DETECTION OF B LYMPHOMA CELLS UNDERGOING APOPTOSIS BY ANNEXIN-V ASSAY[J]. Chinese medical sciences journal, 2002, 0(1): 17-21.
DOI:
DETECTION OF B LYMPHOMA CELLS UNDERGOING APOPTOSIS BY ANNEXIN-V ASSAY[J]. Chinese medical sciences journal, 2002, 0(1): 17-21.DOI:
DETECTION OF B LYMPHOMA CELLS UNDERGOING APOPTOSIS BY ANNEXIN-V ASSAY
摘要
Abstract
To quantitatively analyze apoptotic and secondary necrotic cells unde r apoptosis conditions. Methods. The cells of Burkitt lymphoma (BL) cell line Raji were incubated with 1 .0 μmol/L dexamethasone (DEX) for 2
4 and 8 h respectively
then stained with Annexin V FITC (fluorescein isothiocyanate conjugated) which was used to detec t the exposed phosphatidylserine (PS) on the epimembrane resulting from a loss o f phospholipid asymmetry in the early stage of apoptosis
and also stained with propidium iodide (PI) which allows analysis of secondary necrotic cells related with cell membrane and DNA damage that probably represent late stage of apoptosi s
then apoptotic cells were quantified by flow cytometry (FCM). Furthermore
An nexin+/PI and Annexin+/PI+ cells were sorted by fluoresence activated cell sorter (FACS)
and identified by electron microscopy (EM) and DNA gel electroph oresis. Results. The percentage of apoptotic cells was found to increase with the incuba tion time (r=0.97). This method was sensitive with low detection limit (0.02%)
and was reproducible with low coefficient variance (CV)(4.2%). Meanwhile
the Annexin+/PI and Annexin+/PI+ cells were identified as apoptotic and necroti c cells under EM
and DNA extracted from the Annexin+/PI cells was characteri zed by “ladder pattern”. Conclusions. Annexin V assay is a specific
sensitive
accurate
reproductive and quantitative method for analyzing apoptotic cells.