Antibody Elution Method for Multiple Immunohistochemistry on Primary Antibodies Raised in the Same Species and of the Same Subtype

Author:

Pirici Daniel12,Mogoanta Laurentiu1,Kumar-Singh Samir2,Pirici Ionica3,Margaritescu Claudiu4,Simionescu Cristina4,Stanescu Radu3

Affiliation:

1. Department of Histology, University of Medicine and Pharmacy Craiova, Craiova, Romania

2. Neurodegenerative Brain Diseases Group, Vlaams Instituut voor Biotechnologie, Department of Molecular Genetics, Laboratory of Neurogenetics, Institute Born Bunge, University of Antwerp, Antwerp, Belgium

3. Emergency County Hospital 1, Craiova, Romania

4. Department of Pathology, University of Medicine and Pharmacy Craiova, Craiova, Romania

Abstract

Double or multiple antigen labeling in IHC classically relies on the existence of primary antibodies raised in different species or of different IgG isotypes to ensure the specific labeling with the secondary detection systems. However, suitable pairs of primary antibodies are not always available or the best choice (e.g., as diagnostic tools). During the last few years, several methods have been proposed to overcome this, but none of them offers the flexibility needed for reliable double or multiple enzymatic or fluorescent IHC. We present here a procedure that elutes the antibodies after a first round of immunolabeling, which, in combination with precipitation-based detection systems, allows multiple IHC rounds even for primary antibodies raised in the same species and IgG isotype. Compared with other proposed methods, this procedure ensures a reliable enzymatic or fluorescent staining without cross-reactivity and without loss of tissue antigenicity, thus offering a flexible tool for colocalization studies and pathological diagnosis. This manuscript contains online supplemental material at http://www.jhc.org . Please visit this article online to view these materials.

Publisher

SAGE Publications

Subject

Histology,Anatomy

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