Human muscle specific enolase (hMSE) plays an important role in the diagnosis of acute myomuscle infarction and muscular diseases. A hMSE RIA is developed.An affinity coefficient of antiserum is 5.1×10⁹. ¹²⁵I-hMSE as the tracer is prepared using improved BHR method and its binding rate with antiserum(1∶1000) is up to 50.16%. The variable coefficients of intra and inter-assay are 8.6% and 13%, respectively. The recovery rate is 95%-105%. The standards range from 5 to 320μg/L and the minimum detectable value is 5μg/L. In the assay there is an optimal reaction condition: 0.1mol/L pH7.4 PBS(5mmol/L MgSO₄，0.1% BSA， 0.1% Toween-20, 0.1% NaN₃，0.5% NaCl);and incabation temperature and times: 37℃，0.5h and 4℃，0.5h (as rapid measurement) or 4℃ 24h. On the basis of 65 healthy control，the hMSE levels are 23.9±10.9μg/L (x±s).A positive value is higher than 57μg/L. The hMSE levels are signifcantly increased in the patients with both acute myocardial infarction and muscular diseases.