Biomedical subjects

C C Uphoff

Publications and source records attributed to C C Uphoff.

41 records · Page 3Linked to original sources

PubMed1993

Expression of the monocyte-specific esterase gene in leukemia-lymphoma cell lines.

The expression of the monocyte esterase was examined in a panel of 77 continuous human leukemia-lymphoma cell lines representing all hematopoietic cell lineages and in 16 other cell lines. Accumulation of mRNA, determined by Northern blotting with the cDNA probe HMSE-1, and production of the protein, shown by isoelectric focusing on polyacrylamide gels, correlated with differentiation of the cells along the monocytic lineage. None of the lymphoid, erythroid, megakaryoblastic or Hodgkin's disease derived cell lines or the non-hematopoietic human tumor cell lines expressed the full-length mRNA of 2.0 kb. These results support the notion that this enzyme, a serine hydrolase with still unknown physiological functions, is specifically expressed in cells committed to the monocyte/macrophage cell lineage.

Animals↗

PubMed1992

Mycoplasma contamination in human leukemia cell lines. I. Comparison of various detection methods.

The sensitivity and reliability of seven assays for mycoplasma detection were tested on a panel of leukemia cell lines. The assays used were: microbiological cultivation on broth and agar, immunofluorescent visualization of mycoplasmal DNA using DAPI (both direct staining and after multiplication of the contaminants on an indicator cell line), a nucleic acid hybridization assay with a radioactive probe specific for mycoplasmal rRNA, and ELISA with mycoplasma-specific polyclonal antibodies, a biochemical method utilizing 6-MPDR, and a mycoplasma-specific monoclonal antibody in immunofluorescence staining. The broth-agar method, the two DAPI tests and the RNA hybridization assay produced the highest detection rates; a number of false-negative cases were recorded by the other tests. The detection rates, costs, requirement for specialized equipment and other characteristics were evaluated for each method. Since each technique also has disadvantages and certain limitations and since no method can be regarded as the 'gold standard', at least two procedures should be used in routine screening for mycoplasma in cell cultures.

Antibodies, Monoclonal↗

PubMed1992

Mycoplasma contamination in human leukemia cell lines. II. Elimination with various antibiotics.

19 suspension cell lines were treated with antibiotics for elimination of chronic contamination with mycoplasma. We compared the efficiency, cytotoxicity and cross-resistance of the commercially available antibiotics MRA (Mycoplasma Removal Agent, a quinolone derivative and DNA gyrase inhibitor), Ciprobay (ciprofloxacin, also a quinolone derivative and DNA gyrase inhibitor), and BM-cyclin (a combination of tiamulin, a pleuromutilin derivative, and minocycline, a tetracycline derivative, both inhibitors of protein synthesis on ribosomes). Contaminants were eliminated in all 19 cell lines by BM-Cyclin. Only 74% of the cell lines were cleared of contamination by both MRA and Ciprobay. Successful treatment was monitored by three mycoplasma detection assays. Cross-resistance was noted between MRA and Ciprobay in four of the five cell lines not cleared by either reagent. This resistance could, however, be overcome by consecutive exposure to BM-cyclin. Employed at the recommended concentrations, the antibiotics did not cause marked cytotoxicity, but the growth of the cells was affected to various degrees by some antibiotics. The elimination of mycoplasma from chronically contaminated cell lines is an effective alternative to other treatment protocols, but is cost-intensive and time-consuming; lasting damaging effects of the treatments on the eukaryotic cells cannot be excluded. Long-term post-treatment monitoring is mandatory, since contaminants may only be suppressed and then recur.

Anti-Bacterial Agents↗

PubMed1992

Treatment of mycoplasma-contaminated continuous cell lines with mycoplasma removal agent (MRA).

Thirty-nine continuous adherent or suspension cell lines were treated with a quinolone antibiotic, Mycoplasma Removal Agent (MRA), for the elimination of chronic mycoplasma contamination. In preliminary experiments MRA did not show any cytostatic or cytotoxic effects on mycoplasma-free cell cultures in concentrations up to ten-fold the concentration used for mycoplasma eradication. Twenty-eight cell lines (72%) were effectively cleansed of the mycoplasma contaminants by MRA treatment. The persistent removal of the mycoplasma infection was monitored by three mycoplasma detection assays. In seven cell lines (18%) the mycoplasmas were resistant to treatment with MRA. The resistant species was mainly M. arginini followed by M. orale and A. laidlawii; however, other cell lines harboring these species were cured. Four cell lines (10%) which prior to treatment presented with decreased viability and poor or no cell growth were lost during or shortly after the exposure to the antibiotic. If an antibiotic elimination is attempted it is imperative to closely examine the effectiveness of treatment and possible eukaryotic cytotoxicity. The treated mycoplasma-free cells may also no longer express the original features as a result of treatment or the absence of mycoplasma.

Cell Survival↗

PubMed1992

Sensitivity and specificity of five different mycoplasma detection assays.

The sensitivity and specificity of five different mycoplasma detection tests were evaluated in comparison with the classical microbiological culture assay on agar plates as the reference method: direct fluorochrome DNA staining (direct DAPI), DNA staining of an indicator cell line (indirect DAPI), RNA hybridization with a cDNA specific for ribosomal mycoplasmal RNA, an enzyme-linked immunosorbent assay (ELISA) with mycoplasma-specific antibodies, and a biochemical cytotoxicity assay (6-MPDR). A large panel of continuous cell lines (20 adherent and 233 suspension cell lines, most of the latter were human leukemia-lymphoma cell lines) were analyzed for infection with mycoplasma. The results of the comparative analysis for sensitivity and specificity of the various tests were as follows: 100% and 100% for the indirect DAPI, 100% and 98% for the RNA hybridization assay, 87% and 94% for the direct DAPI, 72% and 100% for the ELISA, 75% and 90% for the biochemical 6-MPDR assay. Each of these approaches has both advantages and disadvantages with regard to cost, time, reliability, specificity, and sensitivity. The best compromise for routine mycoplasma testing is a combination of several techniques (e.g. direct culture on agar, RNA hybridization, and direct or indirect DAPI).

Animals↗