LAUSR

Graphical abstract Figure. No caption available. HighlightsLoading of protein A onto C‐CP fibers is an effective means of generating an efficient IgG capture phase.Quantification of IgG in CHO cell cultures can be affected using either standard calibration curves or standard additions.Intra‐ and inter‐column quantification performance shows a high level of reproducibility. Abstract Polypropylene (PP) capillary‐channeled polymer (C‐CP) fibers loaded with recombinant Staphyloccocus aureus protein A (rSPA) were used as an affinity chromatography stationary phase for the quantitation of immunoglobulin G (IgG) in complex biological matrices. Optimization of the chromatographic method regarding mobile phase components and load/elution conditions was performed. The six‐minute analysis, including a load step with 12 mM phosphate at pH 7.4, an elution step with 0.025% phosphoric acid and a re‐equilibration step, was employed for quantitation of IgG1 from 0.075 to 3.00 mg mL−1 in an IgG‐free CHO cell supernatant matrix. Quantification of IgG1 content in a different CHO cell line was accomplished using the external calibration curve as well as using a standard addition approach. The high level of agreement between the two approaches suggests that the protein A‐modified C‐CP fiber phase is immune from matrix effects due to concomitant species such as host cell proteins (HCPs), host cell DNA, media components and other leachables and extractables. The inter‐day and intra‐day precision of the method were 3.1 and 3.5%RSD respectively for a single column. Column‐to‐column variability was 1.31 and 6.62%RSD for elution time and peak area, respectively, across columns prepared in different batches. The method reported here is well‐suited for IgG analysis in complex harvest cell culture media in both the development and production environments.