Immunoassays are commonly used for clinical diagnosis, although interferences have been well documented. The streptavidin-biotin interaction provides an efficient and convenient method to manipulate assay components and is currently used in several immunoassay platforms. To date, there has been no report in the literature of interference from endogenous anti-streptavidin antibodies; however, such antibodies would potentially affect multiple diagnostic platforms. We report results from a patient being treated for thyroid dysfunction who demonstrated a T-uptake result of less than 0.2 and a nonlinear thyroid stimulating hormone dilution that suggested an immunoassay interference. Protein-A sepharose pretreatment corrected the nonlinear dilution and revealed an interference trend of falsely decreased results, as measured by sandwich assay, and falsely elevated results, as measured by competitive assay. The results of streptavidin-agarose adsorption were comparable to adsorption with protein-A sepharose. To our knowledge, this is the first published description of an endogenous anti-streptavidin antibody interfering with clinical laboratory assays.