Interlaboratory comparison of a multiplex bead assay against the Chlamydia trachomatis antigen Pgp3 using two monoclonal antibody clones

The bacterium Chlamydia trachomatis causes three distinct conditions – urogenital chlamydia, the eye disease trachoma, and lymphogranuloma venereum. Use of serological testing for exposure to C. trachomatis for evaluating ocular disease risk factors and population-based surveillance has expanded over the past decade. We developed chimeric monoclonal antibodies to the C. trachomatis antigen Pgp3 as standardized reagents for Pgp3 serological assays. Here we use two anti-Pgp3 monoclonals (1G1 and 1C3) to conduct an interlaboratory comparison of Pgp3 Luminex-based multiplex bead serological assays in 4 different laboratories (each with their own protocol) with 5 operators using 3 different instrument types. Antibodies spotted onto filter paper were stable for two weeks at room temperature. Accuracy and precision were evaluated at 3 concentrations for each monoclonal (high, medium, and low). Accuracy within 20% was achieved for 11/21 conditions for 1G1 and 18/21 conditions for 1C3; the high concentration resulted in the poorest accuracy. Precision was within 10% for all low and medium concentrations and was above 10% for several conditions at the high concentration. The fluorescence values differed between instruments but showed no difference when extrapolated to concentrations using a standard curve. These data suggest that 1C3 may be the more reliable standard reagent and that the Pgp3 multiplex assays are highly robust when comparing different instruments and assay protocols.