Newborn calves are most susceptible to infection with Mycobacterium avium subsp. paratuberculosis (MAP), the cause of paratuberculosis. Colostrum is an important source of this organism due to direct excretion of MAP within the milk gland or indirect contamination with MAP-infected faeces. The aim of this study was to develop a rapid method based on an immunomagnetic separation (IMS) technique for the sensitive detection of MAP in bovine colostrum. The IMS was performed with mouse monoclonal antibodies (IgG) against lipoarabinomannan (LAM), a component of the cell wall of mycobacteria, secondary coated onto Dynabeads® Pan Mouse IgG. The hybridoma cell line used for the production of the anti-LAM IgG was kindly provided from John T. Belisle, National Institute of Health, Colorado State University, USA. IMS was carried out by adding 10 µl anti-LAM IgG coated beads (ca. 3 x 10⁶ beads) to the centrifuged pellet of 1 ml MAP-spiked colostrum resuspended in 900 µl phosphate buffered saline and incubating for 45 to 60 min at room temperature with gentle rotation. After magnetic separation the cell-bead complexes were resuspended in lysis buffer, transferred to tubes containing glass beads and mechanically disrupted in a Hybaid ribolyser for 3 x 15 s at 6.5 m/s and then incubated with proteinase K for 30 min at 65°C. Following DNA extraction with NucleoSpin Food Kit® (Macherey-Nagel), IS900 PCR was applied for the specific detection of MAP. Experiments showed that IMS in conjunction with IS900 PCR recovered MAP from artificially inoculated colostrum containing between 10 and 100 CFU/ml. This developed method offers a sensitive rapid detection system to screen reserves of colostrum for the presence of MAP as part of a herd-level paratuberculosis control programme.