The effects of selection of pigs on growth rate vs leanness on histochemical characteristics of different muscles.

The microstructure of two type of muscles was studied in a selection experiment conducted with Dutch Large White pigs (boars and gilts) selected for either low backfat thickness (L-line) or fast growth (F-line). Second- and fourth-generation pigs were used to determine effects of selection on fiber type composition, fiber area, and capillary density in the longissimus lumborum (LL) and biceps femoris (BF) muscles. Immediately after slaughter samples were taken from the LL and BF muscles. The latter was divided into an inside (BFi) and outside (BFo) portion, which refer to the red and white portions of the biceps femoris. Serial sections were stained for ATPase (pH 4.60), succinate dehydrogenase (SDH), and alpha-amylase-periodic acid shiff (PAS) to determine fiber type and capillary density. The LL and BFo muscles had predominantly type IIBw fibers, whereas the BFi muscle had a 2 to 4 times higher amount of type I fibers. In most muscles there were more type I and fewer type IIBw fibers in F- than in L-line pigs (P < .05), except in the LL muscle of second-generation pigs and in the red part of the BF muscle of fourth-generation pigs. In both selection lines lower type I and higher type IIBw percentages were found in muscles from gilts than in those from boars (P < .05). Capillary density and fiber area of L- and F-lines showed minor differences, which could be explained by differences in weight and age of the pigs of both lines. The results suggest that selection for low backfat thickness in pig breeding compared with increased growth rate resulted in fewer oxidative and more glycolytic muscle fibers. The magnitude of the effect depended on muscle type and duration of the selection period.

The production of anti-H-Y monoclonal antibodies: their potential use in a sex test for bovine embryos.

In an attempt to improve the accuracy of sexing bovine embryos, new anti-H-Y monoclonal antibodies were produced and selected, using an extended screening procedure. In addition to the commonly used screening of soluble H-Y antigen sources, such as testis supernatant and Daudi supernatant, the binding specificity to cell surface H-Y antigen was tested also. A radioimmunoassay (RIA) employing male and, as a control, female bovine lymphocytes, and enzyme-linked immunosorbent assays (ELISAs) on solubilized membrane fractions resulted in the selection of a number of clones producing monoclonal antibody (mAb) with male-enhanced binding. Four of the anti-H-Y mAb were assessed for binding to Day 7 or 8 bovine embryos. The accuracy of sexing bovine embryos ranged from 58% to 71%. Two of the four antibodies did not react with presumed soluble H-Y antigen-containing sources in an ELISA. These results raise doubts about the suitability of the presumed soluble H-Y antigen sources, Daudi, TM4 and testis supernatant, to be used in screening tests for anti-H-Y antibodies.

Binding of anti-H-Y monoclonal antibodies to separated X and Y chromosome-bearing porcine and bovine sperm.

Studies designed to answer the question whether or not H-Y antigen is preferentially expressed on Y chromosome bearing sperm have resulted in conflicting results. This is probably due to the absence of reliable methods for estimating the percentage of X and Y chromosome bearing sperm in fractions, enriched or depleted for H-Y antigen positive sperm. In recent years a reliable method for separating X and Y chromosome bearing sperm has been published. With this method, separation is achieved by using a flow cytometer/cell sorter, which detects differences in DNA content. This technique provided the first opportunity for testing anti-H-Y antibody binding to fractions enriched for X and Y chromosome bearing sperm, directly. A total of 7 anti-H-Y monoclonal antibodies were tested using sorted porcine sperm and in one experiment also sorted bovine sperm. All monoclonal antibodies bound only a fraction of the sperm (20 to 50%). However, no difference in binding to the X and Y sperm enriched fractions was found. Therefore, the present experiments do not yield evidence that H-Y antigen is preferentially expressed in Y chromosome bearing sperm.

Construction of a bovine-murine heteromyeloma cell line; production of bovine monoclonal antibodies against rotavirus and pregnant mare serum gonadotrophin.

Bovine-murine heteromyeloma cell lines were prepared by fusing lymphoid cells from a bovine leukemia virus (BLV)-infected cow with mouse myeloma cells. Selection of hybrid cell colonies was based on the ratio of bovine and murine chromosomes, the presence of cell-surface immunoglobulins and growth characteristics. First-generation fusion partners were compared for fusion efficiency and the number of antigen-specific antibody-producing clones generated. Hybrid cell colonies that initially secreted antibodies were selected from first-generation heteromyelomas to function as second-generation fusion partners. Although fusion efficiencies for both generations did not differ, the second-generation heteromyelomas yielded a higher number of specific antibody-producing clones. Fusion of hteromyelomas with either lymph node cells or splenocytes indicated that fusion with lymph node cells results in a higher number of specific antibody-producing clones, whereas fusion efficiency was found to be higher with splenocytes. The optimal time intervals between the final booster injection and fusion were found to be 4 days for splenocytes and 7 days for lymph node cells. Finally, the characterization of bovine monoclonal antibodies against bovine rotavirus and pregnant mare serum gonadotrophin and their neutralizing capacities in vitro are described.

Production and characterization of monoclonal antibodies against the H-Y antigen.

Monoclonal antibodies against the H-Y antigen were produced using spleen cells from female C57BL/6 mice hyperimmunized with cells from syngeneic males. Anti-H-Y positive clones were detected by enzyme immunoassays. Supernatant fluids from Daudi cell cultures and testicular cell preparations taken from mice, rabbits or calves served as presumptive sources of H-Y antigen. In addition, testis supernatant from genetically sterile mice was used. Male specificity was ascertained by the fact that the antibodies could be absorbed with spleen cells from male but not from female mice. Binding of the antibodies to H-Y antigen on the surface of male and female cells, obtained from a number of tissues and species, was confirmed by an indirect immunofluorescence assay. Several monoclonal antibodies appeared to be positive in all assays tested, suggesting that the molecule conferring the H-Y antigenicity lacks species-specificity and appears to be identical for soluble and membrane-bound H-Y antigen.

Purification of phospholipids by preparative high pressure liquid chromatography.

A method is described for the purification of a number of phospholipids by preparative high performance liquid chromatography (HPLC). Purification of digalactosyl-diglyceride from spinach and egg phosphatidylcholine, 1,2-dioleoyl-sn-glycero-3-phosphocholine and 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine from its reaction mixture have been resolved. The lipid separation is performed on a polygosil column and the individual compounds are monitored directly by refractive index detection. Chloroform/methanol mixtures are used as eluent systems, providing a wide polarity range to separate the classes of lipids. The developed equipment can be used for columns between 10 and 50 cm long and 4 and 50 mm inner diameter. The flow rate could be varied between 1 and 100 ml/min and applied pressures between 10 and 450 bars.