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Heart-type fatty acid-binding protein (H-FABP) is a low molecular weight cytoplasmic protein and present abundantly in the myocardium. When the myocardium is injured, as in the case of myocardial infarction, low molecular weight cytoplasmic proteins including H-FABP are released into the circulation and H-FABP is detectable in a blood sample. We have already developed a direct sandwich-ELISA for quantification of human H-FABP using two distinct types of monoclonal antibodies specific for human H-FABP. In this study we investigated the clinical validity of H-FABP as a biochemical diagnostic marker in the early phase of acute myocardial infarction (AMI). To evaluate the diagnostic usefulness of H-FABP in the early phase of AMI, blood samples were obtained from the following patients within 12 hours after the appearance of symptoms, and serum levels of H-FABP were compared with those of conventional diagnostic markers, such as myoglobin and creatine kinase isoenzyme MB (CK-MB). Blood samples were collected from patients with confirmed AMI (n=140), patients with chest pain who were afterwards not classified as AMI by normal CK-MB levels (non-AMI) (n=49) and normal healthy volunteers (n=75). The serum concentration of H-FABP was quantified with our direct sandwich-ELISA. The concentration of myoglobin mass was measured with a commercial RIA kit. The serum CK-MB activity was determined with an immuno-inhibition assay kit. The overall sensitivity of H-FABP, within 12 hours after the appearance of symptoms, was 92.9%, while it was 88.6% with myoglobin and 18.6% with CK-MB. The overall specificity of H-FABP was 67.3%, while it was 57.1% with myoglobin and 98.0% with CK-MB. The diagnostic efficacy rates with these markers were 86.2% (H-FABP), 80.4% (myoglobin) and 39.2% (CK-MB), respectively. The diagnostic validity of H-FABP was further assessed by receiver operating characteristic (ROC) curve analysis. The area under the curve (AUC) of H-FABP was 0.921, which was significantly greater than with myoglobin (AUC: 0.843) and CK-MB (AUC: 0.654). These parameters, such as sensitivity, specificity, diagnostic efficacy and diagnostic accuracy, obtained for patients with chest pain within 3 hours and/or 6 hours after the onset of symptoms were almost the same as those for patients within 12 hours after symptoms. H-FABP is more sensitive than both myoglobin and CK-MB, more specific than myoglobin for detecting AMI within 12 hours after the onset of symptoms, and shows the highest values for both diagnostic efficacy and ROC curve analysis. Thus, H-FABP has great potential as an excellent biochemical cardiac marker for the diagnosis of AMI in the early phase.

Long-chain acyl-coenzyme-A synthetase from the microsomes as well as from the mitochondrial fraction of rat liver has been purified to homogeneity as evidenced by dodecylsulfate/polyacrylamide gel electrophoresis, amino-terminal analysis and the elution profile at the final chromatography step. The purification procedure involves resolution of the cellular particles with Triton X-100 and chromatography on Blue-Sepharose, hydroxyapatite and phosphocellulose. The purified enzymes from both sources have a specific activity of 26-29 units/mg protein at 35 'C, which is more than 100-fold higher than those of long-chain acyl-CoA synthetases of animal and bacterial origin hitherto reported. The purified enzymes exhibit a molecular weight of approximately 76 000 as estimated by dodecylsulfate/polyacrylamide gel electrophoresis and catalyze the activation of saturated fatty acids with 10-18 carbon atoms and unsaturated fatty acids with 16-20 carbon atoms most efficiently. The purified enzyme from the microsomes and that from the mitochondrial fraction, which are obtained by essentially identical procedures, are indistinguishable from each other with respect to all molecular and catalytic properties examined, including molecular weight, amino acid composition, amino-terminal residue, heat stability, specific activity, pH optimum and substrate specificity regarding fatty acid, acyl acceptor and nucleoside 5'-triphosphate.

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