Goat Anti-Mouse IgG (H+L) Secondary Antibody, Biotin Conjugated

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Background

Most commonly, secondary antibodies are generated by immunizing the host animal with a pooled population of immunoglobulins from the target species. The host antiserum is then purified through immunoaffinity chromatography to remove all host serum proteins, except the specific antibody of interest, and then modified with antibody fragmentation, label conjugation, etc. to achieve highly specific reagents for molecular detection. Secondary antibodies can be conjugated to a large number of labels, including enzymes, biotin, and fluorescent dyes/proteins. Here, the antibody provides the specificity to locate the protein of interest by recognizing a primary antibody that targets a particular antigen, and the label generates a detectable signal in the area of the formed immune complexes. The label of choice depends upon the experimental application.
Biotinylated antibodies are widely used in systems where signal amplification is desired. Often 15-20 biotin moieties are coupled to a single IgG secondary antibody. Biotin binds avidin, streptavidin, or neutravidin with a high degree of affinity and specificity. In immunoassays avidin/streptavidin-biotin binding is used as a bridge between antibodies and reporters like enzymes (HRP, AP), fluorophores, chromophores, etc. Both avidin and streptavidin are tetrameric proteins capable of binding 4 biotin groups to each molecule of avidin or streptavidin, thus amplifying the signal intensity and detection sensitivity by increasing the concentration of reporters at the antigenic site. Two main biotin-binding detection systems have been widely utilized: Avidin-Biotin Complex (ABC) and Labeled Streptavidin Biotin (LSAB) methods. In the ABC method free avidin (or streptavidin) is used as a bridge/link between the biotinylated antibody and а biotinylated reporter molecule, resulting in three reporter molecules coupled to the biotinylated antibody. The LSAB method employs a reporter-labeled streptavidin (avidin or neutravidin can alternatively be used) to detect the bound biotinylated-secondary antibody on the tissue section, blotting membrane or ELISA plate, improving the sensitivity of detection by 8-fold. The LSAB method is used when the avidin-biotin-enzyme complex in the ABC method becomes too large to penetrate the tissue.

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NEFH was detected in paraffin-embedded sections of rat brain tissues using mouse anti-NEFH Antigen Affinity purified monoclonal antibody (Catalog # MA1071) at 1 μg/mL. Biotin Conjugated goat anti-mouse IgG (H+L) secondary antibody (Catalog # BA1001) was used to detect the primary antibody at 10μg/mL.

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PCNA was detected in paraffin-embedded sections of human intestinal cancer tissues using mouse anti-PCNA Antigen Affinity purified monoclonal antibody (Catalog # MA1083) at 1 μg/mL. Biotin Conjugated goat anti-mouse IgG (H+L) secondary antibody (Catalog # BA1001) was used to detect the primary antibody at 10μg/mL.