Application of Ethidium Bromide Fluorescence Assay to Measurement of Blastogenesis of Swine Lymphocytes

Abstract

The ethidium bromide (EB) fluorescence assay was applied to evaluate blastogenesis of swine lymphocytes by mitogens. The optimal responses were observed in cultures of 1.0×10⁶ cells for 72hr with the final concentration of PHA-P of 5μl/ml, Con A of 25μg/ml or PWM of 0.25μg/ml. At the optimal conditions, values of stimulation index by the EB fluorescence assay (SI-FA) correlated highly with those of stimulation index by the radioisotope incorporation assay (SI-RI) (r=0.752, n=21, P<0.01). The correlation coefficients were r=0.863 in PHA-P and Con A (n=7, P<0.05) and r=0.710 in PWM (n=7, P>0.05). From these results, this EB fluorescence assay is thought to be possible to be used in place of the radioisotope incorporation assay for evaluation of blastogenic response of swine lymphocytes. The SI-FA values of blood lymphocytes of healthy pigs at about 6 months old were 1.54±0.26 to PHA-P, 1.60±0.23 to Con A and 1.56±0.28 to PWM.