Biomedical Engineering Reference
In-Depth Information
3. Protein localization by immunohistochemistry can be visualized by
conventional microscopy, l uorescence microscopy, confocal microscopy
or any other method that can visualize the dye used.
4. Dif erent proteins can be visualized simultaneously by using dif erent
colored dyes for the secondary antibodies.
Definition of Terms
Antibody:
A protein that reacts with or binds to another protein called an antigen.
Antigen:
A protein that attracts and binds with an antibody.
Avidin:
A glycoprotein that has a very strong ai nity for the protein biotin, so there is a
strong binding reaction between the streptavidin and the secondary antibody.
Avidin-biotin blocker:
A chemical that blocks any background biotin in the tissue so it will
not react with the streptavidin step of the process.
Background Sniper:
A casein and protein mixture that is required to reduce nonspecii c
protein background staining.
Biotin:
A B-vitamin that has a strong binding ai nity for avidin, thus allowing for a strong
attachment between streptavidin and a secondary protein.
Conjugated streptavidin:
A compound containing streptavidin and a horseradish peroxidase
tightly bound together
Endogenous:
Normally found within the tissue.
Epitope retrieval:
Exposure of the protein or portion of protein of interest so as to make it
available to bind with the antibody.
Fluorescent dye:
A dye that is visible under l uorescent light.
Horseradish peroxidase:
An enzyme attached to streptavidin so that it will more readily bind
to the dye and antibody.
Parai n-embedded sections:
Sections in which all water has been replaced with wax or
parai n.
Peroxidase:
An enzyme commonly found in mammalian tissues that could bind to
the streptavidin as background, making specii c detection of a particular protein
impossible.
Primary antibody:
h e antibody that attaches to the specii c protein that is being sought.
Secondary antibody:
h e antibody that attaches to a streptavidin-dye complex and to the
primary antibody.
References
Cadigan, K.M. and R. Nusse. 1997. Wnt signaling: a common theme in animal development.
Genes Dev. 11: 3286-3305.
Chen, G. and A.D. Akintola, J.M. Catania, M.D. Covington, D.D. Dean, J.P. Trzeciakowski,
R.C. Burghardt, and A.R. Parrish. 2007. Ischemia-induced cleavage of cadherins in
NRK cells is not sui cient for β-catenin transcriptional activity. Cell Comm. Adhesion
14: 111-123.
