Reliable TMB Substrate Kit

University of California, Merced
Molecular and Cell Biology
Graduate Student

Overall

Quality of Results

Ease-of-Optimization

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Company:

Vector Laboratories

Product Name:

TMB Substrate Kit

Catalog Number:

SK-4400

This is an easy to use TMB substrate kit for a number of applications. We use this TMB for use in sandwich ELISAs. Provided your upstream experiments work then this substrate will provide bright clear results.

Experimental Design and Results Summary

Application

ELISA

Starting Material

Antibody ELISAs with an HRP-conjugated secondary antibody and murine cells

Protocol Overview

Mix 2 drops Buffer stock, 3 drops TMB Stock solution, and 2 drops Hydrogen Peroxide solution into 5mL of milliQ water. DO NOT use PBS or other substrates to make substrate. For ELISA, do not use the stabilizing solution that comes in the kit. Pipette 50-100ul into each ELISA well and read when desired results are achieved.

Tips

If you are using a high a concentration of HRP-conjugated secondary, then TMB can react within seconds.

Results Summary

Results can be read with inactive HRP (bright yellow) at A450 after stopping with 1N sulfuric acid at A600.

DOI or PMID #

N/A

Additional Notes

N/A

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Summary

The Good

Once your other assay is optimized this substrate reacts well with bright signal.

The Bad

Can react too quickly with high concentrations of HRP.

The Bottom Line

If your assay is in order then this product will work for you.

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