An immunochemical sol-gel-based immunoaffinity purification (IAP) method for purification and detection of the progestin drug medroxyprogesterone acetate (MPA) was developed. A polyclonal antibody (Ab) for MPA was generated, and two competitive (indirect and direct) sensitive enzyme-linked immunosorbent assays (ELISAs) for its detection were developed and implemented to determine the recovery and efficiency of the sol-gel based IAP method. The detection limits of the assays were 1.4 ± 0.2 ng·mL

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... −1 (n = 4) and 4.0 ± 0.4 ng·mL −1 (n = 25) for the indirect and direct ELISAs, respectively. The Abs did not exhibit cross-reactivity with any other progestin or steroid hormone, with the exception of megestrol acetate, with which the Ab exhibited 76% cross-reactivity. The sol-gel IAP method successfully eliminated serum interference to a degree that enabled ELISA analysis of spiked serum samples. This method was also found fully compatible with subsequent chemical analytical methods, such as liquid chromatography followed by mass spectrometry (LC-MS/MS). The approaches developed in this study form a basis for analysis of MPA in biological samples and may be further used to study population exposure to MPA and to monitor MPA contamination in water samples.