Immunoassays employed at the point-of-care (POC) are often useful for diagnosing acute infections. Many of these assays rely on identification of microbial antigens that are secreted or shed during infection. However, determining which microbial antigens are best to target by immunoassay can be the most difficult aspect of developing a new diagnostic product. Here we describe a novel technique termed “In vivo Microbial Antigen Discovery” or “InMAD” for identification of microbial antigens that may be targeted for the diagnosis of infectious diseases.