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Microscale autoradiographic method for the qualitative and quantitative analysis of apoptotic DNA fragmentation
Author(s) -
Tilly Jonathan L.,
Hsueh Aaron J. W.
Publication year - 1993
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/jcp.1041540310
Subject(s) - microscale chemistry , fragmentation (computing) , dna fragmentation , apoptosis , qualitative analysis , dna , computational biology , microbiology and biotechnology , quantitative analysis (chemistry) , chemistry , biology , biochemistry , programmed cell death , chromatography , qualitative research , mathematics , sociology , ecology , social science , mathematics education
A method combining the advantages of electrophoretic DNA fractionation and autoradiography is described for the qualitative and quantitative analysis of internucleosomal DNA fragmentation that occurs during apoptosis, or “programmed cell death”. This procedure utilizes terminal transferase enzyme to uniformly add one molecule of [α 32 P]‐to the 3′‐of DNA fragments. Following gel electrophoresis and autoradiographic analysis, the total amount of radiolabel incorporated into the low molecular weight DNA fraction can be quantitated and used to estimate the degree of apoptotic DNA fragmentation in any given sample. This method requires as little as 15 ng of total cellular DNA and increases the sensitivity of apoptotic DNA detection by at least 100‐fold over the widely used ethidium bromide staining method. The procedure should prove valuable for the analysis of apoptosis in minute quantities of tissues and cultured cells. © 1993 Wiley‐Liss, Inc.