Discordant and anomalous results among cytotoxicity assays: the confounding properties of eosinophil granule major basic protein on cell viability assays.
J Immunol Methods
; 227(1-2): 1-15, 1999 Jul 30.
Article
em En
| MEDLINE
| ID: mdl-10485250
ABSTRACT
When five cytotoxicity methods compared the toxicity of eosinophil granule major basic protein (MBP) and melittin to K562 and HL-60 cells, strikingly discrepant results were noted. Trypan blue staining, propidium iodide/CellTrackerGreen staining and incorporation of 14C-leucine assays indicated MBP damages > 75% of cells by 1 h. In contrast, 51Cr and lactic dehydrogenase (LDH) release assays indicated MBP damages most cells only at 20 h. All methods indicated melittin damages nearly all cells by 1 h. Further studies showed that without cell transfer, dye staining methods indicated MBP produces < 10% cytotoxicity after 4 h. A modified 14C-leucine assay, employing sodium dodecyl sulfate solubilization and trichloroacetic acid precipitation, showed lower cytotoxicity, 48%, at 4 h. Modified 51Cr and LDH assays showed increased cytotoxicities at 4 h, 34% and 58%, respectively. Overall, MBP's ability to cause molecular and cellular adhesion systematically confounds standard cytotoxicity measurements. However, the modified 14C-leucine assay provides a valid measure of MBP's cytotoxicity and may be useful for analyses of 'sticky' cytotoxins.
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Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Ribonucleases
/
Testes Imunológicos de Citotoxicidade
/
Proteínas Sanguíneas
/
Eosinófilos
Limite:
Humans
Idioma:
En
Ano de publicação:
1999
Tipo de documento:
Article