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Trachoma, caused by repeated ocular infection with Chlamydia trachomatis (Ct), is targeted for elimination as a public health problem. Serological testing for antibodies is promising for surveillance; determining useful thresholds will require collection of serological data from settings with different prevalence of the indicator trachomatous inflammation-follicular (TF). Dried blood spots were collected during trachoma mapping in two districts each of Togo and Democratic Republic of the Congo. Anti-Ct antibodies were detected by multiplex bead assay (MBA) and three different lateral flow assays (LFA) and seroprevalence and seroconversion rate (SCR) were determined. By most tests, the district with > 5% TF (the elimination threshold) had five-sixfold higher seroprevalence and tenfold higher SCR than districts with < 5% TF. The agreement between LFA and MBA was improved using a black latex developing reagent. These data show optimization of antibody tests against Ct to better differentiate districts above or below trachoma elimination thresholds.