title: Her2 immunohistochemical evaluation by traditional microscopy and by digital analysis, and the consequences for FISH testing.
abstract: OBJECTIVE
  Her2 protein is the key marker determining the choice of Herceptin therapy after a diagnosis of breast cancer. Its evaluation is made in most laboratories by immunohistochemistry, and interpreted by a pathologist using an optical microscope, a process subject to inter-observer variability, particularly for samples scored as equivocal (2+). Software analysis products have been introduced, seeking to reduce this variability. In this study, we compared the results of both traditional evaluation and a specific software package (VISIA Imaging) to those from fluorescent in situ hybridization (FISH).
  
  
  METHODS
  We selected 176 cases of invasive breast cancer sampled during 2012-2014 that were classified as equivocal after evaluation of Her2 immunohistochemistry, and that were also evaluated by FISH. Each tissue slide was scanned with a digital D-Sight Fluo 2.0 microscope and analysed with VISIA Imaging S.r.l. software. The final results were categorised as follows: negative (0-1+), equivocal (2+), or positive (3+). Then each result was compared to that obtained by FISH.
  
  
  RESULTS
  The digital method confirmed 85 samples (48.3%) as equivocal (2+), while 23 (15.1%) were reclassified as negative (1+) and 44 (28.9%) as positive (3+). Of the 176 cases, 24 (13.6%) were not suitable for digital analysis (inadequate). Of 67 reclassified cases (1+ or 3+), 62 were in agreement with FISH results (concordance rate 92.5%). The sensitivity and specificity of the digital method were 100% and 82%, respectively.
  
  
  CONCLUSIONS
  The application of this analysis software led to an improvement in the interpretation of cases classified as equivocal, decreasing the need for FISH and increasing diagnostic certainty.
authors.affiliation: Department of Medical, Surgical and Health Science - University of Trieste, Strada di Fiume 447, 34149 Trieste, Italy.
authors.forename: Thomas
authors.initials: T
authors.lastname: Marcuzzo
authors.affiliation: Department of Medical, Surgical and Health Science - University of Trieste, Strada di Fiume 447, 34149 Trieste, Italy. Electronic address: fgiudici@units.it.
authors.forename: Fabiola
authors.initials: F
authors.lastname: Giudici
authors.affiliation: UCO di Anatomia e Istologia Patologica - Azienda Sanitaria Universitaria Integrata di Trieste, Strada di Fiume 447, 34149 Trieste, Italy.
authors.forename: Elisa
authors.initials: E
authors.lastname: Ober
authors.affiliation: Department of Medical, Surgical and Health Science - University of Trieste, Strada di Fiume 447, 34149 Trieste, Italy; UCO di Anatomia e Istologia Patologica - Azienda Sanitaria Universitaria Integrata di Trieste, Strada di Fiume 447, 34149 Trieste, Italy.
authors.forename: Clara
authors.initials: C
authors.lastname: Rizzardi
authors.affiliation: Department of Medical, Surgical and Health Science - University of Trieste, Strada di Fiume 447, 34149 Trieste, Italy.
authors.forename: Cristina
authors.initials: C
authors.lastname: Bottin
authors.affiliation: Department of Medical, Surgical and Health Science - University of Trieste, Strada di Fiume 447, 34149 Trieste, Italy; UCO di Anatomia e Istologia Patologica - Azienda Sanitaria Universitaria Integrata di Trieste, Strada di Fiume 447, 34149 Trieste, Italy.
authors.forename: Fabrizio
authors.initials: F
authors.lastname: Zanconati
mesh_terms: D014408:Biomarkers, Tumor / Q000378:metabolism; D001943:Breast Neoplasms / Q000175:diagnosis* / Q000378:metabolism / Q000473:pathology; D018270:Carcinoma, Ductal, Breast / Q000175:diagnosis* / Q000378:metabolism / Q000473:pathology; D005260:Female; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods; D008853:Microscopy; D015588:Observer Variation; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000378:metabolism*
chemical_list: D014408:Biomarkers, Tumor; D018719:Erb-b2 Receptor Tyrosine Kinases; C508053:ERBB2 protein, human
keywords: Breast cancer; Digital analysis; Fluorescence; Her2; Immunohistochemistry