Table 2HER2 assays used in tissue specimens and serum: clinical trials, clinical practice, and under development (adapted with permission from the American Society of Clinical Oncology; Wolff, Hammond, Schwartz, et al., 2007a and including information from Carlson, Moench, Hammond, et al., 2006)

A. IHC Assays: measure HER2 protein overexpression in tissue
AssayMfrMethodologyScoring CriteriaFDA Status
Clinical Trials AssayDeveloped by independent laboratoryCB11 and 4D5 MAb0 and 1+ negative, 2+ weakly positive, 3+ strongly positiveResearch assay used in trials of trastuzumab in metastatic breast cancer
HercepTest™DAKO*A0485 polyclonal antibodyWeakly positive (2+): weak to moderate complete membrane staining in >10% of tumor cells; strongly positive (3+): strong complete membrane staining in >10% of tumor cells*U.S. Food and Drug Administration (FDA) approved as an aid in the assessment of patients for whom Herceptin™ (trastuzumab) treatment is being considered
PATHWAY™Ventana†CB11 MAbPositive (2+): weak complete staining of the membrane, >10% of cancer cells; positive (3+): intense complete staining of the membrane, >10% of cancer cells†FDA approved as an aid in the assessment of patients for whom Herceptin™ (trastuzumab) treatment is being considered
B. In-Situ Hybridization (ISH) Assays: measure HER2 gene amplification in tissue
PathVysion® HER2 DNA Probe Kit (FISH)Abbott‡Hybridization of fluorescent DNA probes to HER2 gene (orange) and chromosome 17 centromere (green)HER2 amplification: HER2/CEP17 ratio ≥2 on average for 60 cells; results at or near the cut off point (1.8–2.2) should be interpreted with caution (Persons, Tubbs, Cooley, et al., 2006; Dal Lago, Durbecq, Desmedt, et al., 2006)FDA approved as an aid in the assessment of patients for whom Herceptin™ (trastuzumab) treatment is being considered
INFORM HER2/neu Probe (FISH)Ventana§Hybridization of biotin-labeled DNA probe to HER2 gene and fluorescently labeled avidinHER2 amplification: average of >6 HER2 gene copies/nucleus; an average of >4.0 <6.0 gene copies/nucleus for 60 cells described as equivocal in one publication (Dal Lago, Durbecq, Desmedt, et al., 2006; Vera-Roman and Rubio-Martinez, 2004)FDA approved as an adjunct to existing clinical and pathologic information currently used as prognostic indicators in the risk stratification of breast cancer in patients with a primary, invasive, localized, node-negative tumor
HER2 FISH pharmDx™ KitDako▿Hybridization of fluorescent DNA probes to HER2 gene (red) and PNA probes to chromosome 17 centromere (CEN-17; green)Count 20 nuclei per tissue specimen, when possible from distinct tumor areas. Specimens with a HER2/CEN-17 ratio ≥2 should be considered HER2 gene amplified (Kallioniemi, Kallioniemi, Kurisu, et al., 1992; Ellis, Dowsett, Bartlett, et al., 2000; Hanna, 2001; Tsuda, Akiyama, Terasaki, et al., 2001). Results at or near the cut-off (1.8–2.2) should be interpreted with caution. If the ratio is borderline (1.8–2.2), count an additional 20 nuclei and recalculate the ratio for the 40 nucleiFDA approved as an adjunct to clinicopathologic information currently used for estimating prognosis in stage II, node-positive breast cancer patients and as an aid in assessment of patients being considered for Herceptin™ (trastuzumab) treatment
SPoT-Light (CISH)Invitrogen/Zymed¶Hybridization of digoxigenin-labeled DNA probe to HER2 gene; detection via mouse antidigoxigenin antibody followed by antimouse-peroxidaseHigh HER2 amplification defined as >10 dots, or large clusters, (low if >5 dots to 10 dots, or small clusters) or mixture of multiple dots and large clusters of the HER2 gene present per nucleus in >50% tumor cells (Hanna and Kwok, 2006)DNA probe kit not available in the U.S.
EnzMet GenePro (SISH)VentanaHybridization of dinitrophenol-labeled DNA probe to HER2 gene; detection via peroxidase-labeled multimer followed by enzyme metallographyAmplification defined as six or more dots, or large clusters of dots, in 30% or more of invasive tumor cells (Downs-Kelly, Pettay, Hicks, et al., 2005)DNA probe kit not available in the U.S.
C. HER2 Extracellular Domain (ECD) Assays: detect HER2 ECD in serum
Immuno 1®/ADVIA Centaur®BayerEnzyme immunoassay (EIA); primary MAbs NB-3 and TA-1 (one is labeled with fluorescein and the other is either linked to an enzyme or a chemiluminogenic molecule) specific for the ECD of HER2 added to sera; detection via binding of immunocomplex to antifluorescein antibodies in the solid phase, followed by addition of substrate in case of Immuno 1 assayElevated ECD concentrations often defined as >15 ng/mL (Payne, Allard, Anderson-Mauser, et al., 2000; Esteva, Cheli, Fritsche, et al., 2005)FDA approval for followup and monitoring patients with metastatic breast cancer only

From: 1, Introduction

Cover of HER2 Testing to Manage Patients With Breast Cancer or Other Solid Tumors
HER2 Testing to Manage Patients With Breast Cancer or Other Solid Tumors.
Evidence Reports/Technology Assessments, No. 172.
Seidenfeld J, Samson DJ, Rothenberg BM, et al.

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