Isolation of apoptotic mouse fetal oocytes by AnnexinV assay

Expression of phosphotidylserine by fetal oocytes in culture renders significant numbersof such cells able to bind AnnexinV-coated microbeads and allows their separation from Annexin V-negative oocytes on a Magnetic Cell Separation (MACS) column in a magnetic field. The majority of oocytes (>= 75%) which bound Annexin V-coated microbeads were viable, as indicated by their propidiurn iodine (PI) negativity. However, they showed apoptotic morphologies and were found to be TUNEL-positive. On the other hand, AnnexinV- negative oocytes, besides being PI negative, appeared morphologically healthy and TUNEL negative. Moreover, AnnexinV-positive oocytes showed a marked lower ratio of Bcl-xL/Bax transcripts in comparison to AnnexinV-negative oocytes. We conclude that the present method is able to separate fetal oocytes in two distinct populations: AnnexinV-positive oocytes showing features typical of apoptotic cells and AnnexinV-negative oocytes comprising for the most part viable non-apoptotic cells. This procedure should greatly facilitate studies aimed to identify the currently poorly understood molecular pathways governing apoptosis in mammalian fetal oocytes.