'''Immunohistochemistry''' is the process of detection of
antigens in tissue using
antibodies. The antibodies can be polyclonal or monoclonal in origin, the monoclonal ones being more specific in nature. Immunohistochemistry is widely used for diagnosis of
cancers. Specific markers are known for various cancers.
Methods
There are basically two methods for immmunohistochemical demonstration of
antigens in the
tissue, the
direct and the
indirect method.
The "direct method" is a one-step staining method, and involves a labeled
antibody (i.e. FITC conjugated antiserum) reacting directly with the
antigen in
tissue sections. This technique utilizes only one
antibody and the procedure is therefore short and quick. However, it is insensitive due to little signal amplification and rarely used since the introduction of indirect method.
The "indirect method" involves an unlabeled primary antibody (first layer) which reacts with
tissue antigen, and a labeled secondary antibody (second layer) reacts with primary antibody. (The secondary antibody must be against the IgG of the animal species in which the primary antibody has been raised.) This method is more sensitive due to signal amplification through several secondary antibody reactions with different antigenic sites on the primary antibody. The second layer antibody can be labeled with a fluorescent dye such as FITC, rhodamine or Texas red, and this is called indirect
immunofluorescence method. The second layer antibody may be labeled with an
enzyme such as
peroxidase,
alkaline phosphatase or
glucose oxidase, and this is called indirect immunoenzyme method. Offering a substrate to these enzymes will demonstrate presence of the enzyme (and hence the antibody) in a quantifiable fashion.
External links
Category:Anatomical pathology