Horseradish Peroxidase Labeling Service

Horseradish peroxidase (HRP) is one of the most commonly used enzyme labels and is frequently employed as a colorimetric marker in the area of antibody-based antigen detection assays [western blot, enzyme-linked immunosorbent assay (ELISA), and immunohistochemistry (IHC)]. Combining years of antibody or protein labeling expertise and experience, Biologics International Corp. (BIC) provides rapid, efficient, and high-quality custom HRP labeling services for both research and commercial use, saving our customers’ precious time.

For more information, please contact us! Our scientists welcome the opportunity to discuss your projects directly with you.

Highlights

• Comprehensive service: A full range of services from antibody or protein production to labeling.
• Rich experience: Using our stable and mature platforms of antibody or protein production, we have delivered more than 300 labeled proteins or antibodies with high bioactivity and sensitivity.
• Diverse selection: If you require additional labeling selections, we can offer a wide variety of labels, including fluorescein isothiocyanate (FITC), biotin, and isotopes.

HRP

HRP, found in the roots of horseradish, is a 44 KDa glycoprotein which is used extensively in biochemistry applications for its ability to produce a colored, fluorimetric, or luminescent derivative when incubated with the appropriate substrate.

Application of HRP Labeling

HRP is generally used to label water-soluble biomolecules (such as antibodies and proteins), containing reactive and accessible primary amino groups, with peroxidase for use in analytical methods. It is particularly suitable for the coupling with antibodies, as the resulting conjugate is optimal for use in immunochemical detection systems (such as western blot, ELISA, and IHC).

Methods of HRP Labeling

In order to obtain HRP-labeled antibodies or proteins, cross-linkers (such as glutaraldehyde, periodate, and maleimide derivatives) are essential. As a kind of glycoprotein, HRP can be activated for conjugation with antibody by treatment with periodate. Polysaccharide residues on the surface of HRP are oxidized with sodium periodate to produce reactive aldehyde groups that can be conjugated to amino groups of the antibody molecule forming an HRP-antibody or HRP-protein complex.

Color Development with HRP Labeling

The HRP enzyme alone and conjugates thereof are of little value. In the presence of specific substrates, HRP can catalyze the transfer of two electrons from a substrate to hydrogen peroxide to produce an oxidized substrate along with water. The complex of HRP-labeled antibodies and substrates generates a chromogenic, chemiluminescent, or fluorescent signal upon oxidation.

Of note, different substrates will produce signals with different spectral properties, for example, HRP reacts with diaminobenzidine to produce a brown-colored product or with luminol to produce light. Other subtrates like TMB and Amplite Red, can yield signals at 650 nm and 585 nm, respectively.

Advantages of HRP Labeling

With six lysine residues, HRP can be conjugated easily to a antibody or protein. Although HRP label has a bigger size than that of biotin (approximately 244.3 Da), it rarely disrupts antibody or protein function. The reason is that the relatively small size of HRP allows greater penetration into sample tissues and cells and reduces the likelihood of interference with the function of the conjugated antibody or protein. Due to its monomeric nature, HRP is often used in the detection of specific proteins on western blots, or in techniques such as ELISA and immunohistochemistry.

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