title: Optimal Scoring of Brightfield Dual-Color In Situ Hybridization for Evaluation of HER2 Amplification in Breast Carcinoma: How Many Cells Are Enough?
abstract: OBJECTIVE
  The purpose of this study was to determine the optimum number of cells that should be counted when scoring human epidermal growth factor receptor 2 (HER2) brightfield dual-color in situ hybridization (BDISH), including cases with HER2/chromosome 17 (Chr17) ratios in the 1.80 to 2.20 range.
  131 cases of breast carcinoma with HER2 immunohistochemistry and BDISH were included. For cases with a HER2/Chr17 ratio of less than 1.80 or more than 2.20 (n = 115), BDISH scoring was performed for 60 cells using three tumor fields, and for cases with a HER2/Chr17 ratio of 1.80 to 2.20 (n = 16), scoring was performed for 120 cells using six tumor fields. Mean HER2/Chr17 ratio and HER2 copy number were calculated for cumulative cell counts.
  
  
  RESULTS
  The HER2 status as determined by the HER2/Chr17 ratio or HER2 copy number was unchanged following counting of additional cells in 100% of cases with ratio of less than 1.80 or more than 2.20. The HER2 status of two cases with ratios of 1.80 to 2.20 changed from positive to negative following counting of 120 cells.
  
  
  CONCLUSIONS
  Our findings support recommendations to score 20 nuclei in conjunction with careful assessment of immunohistochemistry and scan of the BDISH slide to identify areas of heterogeneity. Scoring of additional cells/fields is likely not of benefit and might be a disadvantage since the scorer moves out of the area of strongest signal.
authors.affiliation: From the Department of Histopathology.
authors.forename: Susan
authors.initials: S
authors.lastname: Prendeville
authors.affiliation: Breast Cancer Research Centre, Cork University Hospital, Cork, Ireland.
authors.forename: Mark A
authors.initials: MA
authors.lastname: Corrigan
authors.affiliation: Breast Cancer Research Centre, Cork University Hospital, Cork, Ireland.
authors.forename: Vicki
authors.initials: V
authors.lastname: Livingstone
authors.affiliation: From the Department of Histopathology.
authors.forename: Linda
authors.initials: L
authors.lastname: Feeley
authors.affiliation: From the Department of Histopathology.
authors.forename: Michael W
authors.initials: MW
authors.lastname: Bennett
authors.affiliation: From the Department of Histopathology.
authors.forename: Tara Jane
authors.initials: TJ
authors.lastname: Browne
authors.affiliation: From the Department of Histopathology, fionnuala.oconnell@hse.ie.
authors.forename: Fionnuala
authors.initials: F
authors.lastname: O'Connell
mesh_terms: D014408:Biomarkers, Tumor / Q000235:genetics*; D001943:Breast Neoplasms / Q000175:diagnosis / Q000235:genetics*; D002452:Cell Count; D002467:Cell Nucleus / Q000235:genetics; D002886:Chromosomes, Human, Pair 17 / Q000235:genetics*; D015331:Cohort Studies; D056915:DNA Copy Number Variations; D005260:Female; D005784:Gene Amplification; D018740:Genetic Heterogeneity; D006801:Humans; D007150:Immunohistochemistry; D017403:In Situ Hybridization / Q000379:methods*; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000235:genetics*
chemical_list: D014408:Biomarkers, Tumor; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: Breast carcinoma; HER2; In situ hybridization