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What is the difference between primary and secondary antibodies?
Posted July 27, 2019
Answer
The difference between primary and secondary antibodies is that secondary antibodies are raised against the species and isotype of the primary antibody. For example, if your primary antibody was raised in mouse, then you will need an anti-mouse secondary antibody raised in a host species other than mouse (e.g. goat anti-mouse IgG). This minimizes non-specific binding that leads to high background interference and false positives.
Advantage of Using Secondary Antibodies
The secondary antibody, which is generally conjugated to a fluorophore, enzyme or biotin, detects and binds to the primary antibody and not to any antigen present in the specimen. The advantage of using secodnary antibodies is the significant imporvement in sensitivity due to the resulting signal amplification from multiple secondary antibodies binding to a single primary antibody.
Applications for Secondary Antibody Conjugates
- Immunofluorescence (IF)
- Immunocytochemistry (ICC)
- Immunohistochemistry (IHC)
- Flow Cytometry
- ELISA
- Western Blot