＜正＞ Mycotoxin deoxynivalenol produced by Giberella zeae is toxic both to human and animals. It is also a virulence factor for the fungal pathogen to infect cereal crops. Monitoring and neutralization of the toxin has long been the major practice in the toxin management. To neutralize its toxicity, expression of high affinitive scFv in planta is a promising approach. Moreover, replacement of monoclonal antibody with scFv in toxin detection based on immuno-assay is also very attractive because of its low cost and non-animal tissue in concern. We isolated the heavy-chain and lambda light-chain variable region genes from an anti-deoxynivalenol hybridoma cell line by reverse transcription PCR and jointed by a DNA linker encoding peptide （Gly4Ser）3 as a single-chain Fv （scFv） DNA fragment. The scFv fragment was cloned into a phagemid （pCANTAB 5E） and expressed as a fusion protein with E-tag and phage M13 p3 in Escherichia coli TG1. In the presence of helper phage M13 KO7, the scFv fusion protein was displayed on the surface of recombinant phages. High-affinity scFv phages were enriched through affinity selection in microtiter wells coated with deoxynivalenol-ovalbumin conjugate. One of the selected clones （pDON1.G2） in HB2151 mainly produced a soluble scFv antibody in periplasm with a high deoxynivalenol-binding affinity, and low cross-activity with deoxynivalenol analogs as that of its parent monoclonal antibody in a competitive direct enzyme-linked immunosorbent assay. Soluble scFv antibody from clone pDON1.G2 was also used to detect DON spiked and naturally contaminated in wheat samples, and no significant difference was observed when compared with its parental monoclonal antibody. High correlation was observed when detected the DON in naturally contaminated wheat samples between scFv and monoclonal antibody.The anti-DON scFv gene was then recloned into a fusion vector for the production of a fusion protein comprising the antibody fragment fused to a potent bacterial alkaline phosphatase variant PhoA. The anti-DON scFv-PhoA fusion protein was bifunctional and possessed both antigen binding capacity and PhoA activity. The recombinant conjugate was directly used, without further purification, for one-step immunodetection. The detection limit was identical and the test was quicker than the conventional two-step procedure with the purified anti-DON MAb and a secondary enzyme-labeled antibody. And the present study shows that the fusion protein provides a new tool for immunodetection which presents easier and quicker production and usage with the same sensitivity and specificity as classical reagents. The recombinant anti-DON scFv-PhoA conjugate is a promising alternative reagent for applications involving the immunodetection, such as the detection and characterization of microorganisms.In summery, the scFv and scFv-PhoA are possibly replacements of its monoclonal antibody in immunoassay, and this method showed its distinguished advantages, and towards a