Generation and characterization of a monoclonal antibody against human BCL6 for immunohistochemical diagnosis

Kunzhi Jia; Danping Zhang; Yanghai Wang; Yaju Liu; Xiangzhu Kong; Qinghai Yang; Huiling Chen; Chengjie Xie; Shihua Wang

doi:10.1371/journal.pone.0216470

Generation and characterization of a monoclonal antibody against human BCL6 for immunohistochemical diagnosis

PLoS One. 2019 May 7;14(5):e0216470. doi: 10.1371/journal.pone.0216470. eCollection 2019.

Authors

Kunzhi Jia^{1

2}, Danping Zhang², Yanghai Wang³, Yaju Liu¹, Xiangzhu Kong², Qinghai Yang⁴, Huiling Chen⁴, Chengjie Xie², Shihua Wang^{1

2}

Affiliations

¹ Key Laboratory of Pathogenic Fungi and Mycotoxins of Fujian Province, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
² Key Laboratory of Biopesticide and Chemical Biology of Education Ministry, School of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
³ Fuzhou No.1 High School, Fuzhou, China.
⁴ Fuzhou Maixin Biotech. Co., Ltd, Fuzhou, China.

Abstract

Background: Human B-cell lymphoma 6 (BCL6) gene, usually coding protein of 706 amino acids, is closely associated with large B cell lymphoma. Researches showed that protein mutation or change of expression levels usually happened in the mounting non-hodgkin lymphoma (NHL). Thus BCL6 is considered to be involved in germinal center (GC)-derived lymphoma.

Results: The BCL61-350 gene codons were optimized for prokaryotic system. After expression of BCL61-350 in E. coli, the BCL61-350 protein was purified with Ni column. Then the BCL61-350 protein, mixing with QuickAntibody-Mouse5W adjuvant, was injected into Balb/c mice. After immunization and cell fusion, a stable cell line named 1E6A4, which can secrete anti-BCL6 antibody, was obtained. The isotype of 1E6A4 mAb was determined as IgG2a, and the affinity constant reached 5.12×1010 L/mol. Furthermore, the specificity of the mAb was determined with ELISA, western blot and immunohistochemistry. Results indicated that the 1E6A4 mAb was able to detect BCL6 specifically and sensitively.

Conclusions: BCL61-350 antigen has been successfully generated with an effective and feasible method, and a highly specific antibody named 1E6A4 against BCL6 has been screened and characterized in this study, which was valuable in clinical diagnosis.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Antibodies, Monoclonal, Murine-Derived* / chemistry
Antibodies, Monoclonal, Murine-Derived* / immunology
Enzyme-Linked Immunosorbent Assay
Female
Germinal Center / immunology
Germinal Center / metabolism
Germinal Center / pathology
Humans
Immunoglobulin G* / chemistry
Immunoglobulin G* / immunology
Immunohistochemistry
Lymphoma, B-Cell / diagnosis*
Lymphoma, B-Cell / immunology*
Lymphoma, B-Cell / metabolism
Lymphoma, B-Cell / pathology
Mice
Mice, Inbred BALB C
Proto-Oncogene Proteins c-bcl-6 / biosynthesis
Proto-Oncogene Proteins c-bcl-6 / immunology*

Substances

Antibodies, Monoclonal, Murine-Derived
BCL6 protein, human
Immunoglobulin G
Proto-Oncogene Proteins c-bcl-6

Grants and funding

This work was supported by the Science and Technology Project in Fujian Province (2016Y0001, 2015J05052), the project of Fujian marine and fishery department (Fujia marine hi-tech [2015] 26), and Education and research projects for young teachers in Fujian provincial Education Hall (JT180137). Qinghai Yang and Huiling Chen are employed by Fuzhou Maixin Biotech. Co., Ltd. Fuzhou Maixin Biotech. Co., Ltd provided support in the form of salaries for authors QY,HC, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors is articulated in the ‘author contributions’ section.