Measurement of endogenous and TNF alpha-mediated H2O2 production in supernatants of SK-N-SH neuroblastoma cells with an enhanced chemiluminescence assay.
A sensitive peroxidase-dependent luminol-enhanced chemiluminescence (ECL) assay for determination of hydrogen peroxide (H2O2) generation by tumour cells was established. This test system allows determination of H2O2 in concentrations as low as 25 pmol (50 nmol/L) and yields results which are comparable to those obtained using a less sensitive photometric method and a previously described scopoletin fluorescence assay. After 3 h incubation time 10(4) SK-N-SH neuroblastoma cells released 60 +/- 5 pmol H2O2 in the supernatant and this level was significantly (p < 0.025) increased by about 70% in the presence of 5 pmol (100 ng/mL) recombinant tumour necrosis factor alpha (TNF alpha). In contrast, H2O2 production was slightly reduced by TNF alpha at a very low concentration of 0.5 fmol (0.01 ng/mL).
Carmine
T C
TC
Children's University Hospital Tübingen, Germany.
Bruchelt
G
G
Hahn
T
T
Niethammer
D
D
D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D006801:Humans; D006861:Hydrogen Peroxide / Q000032:analysis / Q000378:metabolism*; D007202:Indicators and Reagents; D007700:Kinetics; D008163:Luminescent Measurements; D008545:Melanoma; D008832:Microchemistry; D009447:Neuroblastoma / Q000378:metabolism*; D012680:Sensitivity and Specificity; D014407:Tumor Cells, Cultured; D014409:Tumor Necrosis Factor-alpha / Q000494:pharmacology*
D007202:Indicators and Reagents; D014409:Tumor Necrosis Factor-alpha; D006861:Hydrogen Peroxide