![]() ![]() These results suggest that long-lasting ICAM-1 message upregulation in response to cytokines or PMA may be due to transcriptional upregulation in the early phase and stabilization of ICAM-1 message in the later phase (after 4 h). ICAM-1 message induction-blocking studies suggested that primary upregulation of ICAM-1 mRNA may be caused by transcriptional upregulation. In order to investigate whether increased gene transcription contributes to ICAM-1 upregulation, RCC cells were treated with TNFα, IFNγ, or PMA with or without simultaneous addition of actinomycin D. ICAM-1 mRNA stability assays showed that both unstimulated and stimulated RCC cells had very stable ICAM-1 mRNA up to 24 h. Although ICAM-1 mRNA in NKPT cells was upregulated by these cytokines, their messages returned to basal levels after 24 h. Time course studies showed that ICAM-1 mRNA was upregulated by INFγ, TNFα, and PMA, plateaued after 2 h, and remained increased for up to 24 h. The kinetics of ICAM-1 message induction was studied by Northern analysis of total RNA extracted from RCC and normal kidney proximal tubular (NKPT) cells. Of the various cytokines, tumor necrosis factor αa (TNFα), interferon-γ (IFNγ), and phorbol myristate acetate (PMA) strongly upregulated ICAM-1 protein expression on RCC. #Qlab quintiles Activator#We report an investigation on ICAM-1 expression and molecular regulation by cytokines and protein kinase C activator on RCC cell lines. Since ICAM-1 is expressed by most renal cell carcinomas (RCC), the regulation of ICAM-1 expression is important in understanding the biological behavior of RCC. aspects of tumor biology, namely enhancement of tumor metastasis and mediation of host defense mechanisms such as lymphocyte-mediated tumor cytotoxicity. Intercellular adhesion molecule-1 (ICAM-1) mediates two important functional. However, the cocktails employed in this study enabled several different applications to be run and established that multicolor reagent mixtures containing V450–antibody conjugates are functional and stable. Such cocktails can frequently be unstable due to the tendency of one or more components to lose structural integrity, photobleach, or develop unwanted affinities for another component. Additional studies were performed, with BD Horizon V450–antibody conjugates being included in eight-color cocktails aimed at subsetting lymphocytes and myeloid cells. A monochlorinated hydroxycoumarin was found to have a high quantum yield (∼0.98), and human leucocyte-specific monoclonal antibodies (CD3, CD4, and CD45) conjugated with this dye exhibited reliable performance in flow cytometry assays. In our search for new violet-excitable dyes with improved photophysical and photochemical properties, we examined several halogen-substituted hydroxycoumarins and found that chlorinated derivatives are at least as bright as their fluorinated analogs. We conclude that there is some flexibility in sample processing methods for quantitative CD38 determination however, it is preferable for a laboratory to employ one method of fluorescence quantitation calculation consistently because small differences are detected between different methods. In many cases, however, these differences are relatively small and were more pronounced in certain laboratories. However, for all sample processing methods above, the CD4 biologic calibrator and QuantiBRITE bead methods gave significantly different estimates of CD38 intensity. The only significant difference in fluorescent intensity and CD38 antibodies bound per cell (ABC) was encountered when whole blood was held for 24 h prior to staining and fixation and then acquired after another 24-h hold. ![]() Neither lysing reagent (ammonium chloride versus BD FACSlyse), fixation (paraformaldehyde versus no final fixation step), nor acquisition delay (acquisition within 6 h after fixation versus 24 h after fixation) had a significant effect on CD38 relative fluorescent intensity or CD38 quantitative estimates (RFI or antibodies bound per cell). We evaluated the effect of specimen processing variations and quantitation methods on quantitative determination of CD38 expression on CD8 T lymphocytes. ![]()
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