Immunohistochemical heterogeneity of alpha-tubulin in human epithelia revealed with monoclonal antibodies.

Four monoclonal antibodies raised to alpha subunit of pig brain tubulin (TU-01, TU-02, TU-03, TU-04) were used to study immunohistochemical heterogeneity of alpha tubulin in human epithelia. Selective reactivity was detected in the skin and trachea/bronchi, whereas all other epithelia investigated reacted uniformly with all four monoclonal antibodies. In the skin TU-01 reacted very strongly with all layers except the basal layer; TU-02 reacted strongly with granular layer and was unreactive or only weakly reactive with others; TU-03 reacted very strongly with basal layer and weakly to moderately with superficial layers; TU-04 reacted strongly with the granular layer of epidermis and was unreactive with other layers.

Cellular distribution of a protein related to neuronal microtubule-associated protein MAP-2 in Leydig cells.

A monoclonal antibody to neuronal microtubule-associated protein MAP-2 was produced. Immunoblotting of lysates of cultured cells revealed that the antibody, called MA-01, bound to a protein of Mr 210 kDa. Double immunofluorescence microscopy showed that the antibody stained microtubules.

Heterogeneity of microtubules recognized by monoclonal antibodies to alpha-tubulin.

A set of four monoclonal antibodies against tubulin (TU-01, TU-02, TU-03, and TU-04) were produced using pig brain microtubule protein as antigen. Their characterization shows that all recognize antigenic determinants located on the tubulin alpha-subunit. However, peptide mapping of isolated alpha-tubulin, followed by immunoblotting with the monoclonal antibodies, shows that the antigenic determinants are located on different peptide fragments in at least three cases.

A common antigenic determinant of vimentin and desmin defined by monoclonal antibody.

A monoclonal antibody, VI-01, to vimentin and desmin has been isolated from a fusion of mouse myeloma cells with immune spleen cells. The antibody appeared specific for vimentin of various species and smooth muscle desmin and did not recognize other intermediate filament proteins when cell lines and tissue sections were assayed in immunofluorescence and cell lysates in immunoblotting. The binding test with purified vimentin and desmin confirmed these results.

Antibodies against surface antigens common to spermatogenic and F9 teratocarcinoma cells in mice immunized with blood group substances from human saliva.

Significant inhibition of spermatogenesis and production of antibodies against membrane antigens of spermatogenic and F9 teratocarcinoma cells were observed in mice of the strain 129/Sv after immunization with human blood group substances from saliva of A, B or H secretors. Absorption of the mouse anti-ABH sera with appropriate human erythrocytes did not change their reactivity with testicular and F9 cells, whereas absorption with F9 cells eliminated the reactivity with both F9 and spermatogenic cells. This pattern of reactivity, together with higher binding of the anti-ABH sera to the cells expressing stage-specific embryonic antigen 1 (SSEA-1), suggests that these antisera contain antibodies against developmentally regulated carbohydrate antigens.

Developmentally regulated surface structures of teratocarcinoma stem cells studied by mutant cell lines.

Monoclonal antibodies TEC-01, TEC-02, and TEC-03, which define three developmentally regulated antigens TEC-1 (SSEA-1-like), TEC-2, and TEC-3, have been used to isolate and characterize teratocarcinoma stem cell mutants with altered expression of surface glycoconjugates. Mutants lacking TEC-1 antigen have been isolated by exposing mutagenized P19S1801A1 cells to TEC-01 antibody, which was conjugated to the toxin from Ricinus communis. None of the mutants exhibits significant changes in the expression of TEC-3 antigen, but some are defective in the expression of TEC-2 antigen.

Differentiation antigens of mouse teratocarcinoma stem cells defined by monoclonal antibodies.

Three differentiation antigens of mouse teratocarcinoma stem cells are defined using a panel of ten IgM-class monoclonal antibodies raised against teratocarcinoma F9 cells. TEC-01 and four other antibodies define an antigen that corresponds to SSEA-1. TEC-02 antibody defines an antigen that is expressed on teratocarcinoma stem cells, parietal yolk sac cells PYS-2, unfertilized eggs including the zona pellucida and blastocysts.

Localization of mouse TEC-1,2,3 antigens in adult and fetal tissues in the rat and human.

Cryostat sections of adult and fetal tissues of the rat and human were investigated by a double immunoperoxidase technique for the presence of three developmentally-regulated mouse antigens, TEC-1,2,3. In the adult rats the TEC-1 antigen was detected on the epithelium of the isthmic portion of the oviduct, in some kidney tubuli, and in the brain; the TEC-2 antigen was distributed on the epithelium of various tissues, but it was also present on cells that may correspond to phagocytizing macrophages; the TEC-3 antigen was detected on endothelia of some vessels or capillaries. In rat fetuses only the TEC-1 antigen was detected; it was found on digestive tract epithelium and some kidney tubuli.

Relationship of F9 antigens to spermatogenic cell antigens.

Mice immunized with teratocarcinoma F9 cells or human blood group substances A, B or H exhibited significant inhibition of spermatogenesis comparable to the inhibition induced by immunization with testicular cells. All these immunization schemes resulted in the production of antibodies which recognize antigens common to F9 and spermatogenic cells. In addition to these antigens, both anti-F9 and anti-ABH sera also recognize antigens which are specific for F9 cells.

Isolation and partial characterization of lectin-resistant F9 cells.

Cytotoxic plant lectins have been used for the single-step selection of mouse embryonal carcinoma cell mutants with altered expression of surface glycoconjugates. Following mutagenesis, several F9 and OTF9-63 cell lines resistant to the lectins from Triticum vulgaris or Ricinus communis were obtained. At least five distinct lectin-resistant (LecR) phenotypes have been identified on the basis of their relative sensitivities to four different plant lectins and their altered lectin-binding properties.

Cytotoxicity of plant lectins for mouse embryonal carcinoma cells.

The cytotoxicity of 10 plant lectins with different carbohydrate recognition properties towards a number of mouse embryonal carcinoma (EC) cell lines (F9, OTF9-63, PCC4, PCC3/A/1, P19, and P19S1801A1) has been examined. Six of the lectins are toxic for the majority of the cell types at concentrations of less than or equal to 100 micrograms/ml and should be useful as direct selective agents for the isolation of EC glycosylation mutants (see accompanying manuscript). However, the concentration of the various lectins required to kill 90% of the cell population differs markedly between EC cell lines, the greatest variation being observed with the lectins from T.

Induction of autoimmune damage to spermatogenesis in guinea pigs by passive transfer of anti-F9 mouse teratocarcinoma stem cell serum fraction.

The anti-F9 and anti-spermatogenic cell sera with known cytotoxic and immunofluorescent activities were produced in albino guinea pigs immunized with F9 or spermatogenic cells emulsified with Freund's complete adjuvant. Short-term administration of the globulin fractions (3 x 50 mg at 24-h intervals + 1 x FCA on the first day) obtained from these antisera to adult albino guinea pigs resulted in a significant inhibition of spermatogenesis and appearance in the sera of marked cytotoxic activity against F9 cells. These results together with the impairment of fertility in male and female mice immunized with teratocarcinoma stem cells described by other authors are interpreted as supporting the hypothesis that the F9 antigens may play an essential role in spermatogenesis and function of spermatozoa.

Immunological tolerance to spermatogenic cell antigens induced by teratocarcinoma stem cell antigens.

Repeated intraperitoneal administration of F9 teratocarcinoma stem cells (first dose of 3.8 X 10(8) + 3 doses of 7.5 X 10(8) cells at two-week intervals) to guinea pigs starting from birth prevented in more than one third of them (in 10 out of 28) the induction of autoimmune aspermatogenesis by subsequent immunization with spermatogenic cells emulsified with FCA.

Production and characterization of monoclonal antibodies against human transferrin.

Twenty-one hybridoma lines were established, producing monoclonal antibodies against human transferrin and growing well in the peritoneal cavity of the mouse. Eight of these monoclonal antibodies were characterized. All of them are of IgG1 subclass with K light chains.

Monoclonal antibodies against horseradish peroxidase.

Four hybridomas making monoclonal antibodies against horseradish peroxidase have been established. Two of them are IgM and two are IgG1 with kappa light chains. Antibody HP-03 has been selected as the most suitable one for preparation of the peroxidase-anti-peroxidase complex.

Cross-reactivity between tubulin and denatured human serum albumin demonstrated by monoclonal antibody TU-01.

Murine monoclonal antibody TU-01 specific for tubulin (Viklický et al., 1982, Cell. Biol.

Autoimmune damage to spermatogenesis in rodents immunized with mouse F9 embryonic carcinoma cells.

Significant inhibition of spermatogenesis and appearance of antibodies against spermatogenic cells identified by cytotoxicity and immunofluorescence reactions were observed in mice of inbred strains 129/Sv and BALB/c and in albino guinea pigs after syngeneic, allogeneic, and xenogeneic immunization with mouse F9 embryonic carcinoma cells and Freund's complete adjuvant. A similar syngeneic immunization with PYS-2 cells was ineffective. Appropriate absorption experiments confirmed the similarity between the antigens of F9 and spermatogenic cells and the absence of such a similarity with antigens of PYS-2 cells.

Monoclonal antibodies against transferrin. Precipitating mixtures and lack of inter-species cross-reactivity.

Five stable hybridoma lines were prepared using the myeloma cell line P3-X63-Ag.653 and spleen cells of mice hyperimmunized by pig transferrin. All hybridomas grew well in mouse peritoneal cavity and produced antibodies of the IgG1 subclass.

Biological activity of hormonally active and non-active androgen derivatives.

Androgen derivatives appeared to have different biological activities in vivo and in vitro. Testosterone-17-isobutyrate given in three doses of 50 or 200 microgram increased significantly the weight of seminal vesicles and reduced thymus weight in castrated males, whereas testosterone-17-hemisuccinate, testosterone-D-beta-glucoside and testosterone-3-(O-carboxymethyl)-oxime had no such effects. Similarly, ten 1-mg doses of testosterone-17-isobutyrate, unlike testosterone-17-hemisuccinate, resulted in a marked reduction of thymus weight in non-castrated males and in a significant inhibitory effect on the activity of spermatogenesis.

Mechanism of the inhibitory effect of hydrocortisone on concanavalin A-induced activation of thymus cells. Role of interleukins.

Concanavalin A-induced activation of thymus cells compared to spleen or lymph node activation is extremely sensitive to the inhibitory effect of hydrocortisone (HC). This inhibitory effect of thymus cells, as observed at concentrations of 10(-7) M and higher, can be abrogated (at concentrations of up to 10(-5) M HC) by the addition of culture supernatants containing the product of activated T cells--Interleukin 2 (IL2), but not the supernatants containing the product of activated macrophages--Interleukin 1 (IL1). The protective effect of the IL2-containing supernatants can in part be removed by absorption with T cell blasts, but not B cell blasts.

Monoclonal IgG3 (kappa) antibodies against murine Thy-1.2 antigen produced by murine hybridomas. Differences in the specificity of the antigen binding site and in the structure of the hinge region.

Two monoclonal IgG3 (kappa) antibodies against murine Thy-1.2 antigen produced by murine 1B5 and 1aG4/C5 hybridomas were partially characterized. The 1aG4/C5 antibody has slightly higher affinity for the Thy-1.

Localization of androgen receptors in mouse thymus.

Androgen receptors were found both in cytosol prepared from intact thymuses of the adult castrated B10. A male and in cytosol from thymuses of the castrated males that had been previously given whole-body irradiation with 6.0 Gy (60Co).

Identification and characterization of immature thymocytes responsive to T cell growth factor.

The surface antigenic phenotype of peanut lectin agglutinin (PNA)-separated, high-density ("immature") thymocytes which are the target for the T cell growth factor (TCGF) enabling this otherwise unresponsive cell population to mount a strong proliferative response to concanavalin A (Con A) has been studied. The results obtained indicate that (a) the major population of cells bearing the typical phenotype of immature thymocytes, i.e.