The recent discovery and rapid spread of mobile colistin-resistant gene, mcr-1, among bacteria isolated from a broad range of sources is undermining our ability to treat bacterial infections and threatening human health and safety. To prevent further transfer of colistin resistance, practical and reliable methods for mcr-1-containing bacteria are need. In this study, standards and novel polyclonal and monoclonal antibodies (mAbs) against MCR-1 were developed. Among nine mAbs, three were MCR-1 specific and six cross-reacted with both MCR-1 and MCR-2. A sandwich enzyme-linked immunosorbent assay (ELISA) was established using the polyclonal antibody as a capturer and the mAb MCR-1-7 as a detector. The assay had a limit of detection of 0.01 ng/mL for MCR-1 and 0.1 ng/mL for MCR-2 in buffer with coefficients of variation (CV) less than 15%. When applied to ground beef, chicken and pork, this ELISA identified samples inoculated with less than 0.4 cfu/g of meat, demonstrating its strong tolerance to complex food matrices. To our knowledge, this is the first immunoassay developed for MCR-1 and MCR-2. It should be useful for prompt and reliable screening of meat samples contaminated with plasmid-borne colistin-resistant bacteria, thus reducing human risk of foodborne infections with possibly no antibiotic treatment options.