Cell-based quantitative evaluation of the MTT assay.
OBJECTIVE
To analyze the bioreduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) on a per cell basis and evaluate its modulation as a function of different stages of cell metabolism.


METHODS
Following MTT bioreduction, total optical density (TOD), cell area and specific activity (TOD/area) of V79 cells and cultured macrophages were recorded for individual cells by means of digital image analysis. The effect of different serum (0-10% vol/vol) or genistein (0-100 microM) concentrations was used to modulate the MTT-specific activity response.


RESULTS
As cells in culture are heterogeneous in cell size, the contribution of each cell to the total amount of formazan formed per dish is variable. The production of formazan per cell as a result of MTT bioreduction was found to be proportional to cell size.


CONCLUSIONS
Specific MTT-reducing activity was analyzed in phagocytes and nonphagocyte cells, revealing the utility of this variable in evaluating the MTT assay at the single-cell level.
Molinari
Beatriz L
BL
National Research Council and Radiobiology Department, National Laboratories of Research and Service-Conicet, National Atomic Energy Commission, Avda. Gral. Paz 1499, (1650) San Martín, Pcia Buenos Aires, Argentina. beamolin@cnea.gov.ar
Tasat
Débora R
DR
Palmieri
Mónica A
MA
O'Connor
Silvia E
SE
Cabrini
Rómulo L
RL
D000818:Animals; D004396:Coloring Agents; D003239:Connective Tissue Cells / Q000166:cytology* / Q000378:metabolism* / Q000648:ultrastructure; D006224:Cricetinae; D003581:Cytodiagnosis / Q000379:methods; D003584:Cytological Techniques / Q000379:methods; D003952:Diagnostic Imaging / Q000379:methods; D011992:Endosomes / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D007091:Image Processing, Computer-Assisted / Q000379:methods*; D016676:Macrophages, Alveolar / Q000166:cytology / Q000378:metabolism / Q000648:ultrastructure; D051379:Mice; D008815:Mice, Inbred Strains; D018613:Microscopy, Confocal; D010084:Oxidation-Reduction; D013778:Tetrazolium Salts / Q000378:metabolism* / Q000493:pharmacokinetics; D013844:Thiazoles / Q000097:blood / Q000378:metabolism* / Q000493:pharmacokinetics
D004396:Coloring Agents; D013778:Tetrazolium Salts; D013844:Thiazoles; C022616:thiazolyl blue