The vaccination could induce high titer of anti-SPs antibodies against FMDV while FMDV infection induces both anti-SPs antibodies and anti-NSPs antibodies [4]. To distinguish between infected and vaccinated cattle, it is required to develop assay for detecting NSP-specific antibodies. Several ELISA tests have been described to detect the NSP-specific antibodies, using recombinant 3A [5], [10], [13] and [17], 3B [10] and [17], 3C [5], 2C [5], [14] and [15], 3AB [6] and [16] and 3ABC [5], [6], [7], [8], [9], [10], [11], [12] and [17] as coating antigens.

Among them, 3AB was reported as specific and sensitive coating antigen to distinguish antibodies induce by FMDV infection and vaccination [18]. In the study, we firstly attempted to express recombinant protein 3AB (r3AB) in Escherichia AZD2281 coli. However, the r3AB was mainly expressed in Temozolomide nmr the form of inclusion body, and the purified r3AB existed as a mixture of monomers and dimers. To overcome the disadvantages, a recombinant truncated FMDV 3AB protein, designated as r3aB, resulted from the deletion of 80 amino acid residues from N-terminal of 3AB, was expressed in E. coli. The r3aB was majorly expressed in soluble fraction and presented as homogeneous monomers after purification. Coated with the r3aB, an indirect ELISA was established for distinguishing the

antibodies induced by FMDV infection from those induced by vaccination in cattle. The assay could be potentially used to differentiate the cattle FMDV infected from those vaccinated. (I) Sera from naive cattle:

20 serum samples were collected from the cattle with no virus infection or vaccination. (II) Sera from vaccinated cattle: 137 serum samples were collected at 21 dpv from FMD free cattle after vaccination. Among them, 127 serum samples were collected from the cattle vaccinated with a commercial bivalent vaccine containing FMDV type Asia 1 and type O (Baoling Bio-pharmaceutical Corporation) and 10 serum samples were collected from cattle vaccinated with recombinant FMDV VP1 peptide vaccine. The FMDV VP1 peptide vaccine, designed and produced by Molecular Linifanib (ABT-869) Biology department of Jilin University, China, could induce neutralizing antibodies and protect the cattle from FMDV challenge. (III) Sera from infected cattle: 54 serum samples were collected at 21 dpv from cattle after infection. Among them, 30 and 24 serum samples were collected from cattle infected with FMDV strain of type Asia 1 and type O, respectively. The coding sequences of 3AB and 3aB were amplified using RT-PCR from FMDV (Asia I/Jiangsu/China/2005, GenBank: EF149009.1, provided by Jin Yu Company, Mongolia, China). DNA fragments of 672 bp for 3AB and 432 bp for 3aB were cloned into pET28a plasmid (Novagen) to construct recombinant expression plasmids designated as pET28a-3AB and pET28a-3aB, respectively. The plasmids were transformed into E. coli BL21 (DE3) (Novagen).