Heterologous immunoassay strategy for enhancing detection sensitivity of banned dye rhodamine B in fraudulent food

Abstract

Background The high quality of antibody (Ab) is critical for an immunoassay; usually, an Ab with low affinity is often regarded as a “bad” one in the immunoassay development. How to use a “bad” Ab to develop a highly sensitive immunoassay is still a huge challenge. Methods In this study, a heterologous immunoassay strategy was designed to enhance the sensitivity for the detection of banned dye, rhodamine B (RB), in fraudulent food. The RB Ab could not recognize RB by pairing with homologous coating antigen (Ag). However, the Ab showed unexpected high specificity and sensitivity recognition after being replaced by heterologous coating Ag. Indirect competitive enzyme-linked immunosorbent assay (icELISA) was developed based on the heterologous strategy. Results The detection limit of icELISA for chilli powder, Chinese prickly ash, hot-pot seasoning, and chilli sauce was 0.002 μg/kg, and the recoveries of the four samples ranged from 76.0 to 102.0%, with the coefficient of variation between 3.9 and 18.8%. Parallel experiment for 20 market samples with high-performance liquid chromatography (HPLC) was performed on to confirm the performance of the practical application of the developed icELISA, and the results of the two methods had good correlation. Molecular modeling inferred that the carboxyl group of hapten and its exposure level played an important role in the hapten-Ab recognition. Conclusions The proposed icELISA can be used for the surveillance screening of RB in a range of seasoning foods, and the heterologous strategy is an effective approach to enhance the sensitivity in an immunoassay.

Conjugates of Aminopenicillins with Proteins: Synthesis, Immunogenic Properties, and Binding to the β-Lactam Receptor and Antibodies

“…3) confirmed that antibodies from the antiserum to Amp-pPh-TG were more sensitive to penicillin antibiotics than polyclonal antibodies from the antiserum to Amp-rhLF. In line with observations by Wang et al [21], we found additionally that better immunoanalytical parameters were ensured by heterologous immunogen and conjugated antigen constructs, which differed in the structure of the hapten used for their synthesis. In the case of the antiserum to Amp-pPh-TG, the effect was observed both when the haptenprotein linker differed between the constructs and when the linker was the same, but the constructs included Amp or Amox.…”

Authenticating Emergent Adulterant 6-Benzylaminopurine in Bean Sprouts: Virtual Hapten Similarity Enhanced Immunoassay

“…Since the heterologous coating antigens frequently cause a minor decrease in antibody recognition when compared to the homogeneous coating antigen, analytes can be competed with the heterogeneous coating antigen at lower concentrations and achieve better sensitivity. 30 Similarly, in this study, the inhibition ratios of the antisera to 6-BA was improved by the use of heterologous coating antigens. The highest inhibition ratio of 90% for 6-BA was obtained from hapten 1-BSA when hapten 1-BSA was paired with hapten 2-OVA, which was utilized as the coating antigen (Table S1).…”

“…In addition, it is generally understood that heterologous coating antigens typically lead to enhanced sensitivity and lower immunoreagent dilution in icELISA. Since the heterologous coating antigens frequently cause a minor decrease in antibody recognition when compared to the homogeneous coating antigen, analytes can be competed with the heterogeneous coating antigen at lower concentrations and achieve better sensitivity . Similarly, in this study, the inhibition ratios of the antisera to 6-BA was improved by the use of heterologous coating antigens.…”

Molecularly imprinted polymer-coated hybrid optical waveguides for sub-aM fluorescence sensing

“…However, a thorough safety evaluation has provided sufficient evidence to suggest that Rh B contamination irritates the skin, eyes, and respiratory tract and even causes carcinogenic and teratogenic effects on public health. [11][12][13] To achieve highly sensitive detection of Rh B, various methods have been proposed, such as surface-enhanced Raman scattering (SERS), 14,15 electrochemical detection, 16 enzyme-linked immunosorbent assay (ELISA), 17 and high-performance liquid chromatography (HPLC). 18 Optical evanescent-wave (EW) sensors have attracted considerable attention owing to their features including immunity to electromagnetic interference, electrical insulation, high corrosion resistance, and low attenuation over a long distance.…”