Objective: To prepare the polyclonal antibodies against Golgi protein 73 (GP73) and preliminarily establish the particle-enhanced turbidimetric immunoassay.
Methods: New Zealand rabbits were immunized with recombinant GP73 protein to obtain the polyclonal antibodies. The antibodies were purified and covalently attached to latex particles. Under the optimized reaction conditions, the covalent latex-antibody complexes were prepared to establish the GP73 particle-enhanced turbidimetric immunoassay.
Results: The titer of purified polyclonal antibodies reached 16 000. The best reaction conditions were that the concentration of EDAC/NHS was 10 mg/mL, the reaction solution was PBS (pH6), reaction time was 3 hours at room temperature. Under such conditions, the coupling efficiency was the highest. The particle-enhanced turbidimetric assay showed a good specificity and sensitivity with a good linear relationship in the range of 6.25-200 ng/mL. The determination limit was 10 ng/mL.
Conclusion: The study has successfully purified anti-GP73 antibodies, prepared the covalent latex-antibody complexes and established particle-enhanced turbidimetric immunoassay.