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Eur Surg Res 2004;36:371-375 (DOI: 10.1159/000081647)

  Key Words

• Fluorescent dye
• Homograft
• MTT test
• Viability assay

  Abstract

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. Materials and 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 µg/ml propidium iodide PI and 4 µM/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 µM), the maximal velocity (196 ± 2.2 × 10^-5 µM s^-1) and the velocity constant (6.98 ± 0.2 × 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. Conclusion: The use of MTT in colorimetric assays offers high sensitivity. The assay is simple, inexpensive, and reproducible in vein homograft studies.

Copyright © 2004 S. Karger AG, Basel

  Author Contacts

Barnabás Galambos
Department of Vascular Surgery
Petz Aladar University Teaching Hospital 9024, 2 Vasvari P.
HU-9024 Gyr (Hungary)
Tel. +36 96 507 957, Fax +36 96 418 244/1203, E-Mail gbarna69@axelero.hu

  Article Information

Received: April 5, 2004
Accepted after revision: August 11, 2004
Number of Print Pages : 5
Number of Figures : 3, Number of Tables : 1, Number of References : 24