Evaluation of the presence of a specific histocompatibility protein on equine embryonic cells

An indirect immunofluorescence assay was used to detect the presence of H-Y antigen on equine blastocysts. A total of 33 blastocyst stage horse embryos were collected 6 to 7 days post-ovulation by trans-cervical flush and were immediately evaluated for the presence of H-Y antigen. Additionally, 17 embryos, were collected and cultured for 72 h to the expanded blastocyst stage and similarly evaluated. Embryos were placed in medium containing monoclonal antibodies to H-Y antigen followed by incubation in medium containing 1/10 (v/v) fluorescein isothiocyanate conjugated goat anti-mouse IgM Fc specific antiserum. Embryos were individually evaluated at 400X to identify cell specific fluorescence. Following evaluation, embryonic sex was independently verified with karyotypes to identify sex chromosomes. Of the 50 embryos evaluated, 29 were evaluated as non-fluorescent and 21 fluorescent. Expression of H-Y antigen was determined to be uniform in those embryos classified as fluorescent. Twenty-three of 28 (82%) readable karyotypes corresponded to the predicted sex. These results indicate a specific histocompatibility antigen is expressed and maintained at the blastocyst stage of development. In addition, no segregation of this protein on specific cell types occurs in this species.