OBJECTIVE: Although HER2 has gradually become an important therapeutic target for colorectal cancer (CRC), a unified and standard HER2 scoring system was still not established in CRC, and the debatable results of immunohistochemistry and fluorescence in situ hybridization (FISH) in CRC requires further exploration. METHODS: In this study, we use five immunohistochemical (IHC) scoring criteria (i.e., IRS-p, IRS-m, GEA-s, GEA-b and HERACLES) and two FISH criteria to evaluate HER2 status, and further evaluate the correlation between HER2 status and clinicopathological features, survival in a large, unselected Chinese cohort of 664 CRCs. RESULTS: Finally, we set HER2/CEP17 ratio ≥ 2.0, or an average HER2 copy number ≥ 6.0 as FISH-positive threshold and the amplification rate of HER2 gene was 7.08% (47/664).The HER2 positivity (IHC 3+) was 2.71%, 3.16%, 2.56%, 2.71% and 3.16%, according to the IHC scoring criteria of IRS-p, IRS-m, GEA-s, GEA-b and HERACLES, respectively. Set FISH results as the golden standard; receiver-operating characteristic analysis showed that IRS-p had both high sensitivity and specificity than other IHC scoring systems to evaluate HER2 status. Based on IRS-p criterion, There were significant differences in tumor differentiation (p = 0.038), lymphatic vascular invasion (p = 0.001), pN stage (p value = 0.043), and overall survival (p < 0.001) among IHC score 0-3 + groups. Meanwhile, there were significant differences in pT stage (p = 0.031), pN stage (p = 0.009) and overall survival (p < 0.001) among FISH subgroups. CONCLUSION: The IRS-p criterion was more suitable for assessing the HER2 status in CRC patients than other IHC criteria. Whereas for FISH scoring system, only HER2/CEP17 < 2.0, meanwhile HER2cn < 4.0 and HER2cn ≥ 6.0 were subgroups with unique clinicopathological characteristics.