Production of alloantibodies against bovine B-lymphocyte antigens.

A series of alloimmunizations were carried out between BoLA class I antigen typed bulls, with the aim of generating class II specific reagents. Of the antisera produced, seven demonstrated exclusively B cell reactivity. Another 19 sera reacted with both T and B cells from some animals and with B cells only in other cases.

Biochemical evidence that equine leucocyte antigens W13, W22 and W23 are present on horse major histocompatibility complex class II molecules.

A number of horse alloantisera were characterized biochemically as being directed against MHC class I or class II antigens by immunoprecipitation of the corresponding antigens from lysates of biosynthetically radioactively labelled lymphocytes and determination of their molecular weights by SDS-PAGE and fluorography. Sera recognizing A2 and A3 specificities precipitated antigens of 44,000 Daltons molecular weight (class I heavy chain), whereas sera with specificities W13, W22 and W23 precipitated antigens corresponding to class II dimers (30,000 and 32,000 Daltons). Comparison with antigens precipitated from horse lymphocyte lysates using (cross-reacting) antibodies to human class I and class II MHC molecules confirmed the results obtained.

Restriction fragment length polymorphisms of horse class II MHC genes observed using various human alpha- and beta-chain cDNA probes.

Genomic DNA isolated from 20 horses was digested with up to six restriction endonucleases and subjected to southern blot hybridization analysis using various human class II alpha- and beta-chain cDNA probes. A high degree of restriction fragment length polymorphism (RFLP) was found for the DQ alpha, DP beta, DQ beta and DR beta probes, about 20 polymorphic bands being detected for each. DR alpha showed 2-4 polymorphic bands, whereas no evidence for DP alpha-like genes was found.

Rapid data acquisition from a microtiter plate fluorescence reader and applications in kinetic measurements.

Programs written in Applesoft BASIC for the rapid acquisition and evaluation of data from a commercially available microtiter plate fluorescence reader are presented. Using the data acquisition program, the relative fluorescence readings from all 96 wells of the microtiter plate (one read cycle of the fluorescence reader) can be stored in each of up to 90 consecutively numbered files on a single-sided diskette. A simple timer circuit is described which, when used in conjunction with the above program, initiates the fluorescence reading process at preset time intervals, thus making automatic acquisition of data possible.

Localization of galactosyl- and sialyltransferase by immunofluorescence: evidence for different sites.

Polyclonal rabbit antisera against soluble human milk galactosyltransferase and bovine colostrum sialyltransferase were used to localize by indirect immunofluorescence the respective intracellular enzymes in primary cultures from bovine fetal kidneys and established cell lines of human and bovine fibroblasts. Staining for galactosyltransferase was juxtanuclear and crescent shaped in epitheloid cells; a similar staining, occasionally perinuclear and sparsely distributed in the cytoplasm, was found in fibroblasts. In contrast, staining for sialyltransferase in epitheloid kidney cells derived from the same primary culture was observed predominantly in cytoplasmic vesicles that were spread over the whole cytoplasm.