Abstract: Objective In order to reduce the harm of OPD(O-phenylenediamine) to the operators and pollution to environment, TMB (tetramethylbenzidine) was used as a color reagent to replace OPD in ELISA. Methods The operation was carried out on the basis of GB/T 14926.50-2001, to compare stability, specificity, sensitivity and repeatability of two color systems in detection of mouse hepatitis virus (MHV) antibody by ELISA. Result The TMB is more stable tiian OPD, the relative deviation (15.3%) is smaller than OPD(17.7%); the special antigen P/N value of TMB is higher than OPD and the difference is significant (P＜0.05); The working concentration of Enzyme of TMB is 0.25 times that of OPD, and the detection sensitivity is twice that of OPD; When using TMB, The inter-assay and the intra-assay average coefficient of variation of normal antigen and special antigen is 3.9%513.0% and 4.7%, 11.3% while the OPD is 2.7%, 11.2% and 4.5%,9.5%. Conclusion TMB is better than OPD in stability，reproduction, sensitivity and specificitys. TMB as a color reagent may replace OPD in detection of mouse hepatitis virus (MHV) antibody by ELISA.

Key words: ELISA, TMB, OPD