Abstract
Human plasma contains at least three forms of adiponectin: a trimer, a hexamer, and a high-molecular-weight (HMW) multimer. We purified HMW adiponectin from human plasma using its affinity to gelatin and obtained monoclonal antibodies against it. On Western blot analysis, the reactivity of these monoclonal antibodies was shown to be restricted to a non-heat-denatured form of adiponectin molecules. On heating, the collagen-like domain of adiponectin molecules became denatured, and thus the trimer form could not be maintained. From these, monoclonal antibodies against HMW adiponectin were suggested to react with the intact trimer of adiponectin. With these monoclonal antibodies, we developed a sandwich ELISA system for quantifying adiponectin in human serum. Its specificity was verified by analysis of serum fractions separated by gel-filtration chromatography, and our ELISA system was found to be HMW adiponectin-specific. With this novel ELISA, the HMW adiponectin concentrations were 8.4 +/- 5.5 microg/ml (mean +/- SD) in healthy women and 6.2 +/- 3.6 microg/ml in healthy men. Also, serum with a lower HMW adiponectin concentration was shown to have a lower HMW ratio (i.e., HMW adiponectin/total adiponectin).
Highlights
Human plasma contains at least three forms of adiponectin: a trimer, a hexamer, and a high-molecular-weight (HMW) multimer
Adiponectin mRNA levels are significantly reduced in omental adipose tissue of obese patients with type 2 diabetes compared with lean and obese normoglycemic subjects, and less pronounced, the levels are reduced in subcutaneous adipose tissue of type 2 diabetic patients [8]
Tietge et al [11] reported that plasma adiponectin levels in cirrhosis are increased significantly, because the liver is a major source of adiponectin extraction and the adiponectin levels in cirrhosis do not correlate with parameters of body composition or metabolism but exclusively with reduced liver function and altered hepatic hemodynamics
Summary
Human plasma contains at least three forms of adiponectin: a trimer, a hexamer, and a high-molecular-weight (HMW) multimer. Monoclonal antibodies against HMW adiponectin were suggested to react with the intact trimer of adiponectin. With these monoclonal antibodies, we developed a sandwich ELISA system for quantifying adiponectin in human serum. Its specificity was verified by analysis of serum fractions separated by gel-filtration chromatography, and our ELISA system was found to be HMW adiponectinspecific. With this novel ELISA, the HMW adiponectin concentrations were 8.4 6 5.5 mg/ml (mean 6 SD) in healthy women and 6.2 6 3.6 mg/ml in healthy men.
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