Die Zelle: leben, fressen, sterben

A tetrazolium salt has been used to develop a quantitative colorimetric assay for mammalian cell survival and proliferation. The assay detects living, but not dead cells and the signal generated is dependent on the degree of activation of the cells. This method can therefore be used to measure cytotoxicity, proliferation or activation. The results can be read on a multiwell scanning spectrophotometer (ELISA reader) and show a high degree of precision. No washing steps are used in the assay. The main advantages of the colorimetric assay are its rapidity and precision, and the lack of any radioisotope. We have used the assay to measure proliferative lymphokines, mitogen stimulations and complement-mediated lysis.

Apoptosis is a programmed, physiological mode of cell death that plays an important role in tissue homeostasis. Understanding of the basic mechanisms that underlie apoptosis will point to potentially new targets of therapeutic treatment of diseases that show an imbalance between cell proliferation and cell loss. In order to conduct such research, techniques and tools to reliably identify and enumerate death by apoptosis are essential. This review focuses on a novel technique to detect apoptosis by targeting for the loss of phospholipid asymmetry of the plasma membrane. It was recently shown that loss of plasma membrane asymmetry is an early event in apoptosis, independent of the cell type, resulting in the exposure of phosphatidylserine (PS) residues at the outer plasma membrane leaflet. Annexin V was shown to interact strongly and specifically with PS and can be used to detect apoptosis by targeting for the loss of plasma membrane asymmetry. Labeled annexin V can be applied both in flow cytometry and in light microscopy in both vital and fixed material by using appropriate protocols. The annexin V method is an extension to the current available methods. This review describes the basic mechanisms underlying the loss of membrane asymmetry during apoptosis and discusses the novel annexin V-binding assay.