To prepare citrinin(CIT)-protein antigen, CIT was conjugated with bovine serum albumin (BSA) by 1, 4-butanediol diglycidyl ether. The spleen from the BALB/C mice immunized with CIT-BSA conjugate was used to fuse with the murine SP2/0. By subcloning, a hybridoma cell lines excreting monoclonal antibodies(McAb) against CIT was obtained and named H2-F8. The monoclonal antibodies obtained from hybridoma H2-F8 was of IgM subclass. The affinity constant of the McAb to CIT was 4.17×10⁸ L/mol and the IC₅₀ value was 0.3 μg/L. Its linear range of the assay was between 0.05 μg/L and 1.0 μg/L. The cross-reactivity rates were less than 0.1% of other toxins such as ochratoxin A, aflatoxin B1, deoxynivalenol, zearalenone and were less than 0.01% of rubropunctatine and rubropunctamine. This work would be helpful for establishing the technology and developing the kit to determine CIT-contaminated samples by ELISA.