Characterization and clinical validation of MCM2 and TOP2A monoclonal antibodies in the BD ProEx™ C assay: An immunoassay which detects aberrant S-phase induction in cervical tissue
| Autor: | Stephen G. Simkins, Douglas P. Malinowski, Steven L. Knapp, Lorraine M. King, George H. Brough, Clark M. Whitehead, Karen L. Lenz, Dorian Henderson, Eric P. Dixon, Janice J. Hessling, Charlotte A. Brown, Ramona Nelson |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Pathology medicine.medical_specialty medicine.drug_class Biopsy Blotting Western Immunology Uterine Cervical Neoplasms Cervix Uteri Monoclonal antibody Severity of Illness Index Epitope S Phase Epitopes 03 medical and health sciences 0302 clinical medicine Antigens Neoplasm Predictive Value of Tests Uterine Cervical Dysplasia medicine Humans Immunology and Allergy Poly-ADP-Ribose Binding Proteins Cell Nucleus Tissue microarray biology Antibodies Monoclonal Minichromosome Maintenance Complex Component 2 Immunohistochemistry Molecular biology Staining DNA-Binding Proteins DNA Topoisomerases Type II 030104 developmental biology Epitope mapping Tissue Array Analysis 030220 oncology & carcinogenesis biology.protein Female Antibody Epitope Mapping |
| Zdroj: | Journal of Immunological Methods. 442:35-41 |
| ISSN: | 0022-1759 |
| DOI: | 10.1016/j.jim.2017.01.002 |
| Popis: | Background The Papanicolaou (Pap) screen has been successful in reducing cervical cancer; but exhibits low sensitivity when detecting cervical dysplasia. Use of molecular biomarkers in Pap tests may improve diagnostic accuracy. Design Monoclonal antibodies to Minichromosome Maintenance Protein 2 (MCM2) and DNA Topoisomerase II α (TOP2A) were selected for use in IHC based on their ability to differentiate normal from diseased cervical tissues in tissue microarrays. Enhanced Green Fluorescent Protein Western blot analysis was used to help identify binding epitopes specific to MCM2 and TOP2A antibody clones. Antibody affinity was determined by solution phase affinity measurement and immunohistochemistry was performed using high affinity MCM2 or TOP2A antibodies on serial histological sections. Results Antibody clones to MCM2 and TOP2A clones were selected based on their ability to detect over expression in abnormal cervical epithelia. In IHC, MCM2-27C5.6 and MCM2-26H6.19 demonstrated superior staining in abnormal cervical tissue over the MCM2-CRCT2.1 antibody. A combination of MCM2 and TOP2A antibodies showed greater staining when compared to staining with any of the antibodies alone on serial histological sections. Distinct linear epitopes were elucidated for each of the MCM2 and TOP2A clones. Affinity values (Kd) for MCM2 or TOP2A antibodies had a similar range. In a research study, the MCM2 and TOP2A (BD ProEx™ C) antibody cocktail showed increased epithelia staining with increasing dysplasia. The use of BD ProEx™ C in combination with H&E staining enhanced immunohistochemical discrimination of dysplastic and non-dysplastic FFPE cervical tissue specimens. Conclusions BD ProEx™ C containing MCM2 and TOP2A antibodies showed strong specific nuclear staining that correlated with increased dysplasia and lesion severity. Enhanced performance of the antibodies was linked to their unique topography recognition. BD ProEx™ C incorporates antibodies that enhance detection of CIN2 + cervical disease. |
| Databáze: | OpenAIRE |
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