Quantitative bottom up analysis of infliximab in serum using protein A purification and integrated μLC-electrospray chip IonKey MS/MS technology

Background: TNO Triskelion has applied its general workflow for the development of quantitative LC–MS methods for proteins in biological matrices to the quantification of infliximab in rat serum using bottom up μLC–MS/MS. Results/methodology: The general workflow consists of sample purification, analyte processing and LC–MS analysis. In the development of a quantitative μLC–MS/MS method for infliximab in rat serum the analyte processing part and the LC–MS part were optimized, in order to meet the different sample requirements of μLC–MS as compared with UPLC–MS. Using the optimized μLC–MS/MS method the LOQ was 75 ng/ml. Conclusion: The present study showed that it is possible to gain sensitivity when going to smaller scale LC–MS (UPLC–MS to μLC–MS). Due to the combination of a modified sample preparation approach and the application of μLC–MS a lower LOQ could be achieved for infliximab compared with a previously developed UPLC–MS method.