medric medric
[닫기]
잠시만 기다려 주세요. 로딩중입니다.

Multistaining Optimization for Epstein-Barr Virus?Encoded RNA In Situ Hybridization and Immunohistochemistry of Formalin-Fixed Paraffin-Embedded Tissues Using an Automated Immunostainer

Journal of Pathology and Translational Medicine 2019년 53권 5호 p.317 ~ 326
 ( Ko Jae-Nam ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology

 ( Jung Jin-Kyoung ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology
 ( Park Yun-Ik ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology
 ( Shin Hwa-Jeong ) - University of Ulsan College of Medicine Asan Medical Center Department of Research Support Team
 ( Huh Joo-Ryung ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology
 ( Back Sol ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology
 ( Kim Yu-Jin ) - University of Ulsan College of Medicine Asan Medical Center Department of Research Support Team
 ( Kim Jae-Ho ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology
 ( Go Heoun-Jeong ) - University of Ulsan College of Medicine Asan Medical Center Department of Pathology

Abstract


Background: Single staining is commonly performed for practical pathologic diagnoses. However, this method is limited in its ability to specify cellular morphology and immunophenotype and often requires consumption of limited tissue. This study aimed to describe an optimized protocol for multiple in situ hybridization (ISH) and immunohistochemistry (IHC).

Methods: The quality of multistaining was evaluated by carefully changing each step of ISH and IHC in an angioimmunoblastic T-cell lymphoma (AITL) case on a Ventana BenchMark XT automated immunostainer. The optimized protocols were also performed using another immunostainer and in 15 cases of five Epstein-Barr virus (EBV)?associated malignancies using formalin-fixed paraffin-embedded tissue.

Results: The quality of various ISHIHC staining protocols was semi-quantitatively evaluated. The best EBV-encoded RNA (EBER)-ISH/double IHC staining quality, equivalent to single staining, was obtained using the following considerations: initial EBER-ISH application, use of protease and antigen retrieval reagent (cell conditioning 1 [CC1] treatment time was minimized due to impact on tissue quality), additional baking/ deparaffinization not needed, and reduced dilution ratio and increased reaction time for primary antibody compared with single immunostaining. Furthermore, shorter second CC1 treatment time yielded better results. Multiple staining was the best quality in another immunostainer and for different types of EBV-associated malignancies when it was performed in the same manner as for the Ventana BenchMark XT as determined for AITL.

Conclusions: EBER-ISH and double IHC could be easily used in clinical practice with currently available automated immunostainers and adjustment of reagent treatment time, dilution ratio, and antibody reaction time.

키워드

Protocol optimization of multistaining; In situ hybridization; Immunohistochemistry; Epstein-Barr virus-associated malignancy; Formalin-fixed paraffin-embedded tissue
원문 및 링크아웃 정보
 
등재저널 정보
KCI