journal of Histology & Histopathology

Journal of Histology & Histopathology

ISSN 2055-091X
Original Research

Standardization of concentrated antibodies for use in automated immunohistochemistry

Emily Ferreira Salles Pilar1* and Claudio Galleano Zettler2

*Correspondence: Emily Ferreira Salles Pilar epilar@hcpa.edu.br

1. Experimental Pathology Unit, Hospital de Clínicas de Porto Alegre (HCPA), Ramiro Barcelos, 2350 - Santa Cecilia, Porto Alegre - RS, 90035-007, Brazil.

Author Affiliations

2. Department of Pathology, Santa Casa de Misericórdia de Porto Alegre, Universidade Federal de Ciências da Saúde de Porto Alegre (UFCSPA), Sarmento Leite, 245 - Centro Histórico, Porto Alegre - RS, 90050-170,Brazil.

Abstract

Background: Immunohistochemistry (IHC) is a method of identifying proteins in cells or tissues that is useful for diagnosis and research. In modern pathology, it has assumed an important supporting role in the molecular diagnosis of certain neoplasia, with an exponential contribution in personalized medicine. Automation in immunohistochemistry contributes to reduced test variability through standardization. The transition to automation is a process, and for a laboratory that already has a collection of stockconcentrated primary antibodies, ideally, these antibodies will be compatible with the chosen automated method, as antibodies are of high value to be discarded as a consequence of migration to automated immunohistochemistry.

Methods: 78 concentrated antibodies were tested for use in Ventana Medical Systems’ Benchmark XT automation platform. Thirty-one human tissue samples containing the antigens of interest were used as positive control.

Results: All antibodies tested showed good performance, indicating the feasibility of using these antibodies concentrated in the automation platform in question. The protocol most frequently used was the one with antigenic retrieval with Cell Conditioning 1 for 60 minutes and incubation in the primary antibody for 32 minutes at 42ºC. The dilutions of the primary antibodies in automation ranged from 1:20 to 1:4000.

Conclusion: Under the aforementioned conditions, it was possible to take advantage of the portfolio of concentrated antibodies present in the laboratory at the time of transition from manual to automated immunohistochemistry.

Keywords: Automation, immunohistochemistry, standardization, concentrated antibodies

ISSN 2055-091X
Volume 7
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