The Preliminary Study On Construction And Application Of Rapid Detection System Of Staphylococcus Aureus Based On Biosensor

ObjectivesStaphylococcus aureus is a major pathogen of nosocomial infection, and its toxins and invasive enzymescan cause symptoms of poisoning and severe infections. The traditional means of detection Staphylococcus aureus is a complicated process, which needs long time and sophisticated equipments, not suitable for real-time detection. So we needa means which suitable for scene analysis, real-time monitoring and low time-consuming. This means can realize the real-time detection of Staphylococcus aureus, with high sensitivity and specificity.MethodsThis experiment detected of Staphylococcus aureus using integrated all fiber structure immune fluorescence biosensor, according to the principle of immune sandwich antibody-antigen-fluorescence labeled antibody. Biological groups modified optical fiber probes coupling with Staphylococcus aureus specific antibody, followed by adding a first antibody, biotinylated second antibody and quantum dots labeled with streptavidin, and then were detected by access to the biosensor. The fluorescence intensity reflects the concentration of Staphylococcus aureus. Evaluated the sensitivity and specificity for the detection of the standard strains of Staphylococcus aureus with application of biosensor. And to evaluate the detection ability for Staphylococcus aureus strains isolated from clinical.Results1.After a series of hydroxyl, amino and aldehyde modification, the fiber probes can achieve the coupling of antibodies, and with the increase of the antibody concentration, the number of conjugated antibodies showed an increasing trend. The fiber coupled with specific antibody can specifically bind to the target substance, but not the nonspecific adsorption.2. The bacteria count under the microscope is more convenient and simpler than the plate counting method, and both of them are in an order of magnitude, and can be replaced by the plate counting method. The fiber probe conjugated with the Staphylococcus aureus specific antibody can capture the bacteria, and the capture rate is stable, up to 18.9%.3. With all fiber structure of the bio-sensor and detection method using immunofluorescence and choosing high light stability of quantum dots as fluorescent materials, can realize the detection of the standard strains of Staphylococcus aureus. The minimum detection range is 104cfu/ ml, with very high sensitivity and good ability for quantitative detection of 104-108cfu/ml Staphylococcus aureus. Meanwhile, we also detected the clinical common bacteria, such as epidermal Staphylococcus, Bormann real bacillus, Escherichia coli, Pseudomonas aeruginosa, pneumonia Klebsiella, but results showed no significant difference compared to the negative control group, indicating that has good specificity.4.The above instrument and method were used to detect the clinical isolates of Staphylococcus aureus, the lowest detection range was 103/10 cfu/ml, which indicated a high sensitivity.Conclusions1. Optical fiber probe modified with biological groups can achieve a good coupling with antibody;2. Optical fiber coupled with specific antibody has high capture rate of bacteria;3. The minimum sensitivity for detection of Staphylococcus aureus strains standard is 104cfu/ml by using all fiber immune fluorescence biosensor, and with good specificity, and can realize the detection of clinical isolates of Staphylococcus aureus strains, the minimum detectable limits for 103cfu/ml.