Immunohistochemistry

Prostate Specific Antibodies

A glycoprotein enzyme encoded by the KLK3 gene and is secreted by epithelial cells in the prostate gland, present in small amounts in men with normal prostates. 

Elevated in the presence of prostate cancer. 

Monoclonal antibodies that target the different epitopes on the antigen will identify the molecular makeup of the tumour and guide therapy. 

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Horseradish Peroxidase

Naturally occurring enzyme from the root of the horse-radish plant used to label antibodies

Causes oxidation.  

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Alkaline Phosphatase

Enzyme found in calves intestine and catalyses the reaction of fuchsin substates. Used to label antibodies. 

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PAP method

Immunological sandwich amplification. 

Antiserum is harvested and placed in solution with the enzyme so that immune complexes form that remain soluble. 

The molecular ratio of two molecules of IgG to three molecules of the enzyme. 

Antiperoxidase antibodies are from the same species as that which has produced a primary antibody raised against tissue antigens

2 antibodies: one is directed against the tissue antigen. The other is directed against the peroxidase enzyme.   

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ABC

Avidin-biotin complex. 

Biotin can be conjugated to antibodies. 

The process involves 3 layers:

  • unlabelled primary antibody
  • biotinylated secondary antibody
  • avidin-biotin peroxidase complex. 

Produce different colorimetric end products. 

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ASP

Streptavidin is a substitution of avidin. The electrostatic binding to tissues is eliminated.  It has the same 3 layers as ABC. 

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Two Step Polymer Method

The secondary antibody is conjugated to a polymer. The synthetic peptide is coupled to an enzyme. This reduces the penetrative ability of the detection reagent. 

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