| Conventional apoptosis assays using propidium iodide generate a significant number of false positives that prevent accurate assessment of cell death. | |
| | |
MedLine Citation:
|
PMID: 20381494 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
|
The advent of flow cytometry-based applications has significantly impacted the study of cellular apoptosis. Propidium iodide (PI) is a commonly used viability stain in these studies. Unfortunately, we find that conventional Annexin V/PI protocols lead to a significant number of false positive events (up to 40%), which are associated with PI staining of RNA within the cytoplasmic compartment. Both primary cells and cell lines are affected, with large cells (nuclear: cytoplasmic ratios <0.5) showing the highest occurrence. This distribution spans a wide range of animal models including mice, swine, avian, and teleost fish and potentially affects up to 1016 out of 1019 of peer-reviewed papers published in this area since 1995. We show that the primary ramifications from these findings relate to cells experiencing changes in RNA content. Virally infected cells, for example, are qualified as undergoing apoptosis in response to infection based on conventional staining protocols; in fact, these cells are alive and actively producing viral RNA that can serve to produce additional infectious viral particles. Based on our observations we propose a modified protocol, show that it overcomes previous drawbacks for this technique, and that it will allow for more accurate assessment of cell death across various platforms. |
| | |
Authors:
|
Aja M Rieger; Brian E Hall; Le Thuong Luong; Luis M Schang; Daniel R Barreda |
Related Documents
:
|
15942714 - Apoptosis and inhibition of the phosphatidylinositol 3-kinase/akt signaling pathway in ... 9727944 - Induction of apoptosis in two mammalian cell lines results in increased levels of fruct... 12398804 - Cell cycle synchronization of porcine fetal fibroblasts by serum deprivation initiates ... 20661774 - Ouabain-induced apoptosis and rho kinase: a novel caspase-2 cleavage site and fragment ... 638144 - Potassium transport in dispersed mucosal cells from guinea pig stomach. 3688464 - Localization of calbindin-d28k in calcitonin containing cells of chick ultimobranchial ... |
Publication Detail:
|
Type: Journal Article; Research Support, Non-U.S. Gov't Date: 2010-04-07 |
Journal Detail:
|
Title: Journal of immunological methods Volume: 358 ISSN: 1872-7905 ISO Abbreviation: J. Immunol. Methods Publication Date: 2010 Jun |
Date Detail:
|
Created Date: 2010-05-31 Completed Date: 2010-07-09 Revised Date: - |
Medline Journal Info:
|
Nlm Unique ID: 1305440 Medline TA: J Immunol Methods Country: Netherlands |
Other Details:
|
Languages: eng Pagination: 81-92 Citation Subset: IM |
Copyright Information:
|
Copyright 2010 Elsevier B.V. All rights reserved. |
Affiliation:
|
Department of Biological Sciences, University of Alberta, Edmonton, Alberta, Canada. |
Export Citation:
|
APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
|
Animals Anthraquinones / metabolism Apoptosis* Cell Death Cell Line Cell Line, Tumor Cell Nucleus / metabolism Cell Survival Cells, Cultured Cercopithecus aethiops Chick Embryo Cytoplasm / metabolism DNA / metabolism False Positive Reactions Flow Cytometry Fluorescent Dyes / metabolism Formaldehyde / metabolism Goldfish Humans Mice Mice, Inbred C57BL Microscopy, Confocal Propidium / metabolism* RNA, Double-Stranded / genetics, metabolism Ribonucleases / metabolism Staining and Labeling / methods* Sus scrofa Tissue Fixation Vero Cells Virus Replication / genetics |
| Grant Support | |
ID/Acronym/Agency:
|
//Canadian Institutes of Health Research |
| Chemical | |
Reg. No./Substance:
|
0/1,5-bis((2-(methylamino)ethyl)amino)-4,8-dihydroxyanthracene-9,10-dione; 0/Anthraquinones; 0/Fluorescent Dyes; 0/RNA, Double-Stranded; 36015-30-2/Propidium; 50-00-0/Formaldehyde; 9007-49-2/DNA; EC 3.1.-/Ribonucleases |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
Previous Document: A unique esophageal motor pattern that involves longitudinal muscles is responsible for emptying in ...
Next Document: Development of an immunoassay for ciprofloxacin based on phage displayed antibody fragments.