Allotypes detection

The very sensitive hemagglutination inhibition system is most often used to detect human and certain rabbit allotypes since most antisera are not of very high titer or affinity. In the rabbit, certain antiallotype sera (e.g., those sera directed against the group a and b allotypes) are relatively higher in precipitin titer. 

This conclusion is also consistent with studies of Gm allotypic markers characteristic of human IgGl, which are detectable, among other primates, only in the serum of the great apes (129). In addition, allotypic markers characteristic of human IgG3 are found in the great apes and Old-World monkeys but not in the New-World monkeys (130). 

Allotypy has been studied most extensively in the rabbit, man, and mouse. In each species allotypic variation is seen in both H and L chains. In the rabbit and in man, for which data are available, genes controlling allotypic determinants on H and L chains are unlinked. In the mouse, allotypic variability of L chains has been detected by peptide mapping but, so far, not through antigenic analysis. 

A. Detection and Quantitation of Allotypic Determinants in Rabbit Immunoglobulins... 

Studies of allotypy in hare immunoglobulins were also reported by Landucci-Tosi et al. (119) who used insolubilized antiallotype antisera, prepared against IgG of the domestic rabbit, and radiolabeled hare IgG. In addition to the b5 marker observed by Mandy and Rodkey, they detected b4 and b6 on 70-80% of the IgG molecules in a typical preparation b5 was present on 55%. Since the sum exceeds 100%, two or three markers must be present on the same molecule. The markers were found in the IgG of all hares from a variety of geographical sources this work suggests that the markers are isotypic rather than allotypic in hares. The difference in results obtained by the two laboratories may be attributable to differences in the cross-reactivity of the antisera employed. 

It is informative to summarize some of the other rules employed by Herzenberg et al. in interpreting their data (198). (a) The recipient strain has none of the specificities recognized by the antiserum it produces, (b) The immunizing strain has all of the specificities detected, (c) The antiserum may not recognize all of the donor s allotypic specificities first, because the recipient may have specificities in common with the donor and, second, because some potential determinants may not have induced antibody formation, (d) When the labeled antigen used in the test is from a strain (say, strain X) other than the donor, the only specificities detectable are those shared by the donor and strain X. (e) Two strains, each of which shows partial inhibition, may conceivably share no specificities, (f) The specificities detected always represent a minimum estimate of the true number. 

Harrison et al. showed that suppression of allotype b5 in homozygous rabbits was accompanied by a complete absence of circulating lymphocytes having membrane-bound b5 immunoglobulins (220). During recovery from suppression the appearance of b5 molecules in the serum was preceded by the appearance of lymphocytes with membrane-bound b5 molecules. Determinants on the lymphocyte surface were detected with fluorescent or I-labeled anti-b5 antibodies. 

Yakulis et al. (Ill) were able to produce anti-D antibodies specific for two BALB/c myeloma proteins, MOPC 195 and LPC 1, by immunization of BALB/c mice with proteins polymerized with glu-taraldehyde. The anti-D was detected by passive hemagglutination. The enhancement of immunogenicity through polymerization may be explained in terms of the earlier work of Iverson (112), who was unable to produce anti-D against a BALB/c myeloma protein in a second strain of mouse that shared H chain allotype with BALB/c, but was successful in eliciting anti-D antibodies if the protein was first modified with Dnp groups prior to inoculation. 

Allotypes or genetic markers are antigenic determinants on Ig molecules which are inherited in Mendelian fashion and are detected by immunological methods. They have been identified in most mammalian species. 

A number of experiments have been reported by Bell and Dray (1969, 1970, 1971a, b, 1972, 1973) in which the authors describe conversion of non-immune rabbit spleen cells by RNA from immunized rabbit to produce IgM and IgG of donor light chain allotype. Furthermore, spleen cells from non-immunized rabbits were converted to antibody-forming cells by incubation with RNA extracts of lymph node cells obtained from rabbits immunized with SRC. If the RNA was extracted from lymph nodes of rabbits immunized 5 days previously, IgM anti-SRC antibody-forming cells were produced, while if the donor of RNA was immunized 18 to 24 days previously, IgG anti-SRC antibody-forming cells were detected in the converted spleen cells. 

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