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The human bowel contains a large and dynamic bacterial population that is not only essential for intestinal health, but also critical for the development of diseases such as cancer. In this respect, the Gram-positive bacteriumStreptococcus bovishas been associated with colon cancer for many years. To investigate the clinical importance of this association, an immunocapture mass spectrometry assay was developed that can generate infection-related protein profiles. The composition of these profiles is governed by the capture of specific antigens by serum antibodies from colon cancer patients. This assay showed thatS. bovisantigen profiles could distinguish 11 out of 12 colon cancer patients from 8 control subjects, whereas antigen profiles derived from the gut bacteriumEscherichia coliwere not diagnostic for colon cancer. Moreover,S. bovisantigen profiles were also detected in polyp patients, indicating that infection with this bacterium does occur early during carcinogenesis. Highly accurate tandem mass spectrometry was used to identify one of the diagnostic antigens as a surface-exposed heparin-binding protein, which might be involved in attachment ofS. bovisto tumor cells. Together, these findings corroborate the hypothesis that colonic lesions provide a specific niche forS. bovis, resulting in tumor-associated “silent” infections. These infections, however, only become apparent in colon cancer patients with a compromised immune system (bacteremia) or coincidental cardiac valve lesions (endocarditis). This makes profiling of the humoral immune response against “silent”S. bovisinfections a promising diagnostic tool for the early detection of human colon cancer, which is crucial for the effective treatment of this disease.