To determine if the presence of cardiac light chains in blood could be used to detect acute myocardial infarction, we developed a specific light chain immunoassay. A synthetic peptide sequence specific for human cardiac ventricular myosin light chain 1 (VLC1) was synthesized and designated P348. This peptide coupled to keyhole limpet hemocyanin was used as an immunogen to obtain murine monoclonal antibodies specific for VLC1. Five monoclonal antibodies were obtained. One of these designated Mab-8E3 reacted equally well with both the synthetic peptide and VLC1. Although the 8E3 antibody is specific for VLC1, the use of HPLC purification of skeletal muscle myosin light chain 1 demonstrated that VLC1 is present in human skeletal muscle. The clinical utility of the assay was tested in 18 patients with creatine kinase (CK) and ECG documented acute myocardial infarction. VLC1 was below the limit of detection (< 1 ng/ml) in sera obtained from healthy volunteers and patients without myocardial infarction or chest pain. In contrast VLC1 was elevated in the serum of all 18 patients with acute myocardial infarction. Combining the two test results at the time of admission resulted in 83% of patients having detectable serum levels of one or both markers.