A New Insulin Immunoassay Specific for the Rapid-Acting Insulin Analog, Insulin Aspart, Suitable for Bioavailability, Bioequivalence, and Pharmacokinetic Studies

Objectives: To validate a specific enzyme-linked immunosorbent assay for the rapid-acting human insulin analogue, insulin aspart, in human serum, human plasma, and porcine plasma. Design and methods: For the enzyme-linked immunosorbent assay, two murine monoclonal antibodies were developed that bind to two different epitopes on the insulin aspart molecule. Key parameters for validation were imprecision, accuracy, matrix effects, dilution-linearity, and cross-reactivity. Results: No cross-reactivity was found with human and porcine insulin, human proinsulin, or human C-peptide. The assay is sensitive (limit of QUANTIFICATION = 11.5 pmol/L), accurate (95–107% recovery with human serum, human plasma, and porcine plasma in the range 16–800 pmol/L), and has a 14.7% total imprecision within the entire analytical range. Dilution of samples gave linear results with human serum as the diluent. Conclusions: The insulin aspart-specific enzyme-linked immunosorbent assay described in this study is well suited to study the bioavailability, bioequivalence, and pharmacokinetics of this insulin analogue.