Broad-Specificity Chemiluminescence Enzyme Immunoassay for (Fluoro)quinolones: Hapten Design and Molecular Modeling Study of Antibody Recognition

Abstract

On the basis of the structural features of (fluoro)quinolones (FQs), pazufloxacin was first used as a generic immunizing hapten to raise a broad-specificity antibody. The obtained polyclonal antibody exhibited broad cross-reactivity ranging from 5.19% to 478.77% with 21 FQs. Furthermore, the antibody was able to recognize these FQs below their maximum residue limits (MRLs) in an indirect competitive chemiluminescence enzyme immunoassay (ic-CLEIA), with the limit of detection (LOD) ranging from 0.10 to 33.83 ng/mL. For simply pretreated milk samples with spiked FQs, the ic-CLEIA exhibited an excellent recovery with a range of 84.6-106.9% and an acceptable coefficient of variation below 15%, suggesting its suitability and reliability for the use of a promising tool to detect FQs. Meanwhile, comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) models, with statistically significant correlation coefficients (q(2)CoMFA = 0.559, r(2)CoMFA = 0.999; q(2)CoMSIA = 0.559, r(2)CoMSIA = 0.994), were established to investigate the antibody recognition mechanism. These two models revealed that in the antibody, the active cavity binding FQs' 7-position substituents worked together with another cavity (binding FQs' 1-position groups) to crucially endow the high cross-reactivity. This investigation will be significant for better exploring the recognition mechanism and for designing new haptens.

Development of a multicomponent immunochromatographic test system for the detection of fluoroquinolone and amphenicol antibiotics in dairy products

“…Enzyme‐linked immunosorbent analysis (ELISA) and immunochromatographic assays (ICA) are immunoanalytical methods based on the highly specific reaction between antibodies and the detected analyte and are characterized by cost‐effectiveness and rapidity. Thus, these techniques are often used to control foodstuffs for the presence of various contaminants, including antibiotics …”

Application of quantitative structure‐activity relationship analysis on an antibody and alternariol‐like compounds interaction study

“…The interaction between antigen and antibody of the immunoassay method was influenced by various factors. More and more studies have shown that the specificity of the antibody is of great significance for improving immunoassays . An antibody with high specificity (low cross‐reactivity [CR]) is suitable to be used in the immunoassay detecting a single compound, while an antibody with broad specificity (high CR) is suitable to be used in the immunoassay detecting multiple compounds .…”

“…More and more studies have shown that the specificity of the antibody is of great significance for improving immunoassays. [9][10][11] An antibody with high specificity (low cross-reactivity [CR]) is suitable to be used in the immunoassay detecting a single compound, while an antibody with broad specificity (high CR) is suitable to be used in the immunoassay detecting multiple compounds. [12][13][14][15] However, because of the lack of knowledge about the antibody recognition and the ambiguous structure-activity relationship, artificial control of the specificity of the antibody remains a huge challenge.…”

“…[12][13][14][15] However, because of the lack of knowledge about the antibody recognition and the ambiguous structure-activity relationship, artificial control of the specificity of the antibody remains a huge challenge. 10 Three-dimensional quantitative structure-activity relationship (3D-QSAR), the most common methods of which include comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA), is widely used in the study of antigen-antibody interactions. 16 In the absence of the antibody crystal structure, 3D-QSAR can reasonably explain the difference in the CR and effectively predict and elucidate the mechanisms for antibody recognition.…”

Doses of Immunogen Contribute to Specificity Spectrums of Antibodies against Aflatoxin

“…Due to the lack of understanding of the specific interactions between antibodies and target analytes or haptens, the antibody specificity resulting from the newly-designed hapten is often unpredictable, and this result comes only after laborious and time-consuming animal experiments. Sometimes an apparent rationally-designed “generic hapten” is unable to generate antibodies with the desirable sensitivity and specificity [13,14,15]. The diverse exposure of an antigenic determinant could form a broad spectrum of the antibody specificity.…”