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Biotin Conjugated Antibodies & Reagents

Biotin conjugated (biotinylated) antibodies and reagents are used to detect a wide range of antigens with techniques including Western blot, ELISA, flow cytometry, immunocytochemistry (ICC), and immunohistochemistry (IHC). A common approach involves identifying the target of interest with a biotinylated primary antibody before adding an enzyme- or fluorophore-labeled avidin/streptavidin conjugate to produce a measurable signal. Alternatively, an unlabeled primary antibody can be detected with a biotinylated secondary antibody prior to conjugate addition.

A major advantage of using biotin conjugated antibodies and reagents for antigen detection is that it provides signal amplification, enhancing assay sensitivity. We offer an extensive selection of biotinylated primary antibodies, secondary antibodies, and isotype controls, as well as many different Streptavidin and Neutralite Avidin Conjugates.

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Frequently Asked Questions

Biotin, also known as Vitamin B7, plays a crucial role in the metabolism of carbohydrates, fats, and proteins. The bond between biotin and avidin/streptavidin is the strongest known non-covalent interaction (Kd ~ 10-15 M), capable of withstanding organic solvents, denaturing reagents, and extremes in pH and temperature. As a result, the biotin-avidin/streptavidin interaction has been widely exploited for scientific research, mainly for antigen detection and purification purposes.

Biotinylated secondary antibodies enhance signal detection by increasing the amount of enzyme or fluorescent label that is accumulated on the target of interest. This is accomplished in two main ways. First, multiple secondary antibodies bind to each primary antibody, owing to the fact that most secondary antibodies are polyclonal. Additionally, because each secondary antibody is conjugated to multiple biotin molecules, high numbers of sites are available for avidin/streptavidin binding. Being able to enhance signal detection in this way is especially useful for detecting antigens that have low expression levels.

We offer a comprehensive range of Streptavidin and Neutralite Avidin Conjugates for detecting biotinylated antibodies. These include reagents for enzyme-based detection with horseradish peroxidase (HRP), alkaline phosphatase (AP), and β-galactosidase (BGAL), which should be paired with an appropriate substrate, and a broad array of fluorescent conjugates. Available fluorophores include the fluorescent proteins allophycocyanin (APC) and phycoerythrin (PE), fluorescent dyes such as Cyanine 3 (CY3), Cyanine 5 (CY5), Fluorescein (FITC), Rhodamine (TRITC), and Texas Red® (TXRD), and various tandem dye reagents.

Native avidin has a basic isoelectric point (pI = 10.5) and a high carbohydrate content, which can lead to non-specific binding and high background signal in applications such as Western blotting, ELISA, and IHC. Neutralite avidin, which was developed to overcome this problem, has a near-neutral isoelectric point (pH 6.3), minimizing interactions with negatively-charged structures like DNA/RNA. It is also deglycosylated, decreasing the possibility of lectin binding. Another important benefit of Neutralite Avidin is that it lacks an RYD (Arg-Tyr-Asp) recognition sequence. This gives it an advantage over streptavidin, in which the RYD recognition sequence can result in off-target binding to cell surface receptors.

When using biotin-streptavidin systems for applications such as Western blotting and ELISA, researchers are advised to avoid non-fat milk for blocking. This is because milk contains endogenous biotin that can interfere with the detection process. Recommended alternatives include bovine serum albumin (BSA) and purified (biotin-free) proteins, such as casein, although the blocking step should always be optimized during assay development. For ICC and IHC, the presence of endogenous biotin, biotin receptors, and avidin/streptavidin binding sites in cells and tissue samples can lead to unwanted background signal. Commercial avidin/streptavidin and biotin blocking kits have been developed for these applications.

Related Product Categories

Conjugated Antibodies

For the direct detection of your target, explore our offering of over 25 enzyme and fluorochrome conjugated primary antibodies.

Primary Antibodies

Find a high-quality primary antibody from our fast-shipping selection, all produced to our exacting standards in our own facility.

Secondary Antibodies & Reagents

With a wide range of ready-to-ship secondary antibodies that are second to none, SouthernBiotech will keep your assays running and projects on schedule.

Streptavidin & Neutralite Avidin

Our streptavidin and avidin provide exceptional performance in a range of applications requiring biotin-binding proteins for detection.