Comparing reverse phase high-performance liquid chromatography and enzyme-linked immunosorbent assay techniques for the quantification of κ-casein B in bovine milk

Among κ-CN variants in milk, κ-CN B is the most reactive to chymosin activity, enhancing the quality, profitability, and sustainability of the final cheese product. The aim of this study was to assess the agreement between reverse phase HPLC (RP-HPLC) reference method and a rapid ELISA technique for the quantification of κ-CN B in individual bovine milk samples. Chromatographic and immunoenzymatic analyses were performed on individual milk samples from 933 Brown Swiss cows, with κ-CN B expressed as (1) milligrams per milliliter of milk, (2) percentage of κ-CN B over total milk protein content, and (3) grams of κ-CN B yielded on a milking event. The agreement between κ-CN B phenotypes measured through RP-HPLC and ELISA was evaluated through r and z-scores. Results suggested a general agreement between the 2 techniques, with r ranging from 0.88 for κ-CN B expressed in milligrams per milliliter and as a percentage to 0.90 for κ-CN B expressed in grams. This is further supported by relatively low z-scores (<0.5), which suggested the absence of significant differences between the values obtained from RP-HPLC and ELISA. Observed discrepancies were likely because ELISA does not provide quantitative results for concentrations of κ-CN B >10 mg/mL, and to the limited sensitivity of the ELISA at low concentrations of κ-CN B. Overall, findings of the present study demonstrated a strong agreement between the 2 techniques.