Quantitative reduction of methyl tetrazolium by fresh vein homograft biopsies in vitro is an index of viability.
OBJECTIVE
The color density of the methyl tetrazolium (MTT) test is proportional to mitochondrial enzyme activity thus reflecting cellular viability. The aim was to evaluate the MTT test as a viability assay for vein homograft studies.


METHODS
Fresh intact vein samples were harvested during multi-organ procurement. The reliability of the MTT assay was tested by a fluorescent dye combination (1 microg/ml propidium iodide PI and 4 microM/ml SYTO-16 stains). The enzyme kinetics of the reaction was also investigated. The optimal reagent concentration, biopsy size and incubation period were established.


RESULTS
There was a linear relationship between the vein homograft's weight and the pigment production activity. A nonspecific reaction (8.6%) was observed in negative controls. The MTT cleavage up to 0.1% (w/v) follows the Michaelis kinetics. The Michaelis constant (2,805 +/- 130 microM), the maximal velocity (196 +/- 2.2 x 10(-5 )microM s(-1)) and the velocity constant (6.98 +/- 0.2 x 10(-7) s(-1)) was calculated. The viability assessed by fluorescent dyes simultaneously visualized the live/dead cell ratio, which can be calculated by image analysis software.


CONCLUSIONS
The use of MTT in colorimetric assays offers high sensitivity. The assay is simple, inexpensive, and reproducible in vein homograft studies.
Galambos
B
B
Department of Vascular Surgery, Petz Aladar University Teaching Hospital, Gyor, Hungary. gbarna69@axelero.hu
Csönge
L
L
Olah
A
A
von Versen
R
R
Tamas
L
L
Zsoldos
P
P
D000328:Adult; D001706:Biopsy; D001807:Blood Vessel Prosthesis; D002470:Cell Survival; D006801:Humans; D066298:In Vitro Techniques; D007202:Indicators and Reagents / Q000494:pharmacology*; D010084:Oxidation-Reduction; D015203:Reproducibility of Results; D012501:Saphenous Vein / Q000187:drug effects* / Q000473:pathology / Q000502:physiology; D013778:Tetrazolium Salts / Q000494:pharmacology*; D014022:Tissue Survival
D007202:Indicators and Reagents; D013778:Tetrazolium Salts