Properties of a panel of monoclonal antibodies which react with the human T cell antigen receptor on the leukemic line HPB-ALL and a subset of normal peripheral blood T lymphocytes

Five Mab raised against the T cell antigen receptor of the human T cell line HPB-ALL which react with a subpopulation of normal peripheral blood T cells are described. Three Mab, 3D6, 1C1, and 1C2, react with 3 to 5% of normal PBL and stimulate proliferation of the cells with which they react. An increase in the number of cells which react with all five Mab occurs. Two Mab, 2D4 and 65, react with subsets of the cells which bind 1C1, 1C2, and 3D6 and divide the family into four subgroups, 2D4+ 65+, 2D4+ 65-, 2D4- 65+, and 2D4- 65-. Functional T cell clones in all four subfamilies have been observed. Cytolytic function can be correlated with the TcR phenotype expressed because all of the Mab which react with a particular clone inhibit its ability to lyse a specific target. The epitopes recognized by the panel are closely related because all five block each other's binding to HPB-ALL. In addition, the determinants recognized by 3D6, 1C1, and 1C2 on normal lymphocytes are probably very closely related because all clones examined react with all three Mab.     

Differential amplification and disproportionate expression of five genes in three multidrug-resistant Chinese hamster lung cell lines.

At least five linked genes are amplified in the multidrug-resistant Chinese hamster ovary cell line CHRC5, selected with colchicine (A. M. Van der Bliek, T. Van der Velde-Koerts, V. Ling, and P. Borst, Mol. Cell. Biol. 6:1671-1678, 1986). We report here that only a subset of these, encoding the 170-kilodalton P-glycoprotein, are consistently amplified in three different multidrug-resistant Chinese hamster lung cell lines, selected with vincristine, daunorubicin, or actinomycin D. Within each cell line, genomic sequences homologous to the P-glycoprotein cDNA probe were amplified to different levels. The pattern of differential amplification was consistent with the presence of at least two and possibly three P-glycoprotein genes. In the actinomycin D-selected cell line, these genes were disproportionately overexpressed relative to the associated levels of amplification. These results underline a central role for P-glycoprotein in multidrug resistance. In the daunorubicin-selected cell line, another, as yet uncharacterized, gene was amplified but disproportionately underexpressed. Its amplification was therefore fortuitous. We present a tentative map of the region in the hamster genome that is amplified in the multidrug-resistant cell lines which were analyzed.     

Comparison of various atmospheric conditions for isolation and subcultivation of Mycoplasma hyorhinis from cell cultures

The efficiency of aerobic incubation was compared with incubation under various oxygen and carbon dioxide conditions for the isolation and subcultivation of three strains of Mycoplasma hyorhinis from VERO-cell cultures and subcultivation of three laboratory strains. Under anaerobic conditions with a low oxidation-reduction potential (at or below -115 mV) as obtained in jars, with catalysts, containing mixtures of 5%-10% CO2 in H2, very poor or no growth of any of the six M. hyorhinis strains was observed. When traces of oxygen were present (that is, under conditions with higher oxidation-reduction potentials, e.g. when omitting the catalyst in the above gas mixtures or in 5% CO2 + 95% N2) isolation from cell cultures was successful in most tests, but subcultivation of these primary isolates was seldom possible under these semi-anaerobic conditions. However, in most cases these primary isolates could be subcultivated aerobically, although aerobic conditions were unsatisfactory for isolation in about half of the experiments. Isolation of M. hyorhinis was optimal in 5% O2 + 95% N2, under which condition the isolates could also always be subcultivated. Isolation failed occasionally when 5% O2 + 5% CO2 + 90% N2 was used, thus indicating that 5% CO2 was slightly inhibitory. 5% CO2 in air and 10% CO2 either in air, H2 or N2 were also inadequate for isolation from cell cultures. In contrast to the findings with these cell culture-adapted M. hyorhinis strains, the laboratory strains could be subcultivated easily under all conditions tested except those with an oxidation-reduction potential at or below -115 mV; 100% CO2 was inhibitory for all 6 strains. Our findings may partly explain why M. hyorhinis is often considered "non-cultivable" on artificial media once adapted to cell cultures. The findings emphasize the need to employ also a micro-aerophilic condition (5% O2 in 95% N2) in the examination of cell cultures for mycoplasma.     

Telomere conversion in trypanosomes.

Activation of the gene coding for variant surface glycoprotein (VSG) 118 in Trypanosoma brucei proceeds via a duplicative transposition to a telomeric expression site. The resulting active expression-linked extra copy (ELC) is usually flanked by DNA that lacks sites for most restriction enzymes and that is thought to interfere with the cloning of the ELC as recombinant DNA in Escherichia coli. We have circumvented this problem by cloning an aberrant 118 ELC gene, flanked at the 3'-side by at least 1 kb DNA, that contains restriction enzyme sites. Our analysis shows that this DNA and the 3'-end of the 118 ELC gene are derived from another VSG gene (1.1006) that is permanently located at a telomeric position. We propose that the 3'-end of the 1.1006 gene and (all of) its 3' flanking sequence moved to the expression site by a telomere conversion. Such a telomere conversion can also account for the appearance of an extra copy of the 1.1006 gene detected in a sub-population of our trypanosome strain.     

Size fractionation of Trypanosoma brucei DNA: localization of the 177-bp repeat satellite DNA and a variant surface glycoprotein gene in a mini-chromosomal DNA fraction

We have size-fractionated intact DNA from Trypanosoma brucei into a major large DNA fraction (greater than 350S) and minor middle-sized (60-250S) and small (less than 60S) DNA fractions. Large DNA contains the rRNA genes, the basic copy genes for several variant surface glycoproteins (VSGs), including one which lies near a telomer, and the expression-linked copies of the two VSG genes. The middle-sized DNA contains at least one VSG gene, but the hybridization of this fraction with probes for the conserved repetitive sequences that mark the edges of the transposed segments of VSG genes, suggests that it may contain many VSG genes. The 177-bp repeat satellite DNA is also exclusively found in this fraction.     

Hemagglutination Inhibition with Arboviruses: Relationship Between Titers and Source of Erythrocytes

Antigens for Grand Arbaud, Hazara, and California arboviruses were able to agglutinate goose and either dog, hamster and guinea pig, or hamster red blood cells (RBC) to the same titer at the same pH; in hemagglutination-inhibition (HI) tests, titers for homologous and related sera were the same with these different types of RBC or occasionally one dilution higher with the mammalian cells. Antigens for St. Louis encephalitis and Eastern equine encephalitis viruses required use of lower antigen dilutions with human, guinea pig, and hamster RBC than with goose RBC. The results of comparative HI testing with these latter antigens and types of RBC indicate that HI titer is not directly related to the antigen dilution used with different types of RBC.