Various methods of apoptosis detection

Apoptosis is cell death defined by some ultrastructural characteristics. DN A agarose gel electrophoresis is suitable for cultured cells consisting of homogeneous cells in which apoptosis is relatively easy to induce using app ropriate stimuli, but often fails to detect a typical DNA ladder when tissu es consisting of heterogeneous cells are used and contain only a few apopto tic cells. It is known that the terminal deoxynucleotidyl transferase (TdT) -mediated dUTP-biotin nick end-labeling (TUNEL) method detects both apoptot ic and necrotic cells, although TUNEL can detect also newly yielded free 3' -OH ends of DNA. Fluorescence dyes specifically bind with DNA, clearly show ing fragmented nuclei. Annexin V enables classification of the apoptotic ce lls into different stages, because it can detect the externalization of pho sphatidylserine in the cell membrane which occurs at the early stage of apo ptosis. The disadvantage of fluorescence dyes and annexin V is to be applic able only to unfixed materials. Western blot analysis has several advantage s such as its applicability to both cells and tissues, and semiquantificati on of a protein expressed in materials used, but is unsuitable for analysis of the topographic distribution of cells producing apoptosis-related prote in such as the caspase family. As mentioned above, most of the apoptosis de tection methods focus only on one of the apoptotic characteristics, thereby limiting their application to apoptosis detection. Therefore, it is requir ed to combine several methods for the precise detection of apoptosis.