The direct method of IHC staining is a one-step method. This method uses a primary antibody 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.
FITC-labeled antibodies are most commonly used in clinical applications of the direct method. Direct immunofluorescence staining employing FITC-labeled antibodies is frequently performed on kidney and skin biopsy specimens.
A drawback of direct IHC staining is that it is less sensitive than indirect methods. 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 tissue components. This can complicate microscopic evaluation and affect final diagnoses.
