title: 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.
abstract: 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.
  
  
  
  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.
  
  
  
  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.
  
  
  
  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.
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA. Electronic address: Eric_Dixon@BD.com.
authors.forename: Eric P
authors.initials: EP
authors.lastname: Dixon
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Lorraine M
authors.initials: LM
authors.lastname: King
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Ramona
authors.initials: R
authors.lastname: Nelson
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Simkins
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Steven L
authors.initials: SL
authors.lastname: Knapp
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: George H
authors.initials: GH
authors.lastname: Brough
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Karen L
authors.initials: KL
authors.lastname: Lenz
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Dorian T
authors.initials: DT
authors.lastname: Henderson
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Clark M
authors.initials: CM
authors.lastname: Whitehead
authors.affiliation: Solstas Lab Partners, Greensboro, NC, USA.
authors.forename: Janice
authors.initials: J
authors.lastname: Hessling
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Charlotte A
authors.initials: CA
authors.lastname: Brown
authors.affiliation: BD Diagnostics - Women's Health and Cancer, Durham, NC, USA.
authors.forename: Douglas P
authors.initials: DP
authors.lastname: Malinowski
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology*; D000951:Antigens, Neoplasm / Q000276:immunology*; D001706:Biopsy; D015153:Blotting, Western; D002467:Cell Nucleus / Q000201:enzymology / Q000276:immunology / Q000473:pathology; D002584:Cervix Uteri / Q000201:enzymology / Q000276:immunology* / Q000473:pathology; D004250:DNA Topoisomerases, Type II / Q000276:immunology*; D004268:DNA-Binding Proteins / Q000276:immunology*; D018604:Epitope Mapping / Q000379:methods; D000939:Epitopes; D005260:Female; D006801:Humans; D007150:Immunohistochemistry; D064111:Minichromosome Maintenance Complex Component 2 / Q000276:immunology*; D000075223:Poly-ADP-Ribose Binding Proteins; D011237:Predictive Value of Tests; D016196:S Phase; D012720:Severity of Illness Index; D046888:Tissue Array Analysis / Q000379:methods*; D002578:Uterine Cervical Dysplasia / Q000175:diagnosis* / Q000201:enzymology / Q000276:immunology / Q000473:pathology; D002583:Uterine Cervical Neoplasms / Q000175:diagnosis* / Q000201:enzymology / Q000276:immunology / Q000473:pathology
chemical_list: D000911:Antibodies, Monoclonal; D000951:Antigens, Neoplasm; D004268:DNA-Binding Proteins; D000939:Epitopes; D000075223:Poly-ADP-Ribose Binding Proteins; C084859:MCM2 protein, human; D064111:Minichromosome Maintenance Complex Component 2; D004250:DNA Topoisomerases, Type II; C000618811:TOP2A protein, human
keywords: MCM2; ProEx™ C; SurePath™ liquid-based cytology; TOP2A