Cytotoxic Drug Resistance Mechanisms by Robert Brown, Uta Böger-Brown PDF

By Robert Brown, Uta Böger-Brown

ISBN-10: 0896036030

ISBN-13: 9780896036031

Major medical and laboratory scientists describe state-of-the-art equipment for studying the mechanisms of mobile resistance to anticancer cytotoxics in human tumors. The protocols comprise distinct directions and wide troubleshooting assistance that permit researchers successfully to review a wide selection of drug resistance mechanisms, together with facets of drug-induced phone loss of life, drug uptake/efflux, drug metabolism, and DNA fix. every one approach is designed to aid establish the correlation among molecular and biochemical facts and the medical responses of the sufferer. Cytotoxic Drug Resistance Mechanisms illuminates all of the clinically correct mechanisms used as markers of the organic reaction to anticancer biotherapeutics this present day.

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0, 1 mM EDTA) for 30 minutes at 37°C (see Notes 2 and 3). 4. Wash in sterile distilled water three times, rinse in 70, 90, and 95% alcohol and air-dry. 5. Prepare 40–60 mL ISEL buffer for each section and keep on ice (see Notes 4, 5, and 7). 6. Carefully pipet the mixture over the tissue section, and place a cleaned glass coverslip over the section to prevent evaporation. 7. Incubate at 37°C for 1 h in a moist chamber. 8. Terminate the reaction by washing sections in distilled water three times, being careful not to scratch the tissue surface when removing the coverslip.

A sub-G1 peak is reliably found to be indicative of apoptosis (because of the DNA fragmentation) and the peak size reflects the amount of apoptosis in the sample. The method is easily applied to cell suspen- Apoptosis in Tissue Sections 47 sions and can also be applied to material derived from histological samples, although this is associated with higher ‘background levels’ and, of course, results in the loss of potentially important spatial and micro-anatomical information. The appearance of phosphatidylserine on the cell surface can also be measured by fluorescence activated cell sorter (FACS), through binding of fluorescein isothiocyanate (FITC)-tagged annexin V.

During cell death there is an alteration in the ability of cells to scatter light at a forward angle, reflecting cell size, and at 90°, reflecting cell granularity. These changes can be detected using flow cytometry. Such changes can be used to discriminate between live cells, dead cells, and cells that are undergoing apoptosis (10). As mentioned above apoptotic cells characteristically exhibit DNA strand breaks and these can be detected using terminal deoxynucleotidyl transferase (TdT)-mediated dUTP Nick End Labeling (TUNEL) and flow cytometry (8).

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Cytotoxic Drug Resistance Mechanisms by Robert Brown, Uta Böger-Brown

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