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The Bosterbio OneStep Human AFP Pre-Coated ELISA (Enzyme-Linked Immunosorbent Assay) kit is a solid phase immunoassay specially designed to measure Human AFP with a 96-well strip plate that is pre-coated with antibody specific for AFP. The detection antibody is a HRP conjugated antibody specific for AFP. The capture antibody is monoclonal antibody from mouse, and the detection antibody is monoclonal antibody from mouse. The kit is analytically validated with ready to use reagents. To measure Human AFP, add standards and samples to the wells, then add the HRP conjugated detection antibody. Wash away the unbounded protein and HRP conjugated detection antibody. TMB is substrate to HRP and will be catalyzed to produce a blue color product, which changes into yellow after adding acidic stop solution. Upon addition of the substrate, the density of the yellow product is linearly propotional to Human AFP in the sample. Read the density of the yellow product in each well using a plate reader, and benchmark the sample wells' readings against the standard curve to determine the concentration of Human AFP in the sample. For more information on assay principle, protocols, and troubleshooting tips, see Boster's ELISA Resource Center at https://www.bosterbio.com/elisatechnical-resource-center.
*Our Boster Guarantee covers the use of this product in the above tested applications.
**For positive and negative control design, consult "Tissue specificity" under Protein Target Info.
1. Microplate Reader capable of reading absorbance at 450nm.
2. Automated plate washer (optional)
3. Pipettes and pipette tips capable of precisely dispensing 0.5 µl through 1 ml volumes of aqueous solutions. Multichannel pipettes are recommended for large amount of samples.
4. Deionized or distilledwater.
5. 500ml graduated cylinders.
6. Test tubes for dilution.
You can check the tissue specificity below for information on selecting positive and negative control.
Click the images to enlarge.
It is recommended that all reagents and materials be equilibrated to 37°C/room temperature prior to the experiment (see Preparation Before The Experiment if you have missed this information).
Average the duplicate readings for each standard, sample, and control. Subtract the average blank O.D. reading.
It is unnecessary to set blank control for dual wavelength plate reader.
It is recommended that a standard curve be created using computer software to generate a four parameter logistic (4-PL) curve-fit. A free program capable of generating a four parameter logistic (4-PL) curve-fit can be found online at: www.myassays.com/four-parameter-logistic- curve.assay.Alternatively, plot the mean absorbance for each standard against the concentration. The measured concentration in the sample can be interpolated by using linear regression of each average relative OD against the standard curve generated using curve fitting software. This will generate an adequate but less precise fit of the data.For diluted samples, the concentration reading from the standard curve must be multiplied by the dilution factor.
*EK7020 bulk discount: 20% off for 4+ of same kits, 30% off for 20+ of same kits, applicable only to USA and Canada.
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