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A magnetic‐nanobead‐based, substrate‐free method  for the sensitive detection of spores in an immunoassay format is presented. The method is shown to detect  Bacillus globigii  spores, the non‐pathogenic simulant of  Bacillus anthracis , with a limit‐of‐detection of 50 spores with a reaction time of 135 min. The study shows the versatility of magnetic nanobeads for detection of biological molecules other than DNA.