Ultrasensitive Insulin ELISA英文簡述: INTENDED USE The ALPCO Ultrasensitive Insulin ELISA is designed for the quantitative determination of insulin in human serum and plasma. PRINCIPLE OF THE ASSAY The ALPCO Ultrasensitive Insulin ELISA is a sandwich type immunoassay. The 96-well microplate is coated with a monoclonal antibody specific for insulin. The standards, samples, and the control are added to the microplate wells with the Detection Antibody. The microplate is then incubated at room temperature on a microplate shaker at 700-900 rpm. After the first incubation is complete, the wells are washed with Wash Buffer and blotted dry. TMB Substrate is added, and the microplate is incubated a second time at room temperature on a microplate shaker at 700-900 rpm. Once the second incubation is complete, Stop Solution is added, and the optical density (OD) is measured by a spectrophotometer at 450 nm. The intensity of the color generated is directly proportional to the amount of insulin in the sample. CALCULATION OF RESULTS Construct a standard curve from the standards. The Zero Standard should be used as a blank with its average value subtracted from each well. It is recommended to use a software program to calculate the standard curve and to determine the concentration of the samples. The ALPCO Ultrasensitive Insulin ELISA is a ligand binding assay, with responses exhibiting a sigmoidal relationship to the analyte concentration. Currently accepted reference models for such curves use a 4 or 5 parameter logistic (pl) fit, as these models optimize the accuracy and precision across a greater range. Although cubic spline and other models are acceptable methods, they generally show less intra-assay accuracy and precision at the low and high ends of the range.