The release of soluble H-2 alloantigens during disaggregation of mouse embryo tissue by a chelating agent

Treatment with chelating agents binding divalent cations has been found to effect the dissociation of a variety of tissues of both embryo and adult animals (reviewed in Steinberg, 1958). In the course of dissociation it appears that materials are released from cell surfaces which play a part in their specific adhesion, and which may be shown experimentally to promote selectively the re-aggregation of dissociated cells (Humphreys, 1963; Moscona, 1963). The extracted materials appear to be glycoprotein complexes (Humphreys, 1965), made up of fairly small subunits, estimated to be of 13000-20000 molecular weight (Margoliash et al. 1965). Units of about the same size appear to be the antigenic sites involved in the blocking of sponge cell aggregation by rabbit anti-sponge serum, specific for a given sponge species (MacLennan, 1963). I shall here present evidence that materials of similar molecular weight bearing immunological specificities of the H-2 alloantigen system are released from the tissues of certain mouse embryos during the course of their dissociation by the chelating agent Versene (ethylenediaminetetraacetic acid). Antigens of the H-2 system function as transplantation antigens determining the fate of allogeneic homografts, as immunogens stimulating the production of circulating antibody, and as antigens of erythrocytes and leucocytes reacting with antibody (Stimpfling & Snell, 1961; Snell, Hoecker, Amos & Stimpfling, 1964). The antigens extracted by Versene are most conveniently detected by their ability to inhibit the cytotoxic action of antibody produced by tissue allografts made between mouse strains differing at the H-2 locus. The potency of the antigens extracted is thus expressed in terms of their inhibition of a standard cytotoxic alloantiserum. This technique allows determination of very small amounts of material and it avoids the problems of immunization and quantitative scoring of skin graft survival, which are encountered when H-2