Drosophila salivary chromosomes as test system for antinuclear antibody assay.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
J C Herr
Publications and source records attributed to J C Herr.
Identification and isolation of epididymal luminal proteins of the mouse.
Proteins of mouse cauda epididymal fluid were analyzed by polyacrylamide gel electrophoresis. Fluid expressed from the cauda epididymidis and samples obtained by micropuncture of the epididymal lumen showed very similar patterns with respect to the major proteins they contained, with the exception of a small amount of serum albumin found in expressed caudal fluid. Eight prominent peptides present in both micropuncture fluid and expressed caudal epididymal fluid were selected for further study, and were designated CP 47, 42, 35, 29, 27, 25, 18, and 13 according to their mobility. Six of these were never detected in serum. Periodic acid-Schiff staining indicated that at least three were glycoproteins. The epididymal proteins were purified by preparative polyacrylamide gel electrophoresis and electroelution. Upon re-electrophoresis, the individual purified peptides comigrated with the corresponding bands in whole epididymal fluid, and no additional bands were detected, indicating that the proteins were purified to a high degree of homogeneity. Several of the mouse epididymal peptides resemble in their mobility proteins identified previously in other species, most notably the widely studied 33 Kd and 16 to 18 Kd proteins detected in the rat.
Animals↗
The incidence of spermatic granulomas and their relation to testis weight after vasectomy and vasovasostomy in Lewis rats.
The occurrence of spermatic granulomas of the vas deferens was studied in Lewis rats at intervals up to 7 months after vasectomy or vasectomy followed 3 months later by vasovasostomy. The incidence of granuloma progressed with time to involve one or both tracts in 100% of vasectomized rats. In addition, the majority of animals developed new granulomas after vasovasostomy, even though fluid flow through the reconnected vas deferens was demonstrated in vitro. When individual tracts were analyzed, the weight of the testis was related to ipsilateral spermatic granuloma formation in both vasectomy and vasovasostomy groups at 3 and 4 months after initial operation. Testes were small in the absence of a granuloma but similar to those of sham-operated rats if a granuloma was present. The possible protective effect of spermatic granuloma formation on the testis is discussed.
Animals↗
Temporal appearance of antisperm autoantibodies in Lewis rats following vasectomy.
An indirect enzyme-linked immunosorbent assay (ELISA) was employed to monitor antisperm autoantibodies in 16 Lewis rats for up to 36 weeks following vasectomy. This assay was capable of discriminating all prevasectomy from postvasectomy sera at a 1:16 dilution. Weekly serum samples were obtained for the first 13 weeks and bimonthly samples thereafter. Half of the animals developed a positive antisperm autoantibody response by the end of the first postoperative week. By the end of the second week, 81% of the animals had positive responses. The greatest proportion (88%) of animals having a positive response over the course of the study was found at the end of the seventh postoperative week and the highest mean absorbance value for all 16 animals was observed at this time. Only 25% of the animals had positive responses for antisperm autoantibody at the end of the 35th week of the study. These findings indicate that circulating antisperm autoantibodies arise in the Lewis rat earlier than has been generally appreciated. The time course is similar to that of antibody titers to infectious agents or arising from inoculation of rats with spermatozoa. These findings on autoantibody levels in the Lewis rat are compared with the dynamics of antisperm autoantibody formation in man.
Animals↗
Type-A retrovirus particles in heterohybridomas: precaution for human monoclonal antibody purification.
Fine structural features of the murine myeloma MHFP-1 and two heterohybridomas secreting human IgM monoclonal antibody were examined. Intracisternal type-A retrovirus particles were found in both MHFP-1 and the heterohydribomas constructed by fusing MHFP-1 and human peripheral blood lymphocytes. The implications of this finding for the purification of human monoclonal antibody for therapeutic applications is discussed.
Animals↗
Cell growth and monoclonal antibody production in the presence of antigen and serum.
The impact that the continuous presence in the fermentation broth of the cognate antigen has on the serum-supplemented hybridoma cell cultures was investigated. Both soluble and immobilized antigen at various concentrations was applied. The cell line (ATCC TIB191) was cultured in a serum-supplemented PFHM-II medium in T-flasks. Sepharose gel beads provided the immobilization matrix, and bovine gamma-globulin was the carrier protein upon which the antigen, picric acid, was conjugated. Produced antibody, after elution from the beads by displacement with free picric acid, was measured with an ELISA. Soluble antigen--carrier protein conjugates showed no effect on the cultures, but the immobilized antigen had a strong influence on them. Cell growth rate and total antibody production decreased as the amount of immobilized antigen increased from 16:10 (units are in (mol of Ag/mol of carrier):(mg of carrier/mL of beads) to 36:40. Most importantly, the specific antibody production rate switched from growth phase-independent behavior in the antigen-free cultures to growth phase-dependent behavior in the immobilized antigen cultures. Furthermore, during the early stationary phase of the cultures with immobilized antigen, the specific antibody production was 20-100% higher relative to the production rate in antigen-free cultures. This increased specific antibody production rate would be particularly useful in increasing the volumetric productivity of perfusion-type bioreactors for hybridoma cell cultures.
Animals↗
Molecular genetic analysis of two human sperm fibrous sheath proteins, AKAP4 and AKAP3, in men with dysplasia of the fibrous sheath.
Dysplasia of the fibrous sheath (DFS) is characterized by male infertility, asthenozoospermia, and morphologically abnormal flagella that possess a severely malformed fibrous sheath. In many cases, DFS is familial, suggesting a genetic component. Human AKAP4 and AKAP3 are structural proteins of the fibrous sheath that also function to anchor protein kinase A to this structure via the regulatory subunit of the kinase. We hypothesized that defects in either AKAP4 or AKAP3 might cause DFS. No quantitative or qualitative differences between patients with DFS and normal controls were detected when sperm proteins were analyzed by either silver staining or immunoblot analysis using antibodies raised against AKAP4 and AKAP3. Additionally, AKAP4 and AKAP3 from DFS sperm retained the ability to bind the regulatory subunit of protein kinase A. Localization at the light and electron microscopic levels showed that AKAP3 and AKAP4 localized correctly to the FS of the amorphous flagellum in DFS sperm. Partial sequence analysis of the AKAP4 and AKAP3 genes in patients with DFS did not identify any significant alterations in potential AKAP4/AKAP3 binding regions, suggesting that the two proteins interact normally in DFS sperm. Our results did not find evidence to support the hypothesis that mutations in either gene are responsible for DFS in humans.
Adult↗
Assignment of the gene for human intra-acrosomal protein SP-10 to the p12----q13 region of chromosome 11.
The human sperm antigen SP-10 is a testis-specific, intra-acrosomal protein associated with the membranes of the acrosomal vesicle. The molecule has been designated a "primary vaccine candidate" by a World Health Organization (WHO) Taskforce on Contraceptive Vaccines. cDNA cloning and sequencing have indicated that SP-10 is encoded by a 795-base-pair (bp) reading frame that predicts a 265-amino acid protein of 28.3 kd. In this study, we used a 634-bp fragment (bp 68 through 700, amino acids 3 through 222) of the SP-10 sequence to probe, by Southern blotting, EcoRI-digested DNA from 33 mouse/human somatic cell hybrids involving 16 unrelated human cell lines and 4 mouse cell lines. The hybrids were characterized by karyotypic analysis and by mapped enzyme markers. The presence or absence of positive human bands was scored on the blots and the percent of concordance and discordance with a specific human chromosome was determined. The DNA probe for SP-10 showed a concordance of 31 and a discordancy of 0 for human chromosome 11, mapping SP-10 unequivocally to this chromosome. The hybrid XER-7 with the 11/X translocation: 11p12 or 11p11----11qter:: Xq11----Xqter and the hybrid EXR-5CSAZ with the X/11 translocation: Xpter----Xq22::11q13----11qter localized the SP-10 gene to the p12----q13 region. The SP-10 locus has been assigned the gene symbol ACRV1 (acrosomal vesicle protein-1).
Acrosome↗
Temporal appearance of antisperm antibodies during sexual maturation of rats after obstruction of the vas deferens.
The appearance of serum antisperm antibodies was studied during the maturation of rats in which the vas deferens was obstructed prepubertally. Vasa deferentia were ligated and divided bilaterally in 10-day-old Lewis rats, while control animals received a sham operation at the same age. At ages between 14 and 128 days, blood samples were assayed for antisperm antibodies using an enzyme-linked immunosorbent assay. There were no differences between obstructed and sham-operated groups in mean antisperm antibodies from 14 through 56 days. However, in 91- and 128-day-old rats mean antisperm antibody levels were much higher in obstructed than in control animals. This rise in antisperm antibodies occurred several weeks after the development of spermatids in the testes between 23 and 46 days. The presence of elevated antisperm antibodies in obstructed animals followed temporally the appearance of sperm throughout the lumen of the entire epididymis and the formation of spermatic granulomas. The results in this model suggest that consideration be given to repair of injured or obstructed vas deferens prior to puberty to forestall development of antisperm antibodies, as well as the formation of spermatic granulomas.
Animals↗
Sperm autoantigens recognized by autoantibodies in developing rats following prepubertal obstruction of the vas deferens.
Sperm protein autoantigens recognized by serum antisperm autoantibodies during development following vas deferens obstruction were studied using western blot analysis. At age 10 days, rats in an obstructed group underwent bilateral ligation and division of the vas deferens, whereas animals assigned to a sham group received a sham operation. At ages 14, 21, 35, 56, 91, and 128 days, rats were sacrificed and blood samples were obtained. Sperm antigens were recognized infrequently and with low intensity by most sera obtained at ages 14 through 56 days. Subsequently, the incidence as well as the intensity of staining of antigens in blots increased at 91- and 128-day intervals in obstructed animals. The increase in antisperm antibodies correlated with the appearance of sperm throughout the epididymis at approximately 56 days postnatally. A set of autoantigens including proteins migrating at 82-78, 76-73, 68, 57, 54, 48, 44, 42, 38-42, 36, and 22 kDa were recognized with the highest frequency and intensity after puberty. A 42-kDa protein appeared to be one of the first autoantigens recognized when obstructed animals underwent sexual maturation. Although individual animals recognized different patterns of sperm autoantigens, a repertoire of 10-12 autoantigens dominated the antisperm antibody response when obstructed animals underwent sexual maturation.
Animals↗
Epididymal obstruction during development results in antisperm autoantibodies at puberty in rats.
An autoimmune response to sperm occurs after vasectomy, but there is little information on whether similar reactions occur after obstruction of the male reproductive tract at other points. Male Lewis rats received bilateral ligation of the corpus epididymidis or a sham operation at age 10 days, and the subsequent systemic antisperm autoantibody responses were compared to those observed following obstruction of the vas deferens. After sexual maturation, rats with epididymal ligations had antisperm antibodies on an enzyme-linked immunosorbent assay that were significantly higher than those of sham-operated animals and did not differ from antibody levels in vasectomized rats at the same ages. Western blot analysis showed that certain sperm proteins were recognized by antisperm antibodies after both epididymal ligation and vasectomy, including the previously identified "dominant" autoantigens at 73-83, 68-72, 48, 42, and 22 kDa. On the other hand, sera from rats with epididymal ligations recognized 60 and 52 kDa proteins that were not bound by most postvasectomy sera. Conversely, 42-48 and 38-42 kDa bands were more strongly and frequently stained after vasectomy than after epididymal ligation. The results demonstrate that antisperm antibodies are produced after obstruction of the epididymis and that the magnitude of the response is comparable to that after vasal obstruction. Differences in autoantigens recognized after epididymal and vasal obstructions may reflect maturational changes in sperm components that take place during the passage of spermatozoa through the epididymis.
Animals↗