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ELISA protocol

ELISA kit protocol

Summary of Assay Procedure for Sandwich Kit:

-Prepare all reagents, samples and standards,

- Add 100 ul of standard or sample to each well,

    -Seal the plate and incubate for 2 hrs at 37 °C

-Wash the plate 3 times with wash buffer solution,

-Add 100 ul of Biotin conjugated detection antibody,

    -Seal the plate and incubate for 1 hr at 37 °C

-Wash plate 3 times with wash buffer solution,

-Add 100 ul of HRP conjugated detection antibody,

    -Seal the plate and incubate for 30 mins at 37 °C

-Wash plate 5 times with wash buffer solution,

-Add 90 ul of TMB substrate solution,

    -Seal the plate and incubate for 20 mins at 37 °C

-Add 50 ul of stop solution,

-Read the plate at 450 nm.


Summary of Assay Procedure for Competitive Kit:

-Prepare all reagents, samples and standards,

- Add 50 ul of standard or sample to each well,

-Immediately add 50 ul of Biotin conjugated detection antibody,

    -Seal the plate and incubate for 1 hr at 37 °C

-Wash plate 3 times with wash buffer solution,

-Add 100 ul of HRP conjugated detection antibody,

    -Seal the plate and incubate for 30 mins at 37 °C

-Wash plate 5 times with wash buffer solution,

-Add 90 ul of TMB substrate solution,

    -Seal the plate and incubate for 20 mins at 37 °C

-Add 50 ul of stop solution,

-Read the plate at 450 nm.


Please note quantities of samples, reagents and incubation periods can vary between each kit, please refer to the product manual for further information.