Immunological Studies on Dermatophytes IV. Chemical Structures and Serological Reactivities of Polysaccharides from Microsporum praecox, Trichophyton ferrugineum, Trichophyton sabouraudii, and Trichophyton tonsurans

The chemical structures and serological specificities of polysaccharides isolated from four species of dermatophytes, Microsporum praecox, Trichophyton ferrugineum, T. sabouraudii, and T. tonsurans, were investigated. Each of these species yielded a mixture of crude polysaccharides which could be separated into three water-soluble, neutral polysaccharides free of nitrogen. These were grouped as galactomannan I, galactomannan II, and glucan. The galactomannans I were quite similar in chemical structure. When measured by complement fixation, their serological cross-reactivities were similar with rabbit antisera to each of these species except T. sabouraudii. The differences in their relative reactivities with this antiserum could be correlated with differences in structure and specificity of this antiserum for galactofuranose end groups. The galactomannans II differed both in chemical structure and in their serological reactivities with antisera to each of these species. The galactomannan II from T. ferrugineum differed most in chemical structure and was the least reactive serologically. The glucans also differed in both structure and serological reactivities.     

Ribonucleic acid synthesis by Escherichia coli C3000/L after infection by the ribonucleic acid coliphage ZIK/1, and properties of the coliphage-induced double-stranded ribonucleic acid

1. The efficiency of extracting nucleic acids from Escherichia coli after five methods of obtaining cell lysis was determined. 2. The recovery of various nucleic acid species isolated after chromatography on methylated albumin-coated kieselguhr was also examined. 3. Double-stranded coliphage-induced RNA was isolated from infected bacteria and its resistance to ribonuclease digestion under various conditions determined. 4. The involvement of double-stranded RNA during the infection process was demonstrated. 5. The time-course of the syntheses in infected cells of double-stranded RNA, DNA, single-stranded coliphage and 16s ribosomal RNA, transfer RNA and ribosomal 23s RNA was examined. 6. It was demonstrated that the syntheses of DNA, transfer RNA and ribosomal RNA decreased 10-15min. after infection. 7. Synthesis of coliphage RNA commenced 10-15min. after infection and double-stranded RNA was also synthesized from about 10min. after coliphage adsorption.     

Monoclonal antibody against a melanosomal protein in melanotic and amelanotic human melanoma cells

BALB/c mice were immunized with tyrosinase, partially purified in two stages from a human melanoma cell line. A hybridoma was obtained which produced monoclonal antibody (MoAb 1C11) reactive with 8/10 melanoma cell lines and 10/10 primary cultures of human melanocytes, neval cells, and melanomas. Immunoreactivity correlated to a certain extent with tyrosinase activity but not with melanin content. No crossreactivity was obtained with neuroblastoma, medulloblastoma, fibroblasts, keratinocytes, lymphoid cells, or murine melanomas. Purification of the antigen directly from cell lysates with a MoAb 1C11 CNBr-Sepharose affinity column gave a green-brown protein of 56 kDa with no detectable tyrosinase activity. This protein was therefore different from 60 kDa active tyrosinase, identified by enzyme activity and Western blotting with a MoAb derived previously (MoAb 5C12). Unlike 5C12, 1C11 reactivity was not destroyed by pretreatment of the antigen with periodate. Immunogold labelling showed that the 1C11-reactive antigen was associated with melanosomes, and there was close correlation between 5C12 and 1C11 reactivity in resistance to trypsin and in staining various melanocytic cell populations. MoAb 1C11 may therefore recognise a polypeptide epitope in a molecule closely linked to melanin biosynthesis.     

Differential distribution of antigen-specific helper T cells and cytotoxic T cells after antigenic stimulation in vivo. A functional study using limiting dilution analysis

We have developed modified limiting dilution analysis (LDA) techniques that distinguish in vivo Ag-stimulated murine helper T lymphocytes (HTL) and CTL from unstimulated precursor T cells, even those with the same Ag specificity. We refer to these cells that are detectable in the modified LDA as "Ag-conditioned" T cells (cHTL and cCTL). We have used the modified LDA techniques in conjunction with conventional LDA techniques (which enumerate all Ag-specific T cells) to evaluate the in vivo distribution of Ag-conditioned cHTL and cCTL following in vivo sensitization to alloantigens via sponge matrix or skin allografts. In general, we observed the following regarding the distribution of cHTL and cCTL: 1) Ag-conditioned HTL and CTL were detectable only after in vivo sensitization with alloantigen: 2) not all Ag-reactive T cells became conditioned T cells after in vivo Ag deposition; 3) the percentage of Ag-reactive T cells that converted to conditioned T cells after Ag deposition varied among different lymphoid compartments; 4) a high percentage of cHTL, but a low percentage of cCTL, accumulated in regional lymph nodes and spleen; 5) cHTL accumulated in peripheral blood, whereas cCTL did not; 6) Ag-conditioned cHTL were detectable in various lymphoid tissues for greater than 60 days following Ag deposition, whereas cCTL were detectable for only 14 to 20 days; and 7) unlike the other lymphoid sites, the site of Ag deposition accumulated a high percentage of both Ag-stimulated cHTL and cCTL. Furthermore, cHTL and cCTL appeared to reside in phenotypically distinct T cell subsets in that in vivo treatment with anti-L3T4 mAb abrogated the accumulation of HTL, but not CTL, at the site of Ag deposition. These data demonstrate differential compartmentalization of Ag-conditioned cHTL and cCTL subsequent to in vivo Ag deposition. The implications of these findings regarding the monitoring of in vivo immune responses are discussed.