ABSTRACT
The use of a newly developed monoclonal antibody against Bromodeoxyuridine is demonstrated in a study with V79 Chinese hamster cells. It can replace radioactive material in cell cycle analysis studies in the way that BrdU incorporated into the DNA during S-phase is detected by immunofluorescence. By this technique the proportion of actively synthesizing S-phase cells can be determined. The proportion of such cells is a good measure with regard to the quality of a culture and/or medium in question. The method is fast and offers within 3-5 hours the possibility to analyse growth conditions in monolayers and suspension cultures.
Subject(s)
Antibodies, Monoclonal , Bromodeoxyuridine/immunology , Cell Cycle , Interphase , Animals , Cell Line , Cells, Cultured , Cricetinae , Fluorescent Antibody TechniqueABSTRACT
This presentation reports on the routine use of flow cytometric techniques in suspension cultures. As a model system an antibody secreting mouse hybridoma cell line was used to monitor antibody production (a Bromodeoxyuridine specific antibody) and proliferative status as obtained by flow cytometry (FCM) and cell counting.
Subject(s)
Cell Cycle , Hybridomas/cytology , Animals , Antibodies, Monoclonal/biosynthesis , Flow Cytometry , MiceABSTRACT
We have utilized monoclonal antibody against BrdUrd to detect sister-chromatid exchanges in CHO cells. This technique allows detection of SCEs at very low levels of BrdUrd incorporation. At incorporation level of 0.5%, a frequency of about 2 SCEs/cell/cycle was found. In a UV-sensitive mutant (43-3B) which has an increased spontaneous frequency of SCEs, it is found that this increase is due to incorporated BrdUrd. In MMS- and MMC-treated cells, an influence of BrdUrd on the frequencies of induced SCEs was found only when high concentrations of mutagens were employed.
Subject(s)
Bromodeoxyuridine/pharmacology , Sister Chromatid Exchange/drug effects , Animals , Antibodies, Monoclonal , Bromodeoxyuridine/immunology , Cell Line , Cricetinae , Dose-Response Relationship, Drug , Methyl Methanesulfonate/pharmacology , Mitomycin , Mitomycins/pharmacologyABSTRACT
Evaluation of the proliferative activities of cell populations has mainly been restricted to the use of autoradiography and flow cytometric measurements. The introduction of a new BrdUrd specific antibody makes it possible to determine exactly the DNA synthesizing cells. The BrdUrd technique is safe with respect to handling and the results are obtained within five hours. The suitability of the BrdUrd labelling procedure has been studied in different cell lines and compared with 3H-thymidine autoradiography and flow cytometry.
