Simultaneous in situ detection of apoptosis and necrosis in monolayer cultures by TUNEL and trypan blue staining.
A method for simultaneously detecting membrane permeability (characteristic of necrosis) and DNA fragmentation (characteristic of apoptosis) is described. By combining a common dye-exclusion method (Trypan Blue) with a commercially available terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end-labeling (TUNEL) labeling kit, we have succeeded in developing a novel methodology for obtaining permanently mounted slides of monolayer cell cultures double-labeled for DNA fragmentation and cell lysis. This method should facilitate in situ studies of cell death by allowing for a more accurate quantification of total toxicity in monolayer cell cultures and perhaps further enhance our understanding of the different mechanisms of cell death as well.
Perry
S W
SW
University of Rochester Medical Center, NY, USA.
Epstein
L G
LG
Gelbard
H A
HA
D017209:Apoptosis; D002463:Cell Membrane Permeability / Q000502:physiology; D002470:Cell Survival; D002478:Cells, Cultured; D004396:Coloring Agents; D053938:DNA Fragmentation; D004253:DNA Nucleotidylexotransferase / Q000378:metabolism*; D003856:Deoxyuracil Nucleotides / Q000378:metabolism*; D006801:Humans; D009336:Necrosis; D009474:Neurons / Q000166:cytology; D011933:Reagent Kits, Diagnostic; D016707:Tissue Fixation; D014343:Trypan Blue
D004396:Coloring Agents; D003856:Deoxyuracil Nucleotides; D011933:Reagent Kits, Diagnostic; C027078:deoxyuridine triphosphate; D004253:DNA Nucleotidylexotransferase; D014343:Trypan Blue