Statistical Design and Methodology for Validating a Multiplex Electrochemiluminescent Assay Detecting Multiple Endpoints in Human Sera Samples

• Activity Number: 385 - Biomarkers, Endpoint Validation and Other Topics
• Type: Contributed
• Date/Time: Thursday, August 12, 2021 : 12:00 PM to 1:50 PM
• Sponsor: Biopharmaceutical Section
• Author(s): Mohammad Lutfur Rahman* and Lingyi Zheng
• Companies: Sanofi Pasteur and Sanofi Pasteur
• Keywords: LLOQ: Lower Limit of Quantitation; DT = Diptheria Toxoid; TT = Tetanus Toxoid ; PT = Pertussis Toxin; FHA = Filamentous Haemagglutinin ; PRN = Pertactin

Abstract

The Diphtheria, Tetanus, and Pertussis Electrochemiluminescent method (DTP-ECL) is a multiplexed assay used for the simultaneous detection of antibodies against Diphtheria toxoid, Tetanus toxoid, and four (4) Pertussis antigens (pertussis toxin, PRN, FIM, FHA) in human clinical serum samples. DTP-ECL, was jointly developed by Global Clinical Immunology department of Sanofi Pasteur, Swiftwater, PA and Meso Scale Diagnostics, LLC, Rockville, MD.Multiple statistical design was used to evaluate critical assay parameters within wide analytical range of the assay. Specificity via competition was assessed by competing 10 positive samples and Specificity via Matrix Effects was assessed to prepare in hemolytic, lipidic, icteric, and unrelated matrices and evaluated by percent recovery. Accuracy was evaluated by Percent Recovery. Repeatability and intermediate precision were assessed using 96 samples and each sample was tested by a minimum of two analysts on at least three days and analyzed using mixed model. Precision at LLOQ range was verified between LLOQ/2 and 4xLLOQ. Linearity assessment was performed using Linear regression.DTP-ECL method met acceptance criteria of above parameters.