Affinity determination method in vibrio detection based on aptamer-an oligonucleotide probe screened by systematic evolution of ligands by exponential enrichment

Systematic evolution of ligands by exponential enrichment (SELEX) is a new technique that can screen high affinity oligonucleotide probe, called aptamer which have been widely used in the detections of mciroorganism, proteins, tumor cells and so on. However, as a key step in SELEX process, the affinity determination method is variable for the different targets. In the present paper, an aptamer against Vibrio alginolyticus was used to study and optimize the affinity determination method for the vibrio detection. Based on biotin-avidin system marked by enzyme, the binding temperature, time and binding ratio (vibro/aptamer) were studied and optimized through orthogonal experiment. The results showed that the ideal affinity was obtained after the aptamer bound with V.alginolyticus for 70 minutes at the temperature 32 degrees C by a mole ratio 1:100 (vibrio:aptamer). By this optimal affinity detemination method, the aptamer was used to test the affinity with four kinds of microorganisms and the result showed that the aptamer had the highest affinity with V.alginolyticus, which suggesting its specificity of the aptamer against V.alginolyticus.