Abstract Background The Atellica® CI Analyzer is an automated, high-throughput integrated chemistry and immunoassay analyzer utilizing both Atellica® CH and Atellica® IM Assays. This study evaluated the analytical performance of the Atellica IM breast cancer antigen CA 15-3™ (CA 15-3) and the ovarian cancer antigen CA 125II™ (CA 125II) Assays on the Atellica CI Analyzer. Methods Precision studies were performed according to CLSI EP05-A3 using native and contrived human serum samples as well as contrived horse serum samples for CA 15-3. One aliquot of each sample pool was tested in duplicate in two runs per day ≥2 hours apart on each analyzer for ≥20 days. Precision studies were evaluated with one reagent lot on two systems. Method comparison studies were performed using three reagent lots according to CLSI EP09c. Individual native and contrived human serum samples were analyzed using the Atellica IM CA 15-3 and CA 125II Assays on the Atellica IM Analyzer and the Atellica CI Analyzer. LoB, LoD, and LoQ were determined as described in CLSI EP17-A2. Results Representative precision and method comparison results for each assay are listed in the table. For the two assays tested, repeatability and within-lab %CVs were <2.4% and <6.9%, respectively. Slopes determined by the weighted Deming linear regression model were approximately 1. Detection capability for the Atellica IM CA 15-3 Assay on the Atellica CI Analyzer was estimated to be 1.0 U/mL, 2.0 U/mL and 3.0 U/mL for LoB, LoD, and LoQ, respectively. When using the Atellica IM CA 125II Assay, detection capability was estimated to be 2.0 U/mL, 3.0 U/mL and 3.0 U/mL for LoB, LoD, and LoQ, respectively. Conclusions Evaluation of the Atellica IM CA 15-3 and CA 125II Assays using the Atellica CI Analyzer demonstrated good precision and equivalent performance compared to the same assays on the Atellica IM Analyzer.