Description

Description: Specification Sensitivity: 0.188 ng/mL (The sensitivity of this assay, or Lower Limit of Detection (LLD) was defined as the lowest protein concentration that could be differentiated from zero). Detection Range: 0.313-20 ng/mL Specificity: This kit recognizes Biotin. No significant cross-reactivity or interference was observed. Repeatability: Coefficient of variation is <10%. Test principle This kit uses Competitive-method. The microplate provided in this kit has been pre-coated with Biotin. During the reaction, Biotin in the sample or standard competes with a fixed amount of Biotin on the solid phase supporter for sites on the Avidin-HRP specific to Biotin. Excess Avidin-HRP and unbound sample or standard are washed from the plate. Then a TMB substrate solution is added to each well. The enzyme-substrate reaction is terminated by adding Stop Solution and the color change can be measured spectrophotometrically at a wavelength of 450 nm +/- 2 nm. The concentration of Biotin in samples can be calculated by comparing the OD of the samples with the standard curve. Calculation of results Average the duplicate readings for each standard and samples. Plot a four-parameter logistic curve on log-log graph paper, with standard concentration on the x-axis and OD values on the y-axis. If samples have been diluted, the concentration calculated from the standard curve must be multiplied by the dilution factor. If the OD of the sample surpasses the lower limit of the standard curve, you should re-test it after appropriate dilution. The actual concentration is the calculated concentration multiplied dilution factor. SUMMARY 1. Add 50 µL standard or sample to each well. 2. Immediately add 50 µL Avidin-HRP to each well. 3. Incubate for 30 min at 37°C 4. Aspirate and wash 5 times 5. Add 90 µL Substrate Reagent. Incubate 15 min at 37°C 6. Add 50 µL Stop Solution. Read at 450 nm immediately. 7. Calculation of results Kit components & Storage The unopened kit can be stored at 4°C for 1 month. If the kit is not used within 1 month, store the items separately according to the following conditions since the kit is received. Note: All reagent bottle caps must be tightened to prevent evaporation and microbial pollution. The volume of reagents in partial shipments is a little more than the volume marked on the label, please use in measuring instead of directly pouring. Other supplies required Microplate reader with 450nm wavelength filter High-precision transferpettor, EP tubes and disposable pipette tips 37°C Incubator, Deionized or distilled water Absorbent paper

Additional Text

Detection Method

Colorimetric

Storage Note

2-8°C,12 months