Establishment And Optimization Of Bacteria Detection Conditions Based On The Tenebrio Molitor

Prophenoloxidase activation system is one of an important mechanism of the innate immune in insect. As Tenebrio molitor larva a model, the condition of prophenoloxidase activation and activity assay of phenoloxidase were established and optimized in vitro.As peptidoglycan an inducible factor, the influence of temperature, pH, activators, pre-reaction time and other factors were studied with orthogonal experimental method. The optimal condition of prophenoloxidase activation and activity assay of phenoloxidase as fellow: temperature is 30℃, the amount of activator is 5μ1, reaction pH value is 8 and pre-reaction time is 5minutes. Among several factors, the reaction temperature and pH value have significant influence of the detection. The complete strains of Escherichia coli (G+) and Staphylococcus-aureus (G-) can activate prophenoloxidase activation system under such condition, but the higher concentration and more detection time are needed, also the low sensitivity.Selected these strains and treated them by the physical, biological and chemical ways, then used the prophenoloxidase activation system to detect them and screen the best method, which is the optimized chemical method. Among 10 kinds of tested strains, eight different strains could activate the prophenoloxidase activation system with 10 cfu. According to the results of quantitative analysis, different strains have different reactive time, reactive sensitivity, reactive significance during activation of prophenoloxidase system.