Study On The Reference Materials Preparation Of Staphylococcus Aureus In Chicken

Objective:Staphylococcus aureus(S.aureus)is a Common pathogenic bacteria that cause of food poisoning,and it has great significance in the Public Health.Poultry,eggs and meat products are the main media of the S.aureus,it make a serious impact on the development of aquaculture and human health.Therefore those food that contaminate S.aureus vulnerable before into the market should make rigorous testing.To ensure the inspectors' capability,standardize the management of the kit,and to reduce the leak detection and error detection rate in food,we should increase efforts to develop the Biological Characteristics Reference Materials(RM),so that ensure the results true and reliable.The test launched a series of studies of preparation of S.aureus reference materials in chicken and S.aureus nucleic acid reference materials.On the basis to add matrix in Biological reference materials to make the test conformity to the actual value more.Supply a reference value for the same type of reference materials' preparation in future.Method:Firstly the three counting methods of S.aureus was selected.Using three kinds of counting method included PCA counting method,Baird-Parker dumping method and Baird-Parker dilution-plate method,for counting the sample solution containing the same amount of S.aureus.By analyzing the stability and reproducibility of the results of comparison of three methods,choosing the best method for counting S.aureus.Concentration curve plotted S.aureus.Turbidimetric assay,due to the concentration and the optical density(OD600 value)is proportional to the bacterial suspension,so it can take multifunctional enzyme superficially to infer the concentration of bacteria by determining the optical density of the bacterial suspension.And measured OD600 colonies corresponding to the numerical value of the plotted curve can be plotted in a concentration of bacteria under certain conditions,as a basis for the amount of bacteria added.Detection of antibiotic residues in the Pre-added chicken matrix and detection the matrix impact on S.aureus.After affirm the chicken matrix is meet the requirements,than take sterilization,the chicken matrix take 60Co-? sterilization,make the bacterial concentration in matrix<10CFU/g,to achieve the requirements of sterile matrix.Screening of protection used in Freezed-dried RM.S.aureus screening freeze-dried preservation protective agent composition and the best ratio.The selection test of protection formulation,skimming milk,polyvinylpyrrolidone(PVP),sucrose,L-glutamate single dose of four protection agent,three levels of each single dose design.A orthogonal table-L9(34)was adopted to design the experiments for the best combinational schemes.Make the final choice of protective agents effective in reducing mortality of S.aureus in a freeze-dried to prepare a RM for S.aureus.Lyophilized RM rehydrated before use,and therefore need to add a protective agent for dehydration chicken matrix test to determine the amount of water reuse.Test the uniformity stability,tracking,and assessment of uncertainty of reference material,where the uncertainty is mainly generated on the electronic balance and the method of the valuation reference material,combined with the uncertainty to setting of the reference material final.Results:Counts of Staphylococcus aureus,using Baird-Parker dumping method is simple,stable and good reproducibility.S.aureus-RM in chicken were conducted five batches of preparation,each batch prepared the SE-RM eontaining 70 bottles.Protective agent formulation screening results:20%skim milk+8%PVP+12%Sucrose+1%L-glutamic sodium.Chicken and protection dehydration results:5g chicken and 4mL protection freeze-dried was 2g,dehydration rate is 17.57%.Lyophilized reference material dissolve with water about 7mL.The results of the uniformity experiment showed that,the results of the Analysis of Variance are P>0.05,it indicated that there was no significant difference between the five batches of RM.Stability experiments showed that the RM stored at-20?,the microbial content was stabilized 140CFU/bottle after 120days.The assessment of Uncertainty around with the main experimental steps that make the major impact on RM.The uncertainty of electronic balance is 2.7934mg,so it can be neglected.And the uncertain of the constant value methods is 8,and the reference material final value is:value±8.Conclusion:The Study establish a Preparation method of S.aureus reference material.The uniformity are good,and store at-20?,the reference material have a good short-term stability.