TMB Peroxidase Substrate Systems

In the last 50 years, the ELISA has been used to analyze biochemical substances. In the early 1970s, a scientist named Perlmann described ELISA as an assay used to help determine if there is a presence of ligand.

ELISA has been shown to be an effective tool in diagnostic, biotechnology, quality control, as well as pathology. But, first, what does ELISA stand for? This is short for enzyme-linked immunosorbent assay. In brief, you take a sample, and the antigens that are in that specific sample are located on the surface.

After that, you need to apply an antibody that will connect and bind with the antigen. And in the end, one final step is in order, and that is an adding substance. This substance needs to contain an enzyme that will produce a reaction or a signal. The signal usually is seen by changing the color of the sample. To keep reading, follow the link


Before Perlmann developed ELISA, there were other ways to conduct an immunoassay. The immunoassays are mainly used to see if there is a presence of an analyte. This analyte can be antibodies or even proteins that a certain individual has during an infection. These analytes are produced in that individual’s body as a result of that same infection.

That is why immunoassays are quite important to be developed in a specific way. These immunoassays are a type of biochemical tests and have wide usage in today’s life. Before ELISA was created, people used radioimmunoassay.

The importance of enzyme-linked immunosorbent assay can be clearly seen when you realize how the radioimmunoassay affects human health. They can pose a threat to the health of a certain individual. Then is when scientists discovered that there needs to be another way to conduct these assays.

As mentioned above, in order to finish the ELISA application, you would need a suitable substrate. But what is the best one to use and see the best results? After years of research and development, it was concluded that the best substrate is the TMB peroxide.

TMB peroxide 

TMB, or more specifically Tetramethylbenzidine, is a substrate that helps ELISA be conducted. This is a chromogenic substance that has a green-blue color when mixed with ethyl acetate. It can be degraded by fluorescent lights, as well as sunlight.

This is why it should be kept away from sunlight. The substrate is photosensitive, and it has been in usage when a benzidine replacement was needed. The o-phenylenediamine and benzidine are carcinogenic substrates, and that is why today, the TMB Peroxidase substrate is the best substrate you can use for your immunoassay.

A popular enzyme that can detect the substances, as mentioned earlier, is horseradish peroxidase. You can come across HRP, which is short for horseradish peroxidase. This HRP is a stable enzyme with a small size and can be implemented in a peptide, hapten, or an antibody, for that matter.

You may find different substrates to use along HRP, and they all have additional features. However, the most commonly used is the TMB because of its great characteristics. If you find a good and quality provider, the TMB is the best way to go.

TMB Peroxidase Characteristics 

As mentioned above, you can use different substrates when conducting ELISA. But TMB has several advantages over the other substrates. Some of them are commonly known, but there are others that need to be pointed out.

For example, Tetramethylbenzidine has huge sensitivity. The sensitivity of a substrate is one of the first things you should be looking for if you want a successfully conducted assay. With various kinetic rates, several formulations will meet the range that is required.

Furthermore, TMB has the ability to reach the desired range by quantitation and signal-to-noise ratios. But the increased sensitivity isn’t the only advantage of this substrate. It is super easy to manufacture it too. Read more here.

This is quite an important advantage because it is a reflection of reliability and a trustworthy product. Plus, it is stabler than most of the other products, which means that it can be used for four years. If you want to get rid of the yields in the background, this is for sure what you are looking for.