Effect of postmortem interval on in situ end-labeling of DNA oligonucleosomes.
In situ end-labeling (ISEL) of DNA oligonucleosomes facilitates detection of apoptosis in tissue sections by binding labeled nucleotides to the oligonucleosomal fragments. Although ISEL is used in postmortem material, the effect of autolysis is not specifically known. Accordingly, normal rat brain and intestine were immersed in formalin after postmortem intervals (PMI) from 0 to 72 hours (h) or were fixed by perfusion with ethanol or paraformaldehyde-glutaraldehyde (PF-G). Omission of the binding enzymes or pretreatment with DNAase served as negative and positive controls. With DNA polymerase, material fixed by perfusion or by formalin immersion with PMI of O contained ISEL-positive nuclei only in apical intestinal cells and rare intravascular blood cells in brain. Postmortem intervals from 8 to 72 h did not alter this staining pattern although false-positive ISEL developed at section edges as a result of tissue drying. Terminal deoxynucleotidyl transferase gave similar results in the formalin-fixed material with PMI from 0-48 h but nonspecific nuclear labeling was increased with a PMI of 72 h and was widespread in the PF-G and ethanol-perfused material. This study shows that PMI of at least 72 h do not affect the sensitivity or the specificity of ISEL and confirm the reliability of this procedure in postmortem material. The results also indicate that false-positive ISEL occurs if the tissue is allowed to dry or if certain combinations of fixatives and binding enzymes are used.
Petito
C K
CK
Department of Pathology (Neuropathology), University of Miami School of Medicine, Florida, USA.
Roberts
B
B
D000818:Animals; D017209:Apoptosis; D001921:Brain / Q000473:pathology*; D015342:DNA Probes; D003851:Deoxyribonucleases; D005260:Female; D005557:Formaldehyde; D009841:Oligonucleotides; D051381:Rats; D011916:Rats, Inbred F344; D013997:Time Factors
D015342:DNA Probes; D009841:Oligonucleotides; D005557:Formaldehyde; D003851:Deoxyribonucleases