Direct Method

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The page below is a sample from the LabCE course Immunohistochemistry (IHC) Basics in Histology. Access the complete course and earn ASCLS P.A.C.E.-approved continuing education credits by subscribing online.

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Direct Method

The direct method of IHC staining is a one-step method. This method consists of using a primary antibody that is conjugated directly with a label, such as fluorescein isothiocyanate (FITC) conjugated antiserum, enzyme, or alkaline phosphate. This labeled primary antibody reacts directly with a specific antigen in tissue sections.
A FITC labeled antibody is most commonly used in clinical applications of the direct method. Direct immunofluorescence staining employing FITC labeled antibodies is commonly performed on kidney biopsy specimens and skin biopsy specimens.
A drawback to the direct method of IHC staining is that it is less sensitive than indirect methods of staining. Weak signals can also be observed since the method excludes signal amplification actions. Additionally, when a FITC-labeled primary antibody is used, it will compete with the autofluorescence exhibited naturally by componets of the tissue. This can complicate microscopic evaluation and affect final diagnoses.