Utilization Of A Lateral Flow Assay Based On Surface-enhanced Raman Scattering For Detection Of Antibiotics In Milk

Milk is the natural ideal food catering to people of all ages with its good nutrition.However,in the recent years,milk with remains of antibiotics gradual emergence has get frightened upon hearing of it.Antibiotics has good curative effect in preventing and treating cow diseases,but failure to follow instructions and overuse may cause serious adverse reactions,or even deadly accident,so it is necessary to realize highly sensitive,high-throughput,quick and easy detection.?1?The immunoprobe?NEO m Ab-AuNPs-PATP?was prepared from colloidal gold?AuNPs?conjugated with both a Raman probe molecule 4-aminothiophenol?PATP?and a monoclonal antibody against neomycin?NEO-mAb?.The SERS-LFA method was established and the parameters of the assay were optimized.Based on this,the IC₅₀0 and the limit of detection?LOD?were 0.04 ng/mL and 0.216 pg/mL of NEO,respectively.There is no cross-reactivity?CR?of the assay with other compounds,showing high specificity of the assay.The recoveries for milk samples with added NEO are in the range of 90%-106%with the relative standard deviations?RSD?of 3%-5%?n=3?.At five different concentrations,we randomly selected 10 different spots in each concentration and measured,the RSD of 3.8%,4.4%,2.9%,3.2%and 5.1%.And the comparative experiments for detection NEO reveal SERS-LFA has the highest sensitivity.?2?Based on the?Fe₂O₃@Au magnetic gold nanoflowers,a SERS-LFA for detection of neomycin was established.The experimental parameters are optimized.We obtained the IC₅₀0 of NEO was 0.013 ng/mL and the LOD was 0.17 pg/m L by using SERS-LFA.The recoveries of NEO from spiked milk samples ranged between 84%and 109%,with relative standard deviations?RSD?from 4%to 6%?n=3?.And we evaluated the reproducibility and specificity of the assay.?3?The immunoprobes of NEO mAb-Au NPs-PATP and NOR m Ab-AuNPs-PATP were prepared,and the strip with two test lines for simultaneous detection of neomycin?NEO?and quinolones antibiotics?QNS?was assembled.Under optimized conditions,the IC₅₀0 were 0.03ng/mL for NEO and 0.07 ng/mL for NOR,and the LODs were 0.37 pg/mL for NEO and 0.55pg/mL for NOR by using SERS-LFA.The NEO test line is not interfered by the NEO analogues,but the NOR test line suffers from different degrees of cross-reactivity?CR?to 12 common other QNS,the CRs ranging from 1.5%to 135.14%.The recoveries of NEO and NOR from spiked milk samples ranged between 86%and 121%,with relative standard deviations?RSD?from 3%to 6%?n=3?.And the comparative experiments for detection QNS reveal SERS-LFA have the highest sensitivity.?4?The immunogen?LIN-BSA?and coating antigen?LIN-OVA?for lincomycin were synthesized by sodium periodate method.The LIN monoclonal antibody?LIN m Ab? was successfully screened,and the half inhibitory concentration(IC₅₀)was 0.69 ng/m L by indirect competitive ELISA.Based on the m Abs,the immunoprobes of NEO m Ab-Au NPs-PATP and LIN m Ab-Au NPs-DTNB were prepared,and the strip with a blend test line for one-step detection of neomycin?NEO?and lincomycin?LIN?was assembled.Under optimized conditions,the IC50 were0.038 ng/m L for NEO and 0.024 ng/m L for LIN,and the LODs were 0.33 pg/m L for NEO and 0.29pg/m L for LIN by using SERS-LFA.The recoveries of NEO and LIN from spiked milk samples ranged between 84% and 113%,with relative standard deviations?RSD?from 3% to 8%?n=3?.And we evaluated the feasibility and specificity of the assay.In addition,it was compared with other methods to detect LIN.