Evaluation procedure


prepare all reagents, working standards, blanks, and samples according to the instructions in the preceding sections.refer to the assay layout sheet to calculate the number of wells to place any unused wells and desiccant back into the pouch, reseal the pouch, and store the wells at 4C.add 50 mL of the standard to the testing standard well, followed by 10 mL of the testing sample (sample final dilution is fivefold). Pipette the sample to the wells, avoiding the well walls as much as possible, and mix gently.cover with the included adhesive strip and incubate for 30 min at 37 °C.configure liquid, 30 times dilute wash solution with distilled water to configure liquid.washing, remove the adhesive strip, discard the liquid, and then pipette washing buffer into each well. Allow to stand for 30 s and then drain.add enzymes, pipette 50 l of HRP-conjugate reagent into each well, excluding the well designated as a blank.incubate, operation with fourwashing, using number 6add 50 ml of chromogen solution A and 50 ml of chromogen solution B to each well, avoiding light for 15 min at 37 °C.stop the reaction by pipetting 50 ml of stop solution into each well (the blue change to yellow).Calculate the value of the blank well as zero. Read absorbance at 450 nm 15 min after pipetting stop solution.