Abstract:

BACKGROUND: In recent years, there are many reports on serological identification of antigens by recombinant expression cloning (SEREX) technology. However, there are higher false positive and poor stability problems.
OBJECTIVE: To establish a sensitive and stability system by optimizing the SEREX technology.
METHODS: The SEREX technology was used to compare the serum pretreat methods, isopropyl-beta-D-thiogalactoside adding methods, and the effect of different blocking conditions and medium thickness on the results of the coloration, and the plaque stained depth or not, background clean or not and false positive produce or not were observed.   
RESULTS AND CONCLUSION: The best result was obtained using following conditions: the serum was incubated directly;  nitrocellulose membrane was immersed in isopropyl-beta-D-thiogalactoside and dried in air and then blocked with skimmed-milk; the thickness of the medium could efficiently eliminate the brim effect. It suggests that the serum pretreat methods, isopropyl-beta-D-thiogalactoside adding methods, as well as different blocking conditions and medium thickness play a key role in the selection of antigen by using SEREX technology.