Studies on the blood of an MiV homozygote.

An individual, whose parents are third cousins, has been shown to be homozygous for the rare Mi.V. condition. The proposita's red blood cells type as M-, N+(weak), S-, s+(strong), U+, Mi(a-), Vw-, Hil+; Wr(a-b-). The cells react, albeit less strongly than most other samples, with anti-Ena. However, from studies on the red blood cells of the proposita and on those of another person of the En(a+), Wr(a-b-) phenotype, it is apparent that the term "anti-Ena" actually describes a number of antibodies of differing specificities. Inhibition studies with sialoglycoprotein (SGP) isolates, and tests on protease-modified red blood cells illustrate some of the differences in specificity. Biochemical analyses of the SGPs of the red blood cells of the MiV homozygote and those of her parents confirm that the Mi.V condition is associated with the absence of normal MN SGP (alpha) and normal Ss SGP (delta), the appearance of a hybrid SGP molecule comprised of a portion of the MN SGP at its NH2 terminal end, and a portion of the Ss SGP at its C terminal end.

Another individual (J.R.) whose red blood cells appear to carry a hybrid MNSs sialoglycoprotein.

The serum of J.R. contains anti-Wrb and her red blood cells are of the phenotype Wr(a-b-). Evidence was obtained that suggested that her cells totally lack normal MN and Ss sialoglycoproteins (SGPs), and carry instead an abnormal SGP, which is likely to be a hybrid SGP resembling the MN SGP in its outer portion and the Ss SGP in its inner portion. Although the apparent hybrid SGPs of J.R. and the MiV homozygote are virtually indistinguishable in terms of their electrophoretic mobility (apparent molecular weight 40,000) and staining characteristics, they are not identical. That of J.R. is associated with a weak M and increased S antigen, while that of the MiV homozygote is associated with a very weak N, a greatly exalted s, and the rare antigen, Hil. Like the study on the blood of the MiV homozygote, serologic studies on the red blood cells of J.R. have revealed considerable heterogeneity of what have previously been called the Ena antigen and anti-Ena antibodies.

An auto-anti-Ena, inhibitable by MN sialoglycoprotein.

An auto-anti-Ena, which reacts with trypsin, but not with ficin-treated red blood cells, and which can be totally inhibited with sialoglycoprotein (SGP) isolates from red blood cells, is described. From comparative studies on this antibody and on the four known examples of allo-anti-Ena, it is clear that the term "anti-Ena" describes a heterogeneous group of related but not identical specificities. The specificities contained within the auto-anti-Ena described are different from those within any of the sera containing allo-anti-Ena. Several of the specificities that have been included under the blanket term, anti-Ena, complex with the MN SGP of normal red blood cells, but recognize different portions of that polypeptide.

Absence of autoanti-Jk3 as a component of anti-dl.

Thirty-five examples of anti-dl, autoantibodies initially reacting with all red blood cells against which they were tested, were examined prior to, and following adsorption with Jk(a-b-) red blood cells. None of them was found to contain autoanti-Jk3. One example of autoanti-Jka was identified in an adsorbed eluate. The antibody had been produced by a pregnant Caucasian, in whom laboratory and clinical evaluation showed that the auto-antibody was benign in vivo.

Anti-Rh39--a "new" specificity Rh system antibody.

Two examples of an autoantibody that defines a hitherto unrecognized Rh system antigen are described. Both were produced by C-negative individuals and ostensibly resembled anti-C in specificity. However, adsorption studies showed that the antigen that the autoantibodies define is present on all red blood cells with a "normal" Rh phenotype and on D--/D-- and Dc--/Dc--samples. The antigen detected is not present on Rhnull red blood cells. Serologic studies have shown that the new antibody, that has been named anti-Rh39, has a different specificity from those that define the antigens C, Ce(rhi), G, Hro, Hr, CG, LW, Rh:29, Rh:34 and U. A possible relationship between auto-anti-Rh39 and allo-anti-C, in terms of the immune response, is discussed.

Allo-anti-C in a patient who had previously made an autoantibody mimicking anti-C.

A patient, previously studied by us, who had produced a benign autoantibody with a specificity that mimicked anti-C, has now produced allo-anti-C that is not of the mimicking type. She is no longer producing any serologically demonstrable autoantibody. It is highly probable that conversion from auto to alloantibody production in this patient was prompted by the introduction of additional foreign immunogen on fetal red blood cells during her third pregnancy.

The Can serum: demonstrating further polymorphism of M and N blood group antigens.

An antibody defining an antigen that is very similar in structure to M is described. The reactions of this antibody are considered in the light of what is now known of the biochemical structure of the M and N blood group antigens.

Antibodies that define NANA-independent MN-system antigens.

The hemagglutinating properties of a large proportion of anti-M and anti-N reagents, and sera containing antibodies to MN-related antigens, have been shown to be unaffected by treatment of red blood cells with neuraminidase. These antibodies, which define NANA-independent MN-system structures, provide further evidence that MN blood group specificity may also be determined by moieties other than N-acetylneuraminic acid.

Notes on the Ena and U antigens of Ena(En.OP) heterozygotes, and the U antigen of En(a--) individuals.

Using two anti-Ena antibodies, from which anti-Wrb had been removed, and anti-U antibodies, the authors have failed to demonstrate any difference in the amounts of Ena and U antigens on the red blood cells of individuals with one and two functioning Ena(En.OP) genes. Two En(a--) samples, one from an individual who is En/En (En.op/En.op) and the other En/Mk (En.op/Mk), could not be distinguished from U-positive samples from individuals with two functioning U (U.OP) genes in dosage studies with anti-U.

Demonstration of anti-Wrb in a second serum containing anti-Ena.

Many persons who are of a "deletion" or "null" phenotype with regard to a particular blood group system, form a complex specificity antibody or mixture of antibodies, when immunized. Recently, we demonstrated that M.E.P., the En(a-), Wr(a- b-) proposita in a family described by Darnborough has at least, anti-Ena and anti-Wrb in her serum. We suggested that the same two antibodies might be present in the serum of G.W., the En(a-), Wr(a- b-) propositus in the family reported by Furuhjelm et al. We have now been able to confirm that G.W.'s serum does contain antiEna and anti-Wrb and that the antibodies can be separated by adsorption. Because, at this time, it is not known if En(a-), Wr(a- b-) people lack other common antigens from their red blood cells, the possibility remains that the anti-Ena, separated from the anti-Wrb, might be a mixture of antibodies. These findings are important in that they show that reported typings for Ena were actually performed with mixtures of at least anti-Ena and anti-Wrb. Anti-Ena, lacking antiWrb, can be made only by adsorption of a serum containing the two antibodies, onto En(a+), Wr(a+b-) red blood cells, with recovery of the anti-Ena by elution.

Critical re-examination of the specificity of auto-anti-Rh antibodies in patients with a positive direct antiglobulin test.

Forty-eight autoantibodies with apparent 'simple' anti-Rh specificity (anti-e, -E, -c, -D, -C, -Ce, -G), have been studied by means of multiple absorption tests. The finding that 34 (70.8%) of these antibodies could bind to red blood cells lacking the antigens that the antibodies appeared to define, indicated that the antibodies had different specificities than seemed to be the case in initial antibody identification tests. Those autoantibodies that at first appeared to be directed against the Rh antigens e, E or c, most often had anti-Hr or anti-Hro specificity. These data explain why some apparent anti-Rh autoantibodies can be eluted from the red blood cells of patients negative for the antigens that the antibodi:s appear to define. However, they also illustrate that the phenomenon of autoantibodies mimicking specificities that they do not possess is common in patients positive for the antigens against which their autoantibodies appear to be directed. An explanation for the mode of action of these autoantibodies in complexing with the Rh agglutinogen is proposed, and the significance of the antibodies in transfusion therapy is considered.

A "normal" individual with a positive direct antiglobulin test: case complicated by pregnancy and unusual autoantibody specificity.

A "normal" individual with a positive, direct antiglobulin test is described. In common with many other "normal" persons in whom a similar finding has been made, there was no evidence of an increased rate of in vivo red blood cell destruction in this patient. The patient successfully completed a pregnancy during the time that detailed serologic studies on her autoantibodies were being performed. Although the maternal autoantibodies were demonstrable in both an eluate made from the red blood cells of her newborn infant, and in the cord serum, there was no reason to believe that the antibodies caused red blood cell destruction in the infant. The case was of further interest because of the specificities of some of the autoantibodies. Although the mother and child were both C-negative, eluates from their red blood cells contained what ostensibly appeared to be anti-C. Studies that showed that the antibody could be totally adsorbed with C-negative, as well as C-positive red blood cells, proved that this was another example of an autoantibody mimicking an alloantibody. Although this autoantibody appeared initially to have anti-C specificity it was eventually shown to be more closely related to anti-Hr or anti-Rh34, than to anti-C.

Independence of Wright from many other blood group systems.

The finding that En(a-) red blood cells are Wr(a-b), and thus probably represent a "null" phenotype of the Wright system, has provided evidence of the independence of the Wright blood group system from many others. Detection of blood group antigens on "null" red blood cells almost certainly indicates that those antigens do not belong to the same blood group system as the "null" cells. In the case of Wright, lack of certainty that En(a-) is the "real", or only, null type slightly reduces the weight of the evidence.

The phenotypes En(a-), Wr(a-b-), and En(a+), Wr(a+b-), and further studies on the Wright and En blood group systems.

In 1975, we showed 18, 19 an En(a-) blood sample to be phenotypically Wr(a-b-). In the current report, we describe tests that show that three En(a-) members of a single family, not believed to be related to the family of the previously tested En(a-) person, are also Wr(a-b-). They have red blood cells that neither react with nor adsorb anti-Wra or anti-Wrb. In addition, we have shown that the red blood cells of six EnaEn heterozygotes, in the family tested, are Wr(a-b+) but carry only a single dose of Wrb antigen. Tests on anti-Ena have shown conclusively that one example is a mixture of separable anti-Ena and anti-Wrb and that a second example may well contain the same two antibodies. By various methods, we have demonstrated that the red blood cells of the only known Wr(a+b-) individual are En(a+) and do not display any of the physicochemical abberations of the En(a-) phenotype. It is further shown that neuraminidase and trypsin do not denature the Wra or Wrb antigens in vitro, but that the protease ficin does have a limited ability to denature Wrb. Additional observations on the first reported example of anti-Wrb are included. These various findings have been considered in the light of gene linkage of, or gene interaction between, the En and Wright system genes. It is concluded that the evidence does not exclude the possibility that En is a silent allele at the WraWrb locus so that the genotype EnEn (or WrWr) might result in the phenotype En(a-), Wr(a-b-). However, it is also pointed out that the evidence equally well supports the postulation that the Wra and Wrb genes are unable to function in the absence of an Ena gene. If this latter theory is proved correct, the interaction between Ena and the Wright genes can be thought of as similar to that between the H and ABO or X1r and CDE genes. It is pointed out that if En is a silent allele at the MN locus (current evidence on this point is not conclusive,) En and Wr cannot be synonymous for it is known that the Wra and M and N genes segregate independently. Location of En at the MN locus would not, however, refute the theory that Wra and Wrb cannot function in the absence of En. Finally, it is pointed out that the supposed anti-Wrb is probably just what its name implies but that even if this assumption is later disproved, the high incidence antigen defined by the antibody presently called anti-Wrb is unequivocally associated with Ena.

Anti-Wrb, and other autoantibodies responsible for positive direct antiglobulin tests in 150 individuals.

Eluates from the red blood cells (and sera whenever free autoantibody was present) of 150 individuals with positive direct antiglobulin tests, have been studied for antibody specificity. Of 87 patients with AIHA, 64 had autoantibodies reacting with all red cell samples including Rhnu11. Of these 64 anti-d1 autoantibodies, two were, and 32 contained, auto-anti-Wrb. Of 33 patients being treated with alphamethyldopa, who had developed positive direct antiglobulin tests, 23 had anti-d1 autoantibodies four of which contained auto-anti-Wrb. Of 30 haematologically normal donors with positive direct antiglobulin tests, 23 had anti-d1 autoantibodies, two of which were, and six of which contained, auto-anti-Wrb. The full specificities of autoantibodies, other than anti-Wrb and anti-d1, in the 150 patients are described, as are the natures of the protein red cell coatings that caused the positive direct antiglobulin tests. The presence of free serum autoantibody as a correlate of the three clinical conditions is reported. Several observations on auto-anti-Wrb are documented. The antibody can cause gross red cell destruction in vivo, but can be benign on other occasions; it occurs with approximately the same frequency in AIHA patients and "normal" donors with positive direct antiglobulin tests, but in fewer patients with alphamethydopa induced positive direct antiglobulin tests; it does not activate complement in vivo; and finally it may eventually provide a clue to the aetiology of AIHA.

Hemolytic disease of the newborn due to anti-N.

A Negro woman of phenotype M+, N-, S-, s-, U+, who produced anti-N, is described. The antibody in her serum caused hemolytic disease of the newborn in her M+, N+ infant.

Failure to demonstrate dosage of U antigen.

Tests in which 11 examples of anti-U were used in titration studies against the red blood cells of 9 obligate Uu heterozygotes, from 4 unrelated families, and random Negro and Caucasian donors (many of whom were of the presumptive UU genotype) have failed to demonstrate any dosage of the U antigen.

An En(a-) red cell sample that types as Wr(a-b-).

In the course of investigating a patient with autoimmune hemolytic anemia in which the causative autoantibody had anti-Wrb specificity, it was demonstrated that an En(a-) red blood cell sample typed as Wr(a-b-). The only known example of Wr(a+b-) blood typed as En(a+) so that anti-Wrb and anti-Ena do not have the same specificity.

Successful transfusion of Chido-positive blood to two patients with anti-Chido.

Two cases are described in this report in which patients with anti-Chido in the serum were transfused with Chido-positive blood. Since there was evidence of normal survival of the transfused red blood cells, these findings do not support a suggestion that patients with anti-Chido may require transfusion with Chido-negative blood. In spite of the apparently normal survival of the Chido-positive blood, a previous report in which it was shown that weakly Chido-positive blood can stimulate the production of anti-Chido was confirmed.

Some observations on "Bombay" bloods, with comments on evidence for the existence of two different Oh phenotypes.

Bloods from three individuals, one each of the phenotypes Oh-A, Oh-B and Oh-O have been studied. The work of Dzierzkowa-Borodej, et al.-10 was confirmed when it was shown that all three samples of Oh red blood cells had increased I antigen strength. The i, Sd-a, Le-a and Le-x antigens were not found to be increased. Attempts were made to adsorb and elute anti-A, anti-B and anti-A,B with the Oh red blood cells, using sera that contained high titered anti-I antibodies. This was done in the belief that previously reported positive results in such tests might be due to the high level of I on the Oh red blood cells, anti-I in the sera containing the ABO antibodies, and the Matuhasi-Ogata phenomenon. However, in no instance were we able to adsorb an ABO antibody onto the Oh red blood cells. Contrary to the report of others-10 the titers of anti-A, anti-B and anti-H in the sera of the three Oh individuals studied did not differ significantly. We suggest that the evidence from our findings and the work of others is sufficient to show that at least two forms of the Oh phenotype exist: one representing total suppression of H, A, and B antigens, and the other marked but not total suppression, with partial inhibition of antibody production.