Using molecular descriptors for assisted screening of heterologous competitive antigens to improve the sensitivity of ELISA for detection of enrofloxacin in raw milk

Abstract

The use of the heterologous competitive strategy has become a vital method to improve the sensitivity of ELISA. In this work, we prepared an anti-enrofloxacin (ENR) mAb with ENR-bovine serum albumin (BSA) as immunogen. The molecular descriptors of quinolones were then used to screen heterologous coating antigens for the detection of ENR based on an ensemble learning method to improve the sensitivity of the ELISA. Results indicated that 6 of the 7 selected heterologous competitive antigens could enhance the sensitivity of ELISA. The ELISA sensitivity for the detection of ENR with sarafloxacin-BSA as heterologous coating antigen was improved 10-fold (in PBS) and 6-fold (in milk) compared with that with ENR-BSA as homologous antigen. The strategy can effectively screen suitable heterologous competitive antigens to improve the sensitivity of ELISA, followed by preparation of mAb with no additional modification to the corresponding immunogen.

Immunochromatographic assays for ultrasensitive and high specific determination of enrofloxacin in milk, eggs, honey, and chicken meat

“…In a study by Li et al (2019), either carbon dots or quantum dots were used as donor probes in 2 fluorescence-quenching immunochromatographic assays to monitor ENR with visual limits of detection of 0.5 to 2.5 μg/kg in animal-derived food samples. Other immunoassay methods including indirect competitive-ELISA (ic-ELISA; Huet et al, 2006;Hu et al, 2010Hu et al, , 2019Kim et al, 2015), colloidal gold immunoassay (Zhao et al, 2008), biomimetic enzyme-linked immunosorbent assay (Wang et al, 2017), fluorescence polarization immunoassay (Shen et al, 2019a), and chemiluminescence enzyme immunoassay (Kim and Kim, 2009;Kim et al, 2013;Yu et al, 2014a,b) have also been reported for the determination of ENR. Although the above-mentioned methods are able to meet the qualitative and semiquantitative analysis requirements for some samples, there is still scope for a rapid quantitative analytical test for ENR, especially for egg samples with more sensitive detection requirements.…”

High-flux smartphone-integrated lateral flow assay based on chrysanthemum-like Au@polydopamine for sensitive detection of enrofloxacin in milk

“…The LODs of sLFA based on AuNC@PDA-mAb, AuNP-mAb, AuNC@PDA-mAb′, and vvAuNP-mAb″ were 33.78, 169.99, 89.29, and 216.47 pg mL −1 , respectively. When the same antibody (mAb) was used, the LOD of sLFA based on AuNC@PDA-mAb (33.78 pg mL −1 ) was 5.0-fold smaller than that based on AuNP-mAb (169.99 pg mL −1 ); even when an antibody with slightly poor performance (i.e., mAb′) was used, the LOD of the sLFA based on AuNC@PDA-mAb′ (89.29 pg mL −1 , the preparation methods of the two antibodies from different sources may vary, resulting in a different affinity to ENR, 11 which is one of the key factors affecting the LFA detection performance 30 ) was 1.9-fold smaller than that based on AuNP-mAb, indicating the superiority and universality of AuNC@PDA in the LFA. These might be due to the advantage of morphological and optical properties of AuNC@PDA compared with AuNPs.…”

Sensitive immunoassays based on a monoclonal antibody for detection of marbofloxacin in milk

“…The ic-ELISA was carried out as described earlier (Hu et al, 2019). In brief, the coating antigen was added to a 96-well plate at 100 µL/well.…”