A Research Of Magnetic Nanoparticles Enhanced SPR Immunosensor For Detection Of Salmonella Enteritidis

In this paper, a new approach is developed for the detection of Salmonella enteritidis based on the two-channel surface plasmon resonance (SPR) immunosensor coupled with Fe3O4 magnetic nanoparticles(MNPs) separation. The selectivity, real sample detection and recovery test for Salmonella enteritidis detection were performed by developed SPR immunosensor approach. The main study contents were summarized as follows:1. Carboxyl-coated Fe3O4MNPs was produced using chemical coprecipitation method. The Carboxyl-coated Fe3O4MNPs was functionalized by Anti-Salmonella polyclonal antibody(PAb) using amino coupling method, and the coupling conditions for conjugates (immune MNPs) of the MNPs and PAb were optimized. The results showed that the optimal coupling conditions of MNPs and PAb were achieved when the volume ratio of 51.7 ?/mL MNPs, EDC/NHS(0.4 mol/L/0.1 mol/L) and 1.0mg/ mL PAb was 1:2:2 in the water bath of 37? for 40 min.2. The immunoMNPs were used for the enrichment and isolation of Salmonella enteritidis and the adsorption efficiency were evaluated. The adsorption efficiency will achieve when the volume ratio of 51.7 ?g/mL immune MNPs and 1.4 X 104cfu/mL S.enteritidis was 4:5, and incubating time is 40 min.3. SPR direct detection format for Salmonella enteritidis was achieved by immobilized PAb on the chip surface using amine coulping method, then Salmonella enteritidis of difference concentrationwere flowed over the SPR chip surface for directly detection. A good linear relationship was obtained between SPR signals and Salmonella enteritidis concentrations from 1.4×104 to 1.4×109 cfu/mL. The regression equation was Y=6.53X-22.73, R2=0.9874, the relative standard deviation (RSD) range was 4.02%-12.77%. Salmonella enteritidis concentrations as low as 1.4 ×104 cfu/mL can be readily measured.4. MNPs enhanced SPR immunosensor to detect Salmonella enteritidis was achieved by the enrichment and isolation of Salmonella enteritidis using immunoMNPs. Then S. enteritidis-immunoMNPs conjugates were directly flowed over the sensor surface modified with PAb. The selectivity experiment, real sample detection and recovery test were also performed. A good linear relationship was obtained between SPR signals and Salmonella enteritidis concentrations from 1.4 X 101 to 1.4×109 cfu/mL. The regression equation was Y=5.03X+5.41, R2=0.9956, RSD range was 2.124%- 12.480%. Salmonella enteritidis concentrations as low as 14 cfu/mL can be readily measured, which gives 4 orders of magnitude improvement in the sensitivity towards Salmonella enteritidis compared withdirect detection format. Eggshell as real sample was teken to conduct standard addition recoverey study, and the range of the recovery was 92.76%-113.25%(RSD 3.978%-7.345%).