Journal of Pathology and Translational Medicine

Original Articles

Multistaining Optimization for Epstein-Barr Virus–Encoded RNA In Situ Hybridization and Immunohistochemistry of Formalin-Fixed Paraffin-Embedded Tissues Using an Automated Immunostainer: Jae Nam Ko, Jin Kyoung Jung, Yun Ik Park, Hwa Jeong Shin, Jooryung Huh, Sol Back, Yu Jin Kim, Jae Ho Kim, Heounjeong Go; J Pathol Transl Med. 2019;53(5):317-326. Published online August 27, 2019; DOI: https://doi.org/10.4132/jptm.2019.08.06

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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.

Citations

Citations to this article as recorded by

Ultra High-plex Spatial Proteogenomic Investigation of Giant Cell Glioblastoma Multiforme Immune Infiltrates Reveals Distinct Protein and RNA Expression Profiles
Shilah A. Bonnett, Alyssa B. Rosenbloom, Giang T. Ong, Mark Conner, Aric B.E. Rininger, Daniel Newhouse, Felicia New, Chi Q. Phan, Saskia Ilcisin, Hiromi Sato, John S. Lyssand, Gary Geiss, Joseph M. Beechem
Cancer Research Communications.2023; 3(5): 763. CrossRef
Detection of Epstein–Barr Virus in Periodontitis: A Review of Methodological Approaches
Lilit Tonoyan, Marlène Chevalier, Séverine Vincent-Bugnas, Robert Marsault, Alain Doglio
Microorganisms.2020; 9(1): 72. CrossRef

Prognostic Significance of CD109 Expression in Patients with Ovarian Epithelial Cancer: So Young Kim, Kyung Un Choi, Chungsu Hwang, Hyung Jung Lee, Jung Hee Lee, Dong Hoon Shin, Jee Yeon Kim, Mee Young Sol, Jae Ho Kim, Ki Hyung Kim, Dong Soo Suh, Byung Su Kwon; J Pathol Transl Med. 2019;53(4):244-252. Published online May 2, 2019; DOI: https://doi.org/10.4132/jptm.2019.04.16

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Abstract PDF

Background
Ovarian epithelial cancer (OEC) is the second-most common gynecologic malignancy. CD109 expression is elevated in human tumor cell lines and carcinomas. A previous study showed that CD109 expression is elevated in human tumor cell lines and CD109 plays a role in cancer progression. Therefore, this study aimed to determine whether CD109 is expressed in OEC and can be useful in predicting the prognosis.
Methods
Immunohistochemical staining for CD109 and reverse transcription-quantitative polymerase chain reaction was performed. Then we compared CD109 expression and chemoresistance, overall survival, and recurrence-free survival of OEC patients. Chemoresistance was evaluated by dividing into good-response group and poor-response group by the time to recurrence after chemotherapy.
Results
CD109 expression was associated with overall survival (p = .020), but not recurrence-free survival (p = .290). CD109 expression was not an independent risk factor for overall survival due to its reliability (hazard ratio, 1.58; p = .160; 95% confidence interval, 0.82 to 3.05), although we found that CD109 positivity was related to chemoresistance. The poor-response group showed higher rates of CD109 expression than the good-response group (93.8% vs 66.7%, p = .047). Also, the CD109 mRNA expression level was 2.88 times higher in the poor-response group as compared to the good-response group (p = .001).
Conclusions
Examining the CD109 expression in patients with OEC may be helpful in predicting survival and chemotherapeutic effect.

Citations

Citations to this article as recorded by

CD109 Promotes Drug Resistance in A2780 Ovarian Cancer Cells by Regulating the STAT3-NOTCH1 Signaling Axis
Jun Se Kim, Min Joo Shin, Seo Yul Lee, Dae Kyoung Kim, Kyung-Un Choi, Dong-Soo Suh, Dayea Kim, Jae Ho Kim
International Journal of Molecular Sciences.2023; 24(12): 10306. CrossRef
CD109 facilitates progression and 5-fluorouracil resistance of nasopharyngeal carcinoma
Zhenwei Zhu, Fang Zhou, Cheng Mao
Materials Express.2022; 12(9): 1189. CrossRef
Usefulness of CD109 expression as a prognostic biomarker in patients with cancer
Hyun Min Koh, Hyun Ju Lee, Dong Chul Kim
Medicine.2021; 100(11): e25006. CrossRef
Serum CD109 levels reflect the node metastasis status in head and neck squamous cell carcinoma
Sumitaka Hagiwara, Eiichi Sasaki, Yasuhisa Hasegawa, Hidenori Suzuki, Daisuke Nishikawa, Shintaro Beppu, Hoshino Terada, Michi Sawabe, Masahide Takahashi, Nobuhiro Hanai
Cancer Medicine.2021; 10(4): 1335. CrossRef

Expression of Phospholipase C-gamma1 and gamma2 in Non-Hodgkin's and Hodgkin's Lymphoma.: Dae Woon Eom, Sung Sook Kim, Yeong Ju Woo, Jae Hee Suh, Jooryung Huh, Ae Ran Paik, Jae Ho Kim, Sung Ho Ryu, Pann Ghill Suh; Korean J Pathol. 2000;34(2):113-118.

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Abstract PDF: Phospholipase C (PLC) plays a role in ligand-mediated signal transduction for cellular activity such as proliferation and differentiation. A recent observation that PLC- gamma1 is highly expressed in some kinds of human cancer tissue supports the view that PLC-gamma1 may be involved in proliferation and carcinogenesis. PLC-gamma2 is known to be involved in B cell differentiation and maturation. However, there have been few studies about the expressions of PLC-gamma1 and gamma2 in human lymphoid malignancy. In the present study, we examined the contents of PLC-gamma1 and gamma2 in 10 cases of B cell, 10 cases of T cell non-Hodgkin's lymphoma and 5 cases of Hodgkin's lymphoma to find out whether these enzymes play any role in the carcinogenesis by immunohistochemistry and immunoprecipitation. Immunoprecipitation analysis revealed that in contrast to increased expression of PLC-gamma2 only in B cell lymphoma, a considerably higher level of PLC-gamma1 was detected in both B and T cell lymphoma. Immunohistochemical finding confirmed this observation. PLC-gamma1 and PLC-gamma2 were expressed in the cytoplasm of most tumor cells. PLC-gamma2 was also expressed in mature B cells, while PLC-gamma1 was not expressed in reactive non-tumor cells. These results suggest that PLC-gamma1 mediated signal transduction implicates a significant role in the carcinogenesis of all types of lymphoid tissue, and PLC-gamma2 may play a role in the carcinogenesis of B cell lymphoma as well as B cell differentiation.

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