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Because HER-2 expression in invasive carcinoma of the breast has well-documented ramifications for treatment and prognosis, accurate assessment of HER-2 status is critical. Comparative studies have shown high concordance rates between immunohistochemical analysis and fluorescence in situ hybridization (FISH) in cases with immunohistochemical scores of 0 or 1+ (negative) and 3+ (strongly positive) and low concordance rates among cases with immunohistochemical scores of 2+. The present study was performed to determine concordance rates in a setting more representative of routine clinical practice, in which multiple pathologists submit specimens to a single cytogenetics referral laboratory. We found a higher rate of discordance between immunohistochemical analysis and FISH (approximately 92%) in the groups with immunohistochemical scores of 2+ than reported in other studies. These results strongly support the practice of performing FISH in all cases with immunohistochemical scores of 2+, particularly in routine practice, in which interobserver variability in immunohistochemical scoring among multiple pathologists is likely to be high.