Enzyme-linked immune test steps

Enzyme-linked immune test steps

Step 1: Number: determine the number of well (samples and standards) to be used and store unused wells in 4 ℃. Every sample and standard must be parallel well (2well), and record their location

Step 2: Addtion reaction: Add 50μl standard or sample into marked well, then add 50μl HRP-conjugate solution into each well, next, add 50μl antibody working solution into each well

Step 3: Incubate: Cover with the adhesive Membrane, oscillate gently for 5s and incubate for 30 min at 25℃.

Step 4: Washing: Uncover the adhesive Membrane, discard liquid, pipette 250μl washing buffer to every well, still for 30s then drain, repeat 5 times, Pat dry with a blotting paper

Step 5: Color: Pipette 50ul Substrate A Solution, then pipette 50ul Substrate B Solution to each well, oscillate gently for 5s, avoid the light preservation for 15 min at 25℃

Step 6: Stop the reaction: Pipette Stop Solution 50μl to each well, oscillate gently, stop the reaction (the blue change to yellow).

Step 7: Calculate: Read absorbance at 450nm with microplate reader (Recommend reading the OD value at the dual-wavelength 450/630nm).finish this step within 10min.