1/I0(+)
1/10(±)
+++
++±
++1
+++
!/!()(++) l/10(+±)
!/!()(+)
I/I0(+)
l/10(+±)
1/5(T+±)
l/5(++)
l/5(++)
l/5(+±) l/5(+±)
1:256
1/10(±)
+±
++
!/!()(++)
l/5(++)
1:128
1:6<1
1/I0(+)
1/5(+±)
l/5(+)
1/10(1)
*Thc strength of the reactions is indicated by the number of plus signs, and the fractions before the
parentheses represent the proportions ol erythrocytes agglutinated, e.g., l/3(++) would indicate distinct
agglutination visible 10 the unaided eye.lwiih only one third of the cells taking part in the clumping and
two thirds of the cells free.
As in our previous case of A3) the agglutinogen B 3 has been found in a mother
and son. Presumably, therefore, subgroup
B 3 is determined by a corresponding allelic
gene Bs, analogous to the gene A3 for agglutinogen A3. Evidently the mother (as
well as the son) was heterozygous, genotype
B30, since the proband's sister was group O.
That the B 3 erythrocytes, like subgroup
A3 cells, were strongly agglutinable by antiH lectin was to be expected, in view of the
reciprocal relationship between A-B and H.
That the factor C, shared by agglutinogens
A and B 6 , played no role was evident,
since group O sera gave no stronger reac-
tions with the B 3 erythrocytes than with
anti-B (group A) serum. Again, the situation is comparable for A 3 erythrocytes,
which react about the same with group O
and group B sera, in contrast to so-called
group A0 or A x erythrocytes, better named
group "C", 6 which are agglutinable by
group O sera but not by anti-A or anti-B.
In tests of saliva of individuals of subgroup A3, the presence of A substance was
readily demonstrated by Gammelgaard 3 (p.
71), although the average inhibition titer
was slightly lower than for A2 and Ax saliva. Presumably, therefore, secretor B 3 saliva will also prove to contain B substance
Table 2. Grouping Tests on Blood Donor of Subgroup B3 and His Family
Donor ol
Blood
Specimen
Father*
Mother
Son (proband)
Daughter (sister
ol proband)
Cousin ol proband
Ei yt Inocyies
A - li- O
S' on ps
M-N
type
Kell
type
Rh-Hr
type
Saliva
for
A-B-11
B
B,
B,
MN
M
MN
k
k
k
Rh0
Rho
Rh 2 rh
Secretor
Non-secretor
Non-secrelor
O
O
MN
M
k
k
Rh0
Rho
Secretor
Not available for testing
•Note that die results of the R h - H r tests exclude paternity.
696
A.J.C.P.—Vol.
WIENER AND CIOFFI
58
Table 3. Effect of Injection of Blood Group Substance A on the
Isoantibody Titers of the Proband and His Mother
Serum
from
Proband
Preinjection
Postinjeclion
Mother of proband
Preinjection
Postinjeclion
l i t e r s in Saline Solution for
Titers in Acacia for
A2
B
A,
A2
B
6
48
0
8
0
0
56
160
2
64
0
0
12
192
1
192
0
0
56
256
2
256
0
0
Ai
in a lower inhibition titer for anti-B than
ordinary B saliva, but in our case the proband and his mother were both nonsecretors.
If agglutinogen B 3 is due to a special
allelic gene B3, as appears likely, individuals of subgroup AB 3 must exist, although extremely rarely. Because of the
mutual suppressive action of genes A and
B in individuals of group AB, agglutinogen B 3 would be expected to be much more
weakly expressed in subgroup AB 3 than in
subgroup B 3 . Since subgroup B 3 is so readily
mistaken for group O, subgroup AB 3 would
almost surely be mistaken for group A,
unless reverse grouping were done and no
agglutinins found in the serum, because
the very weak reactions due to the B3 gene
would probably be overlooked. Such an error would almost surely have no serious clinical consequences, because transfusion of
B 3 blood to a group O recipient or transfusion of AB 3 blood to a group A recipient
would hardly be likely to give rise to any
hemolysis; moreover, recipients of group B 3
or AB 3 could undoubtedly receive group O
blood without any reaction.
In line with the reciprocal relationship
between agglutinogens and isoagglutinins,
it is not unusual to find an irregular isoagglutinin in the serum when the corresponding agglutinogen is only weakly reactive. For example, the sera of about 5% of
individuals of subgroup A 2 contain the irregular isoagglutinin anti-Ax, reactive for
Ax but not A 2 cells, and in subgroup A2B
as many as one third of the sera have
anti-Ax isoagglutinins. Similarly, some very
rare weakly reactive group B blood samples
have been detected with serum containing
anti-B reactive for all group B cells except
those of the individual himself. In the
present case, however, no anti-B could be
demonstrated in the sera of the two B 3
individuals.
The most interesting problem is the nature and cause of the picture of partial
agglutination with anti-B reagents given by
the erythrocytes of the two individuals of
subgroup B 3 . The simplest explanation is
that in these individuals some erythrocytes
carry the agglutinogen B on their surface
but most have no agglutinogen B. In fact,
the reactions of B 3 erythrocytes with anti-B
and anti-H reagents could be duplicated
with a mixture of nine parts of group O
and one part of group B erythrocytes (Table 1). We attempted to separate the two
postulated kinds of erythrocytes by differential centrifugation, but without success.
That the proband and his mother are
chimeras is hardly likely; first, neither he
nor his mother was a twin; second, no evidence of partial agglutination occurred in
tests for blood factors other than B. A
plausible explanation is that subgroups A 3
and B 3 are comparable to the piebald trait,
which in man as well as in animals may be
determined by a unit gene. Accordingly, it
may be postulated that genes A3 and B3
December 1972
A GROUP B ANALOGUE OF SUBGROUP A„
give rise through appropriate enzymes to
two different kinds of clones of erythroblasts in the marrow; one clone produces
erythrocytes not agglutinable by either
anti-A or anti-B, whereas the other clone
produces agglutinable erythrocytes. This
conforms to the hypothesis suggested by
Cotterman l for A 3 blood, which he considers to be a mosaic of A2 and O erythrocytes.
Acknowledgments. Nancy Savage, Nancy Shackles,
and Madeline Stever of Eastern Biologicals assisted
in obtaining the blood and saliva samples for this
investigation, and Pat Ryan, Sybil Gordon, and
Dina Schoenwetter assisted in carrying out the
experiments.
2.
3.
4.
5.
6.
References
7.
I. Cotterman CW: Somatic mosaicism for antigen
A2. Abstracts, First Cong H u m Genet 1956, p
697
94. Cited after Race RR, Sanger R R : Blood
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FA Davis, 1968, p 18
Friedenreich V: Eine bisher unbekannte Blutgruppeneigenschaft (A0). Z Immunitaets forsch
89:409-416, 1936
Gamraelgaard A: On rare, weak A antigens (A0,
Aj, A5 and Ax) in man. Edited by Arnold Busck.
Copenhagen, Denmark, A/S, 1942. Reprinted by
Walter Reed Army Institute of Research,
Walter Reed Army Medical Center, Washington, D.C., 1964
Sussman LN, Pretshold H, Laeher MJ: An unusual gene of blood group B. Proc 8th Congr
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143-147
Wiener AS, Silverman IJ: Subdivisions of group
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Wiener AS, Ward FA: T h e serological specificity
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subgroup of the blood group B. Vox Sang 2:
348-356, 1957