The rCsMRP4-NBD1 was separated by 12% sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and cut off alone to obtain specific antigens for mouse immunization. The gel slice was equilibrated and homogenized in pre-cooled 1× PBS by complete grinding. The liquid homogenate containing rCsMRP4-NBD1 was then injected into BALB/c mice according to an immunization method . Blood was drawn from the eye and stored at room temperature for 1 to 2 h. The immune serum was obtained by centrifugation at 4000× rpm for 20 min. In order to examine the antibody titer in the immune serum against rCsMRP4-NBD1 and native CsMRP4, C. sinensis crude extracts were prepared using the Mem-PER Plus Membrane Protein Extraction Kit (Thermo scientific, Rockford, USA) following the manufacturer’s instructions. The crude antigen was then examined to determine its concentration using the Bio-Rad protein assay (Bio-Rad, Hercules, CA, USA) and then stored as aliquots at -70 °C until use.