Use of a colorimetric assay to measure differences in cytotoxicity of Mycobacterium tuberculosis strains.
Several techniques have been used to quantify the cytotoxicity produced by Mycobacterium tuberculosis bacilli on cell monolayers; however, they are semi-quantitative or time consuming. Herein, a method based on crystal violet (CV) uptake by THP-1 cell monolayers is described. This colorimetric method quantifies the cytotoxic effect as a function of the number of remaining cells after the infection with M. tuberculosis. Since this micro-organism is not stained by the dye, it does not produce a background that affects absorbance readings. As determined by CV assay (CVA), M. tuberculosis strain H37Rv destroyed 10.5 % of THP-1 cell monolayers at 24 h and 50.52 % at 72 h, while M. tuberculosis strains lacking the complete phospholipase C locus produced a reduced cytotoxic effect. The damage estimated by microscopy corresponded to the effect quantified by CVA. The results show that the use of CVA is a rapid, sensitive and reliable quantitative assay to measure the cytotoxicity of different M. tuberculosis strains.
Castro-Garza
Jorge
J
División de Biología Celular y Molecular, Centro de Investigación Biomédica del Noreste, Instituto Mexicano del Seguro Social, Monterrey, NL, Mexico.
Barrios-García
Hugo B
HB
Facultad de Ciencias Biológicas, Universidad Autónoma de Nuevo León, San Nicolás de los Garza, NL, Mexico.|División de Biología Celular y Molecular, Centro de Investigación Biomédica del Noreste, Instituto Mexicano del Seguro Social, Monterrey, NL, Mexico.
Cruz-Vega
Delia Elva
DE
División de Biología Celular y Molecular, Centro de Investigación Biomédica del Noreste, Instituto Mexicano del Seguro Social, Monterrey, NL, Mexico.
Said-Fernández
Salvador
S
División de Biología Celular y Molecular, Centro de Investigación Biomédica del Noreste, Instituto Mexicano del Seguro Social, Monterrey, NL, Mexico.
Carranza-Rosales
Pilar
P
División de Biología Celular y Molecular, Centro de Investigación Biomédica del Noreste, Instituto Mexicano del Seguro Social, Monterrey, NL, Mexico.
Molina-Torres
Carmen A
CA
Servicio de Dermatología, Hospital Universitario 'José E. González', Monterrey, NL, Mexico.
Vera-Cabrera
Lucio
L
Servicio de Dermatología, Hospital Universitario 'José E. González', Monterrey, NL, Mexico.
D001426:Bacterial Proteins / Q000235:genetics; D001431:Bacteriological Techniques / Q000379:methods*; D045744:Cell Line, Tumor; D002470:Cell Survival; D003124:Colorimetry / Q000379:methods*; D017353:Gene Deletion; D005840:Gentian Violet / Q000378:metabolism; D006801:Humans; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000472:pathogenicity*; D012680:Sensitivity and Specificity; D010738:Type C Phospholipases / Q000235:genetics
D001426:Bacterial Proteins; D010738:Type C Phospholipases; D005840:Gentian Violet