Ultrasensitive quantification of aflatoxin B1 via nanobody phage -mediated quantitative PCR and digital PCR for agro-products

IF 10.7 1区生物学 Q1 BIOPHYSICS

Biosensors and Bioelectronics Pub Date : 2025-05-27 DOI:10.1016/j.bios.2025.117622

Junjie Zeng , Yanping Li , Qinghua He , Ji Xia , Jiebiao Guo , Zhui Tu

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引用次数: 0

Abstract

Phage display-mediated immuno-polymerase chain reaction (PD-IPCR) combines the strengths of phage display technology and immuno-PCR. The link between genotype and phenotype has been established by displaying antibody fragments on the surface of phage particles, allowing signal transmission. In this study, we utilized aflatoxin B₁ (AFB₁) as a model system to investigate the efficacy of PD-IPCR. Signal amplification was achieved using both real-time fluorescence quantitative PCR (qPCR) and a custom-made droplet digital PCR (dPCR) system. Under optimized conditions, the assay using dPCR achieved a limit of detection of 0.015 ng/mL, a 6-fold improvement over qPCR. The validation of our method using major agricultural produce, such as corn, flour, and soy sauce, shows excellent recoveries ranging from 85.77 % to 108.28 %, with a total assay time of less than 2 h. Furthermore, we compared our results with a high-performance liquid chromatography (HPLC) method for AFB₁ content in corn and flour, revealing consistent outcomes (r = 0.9847). These results demonstrate the potential of nanobody-based PD-IPCR as a reliable tool for detecting mycotoxins.

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利用纳米噬菌体介导的定量PCR和数字PCR对农产品中黄曲霉毒素B1的超灵敏定量

噬菌体展示介导的免疫聚合酶链反应（PD-IPCR）结合了噬菌体展示技术和免疫pcr技术的优势。通过在噬菌体颗粒表面展示抗体片段，建立了基因型和表型之间的联系，从而允许信号传递。本研究以黄曲霉毒素B1 （AFB1）为模型系统，研究PD-IPCR的疗效。利用实时荧光定量PCR （qPCR）和定制的微滴数字PCR （dPCR）系统实现信号扩增。在优化条件下，dPCR的检测限为0.015 ng/mL，比qPCR提高了6倍。以玉米、面粉、酱油等主要农产品为样品，对该方法进行验证，回收率为85.77% ~ 108.28%，总测定时间小于2 h。此外，将该方法与高效液相色谱法测定玉米和面粉中AFB1含量的结果进行比较，结果一致（r = 0.9847）。这些结果证明了基于纳米体的PD-IPCR作为检测真菌毒素的可靠工具的潜力。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Biosensors and Bioelectronics 工程技术-电化学

CiteScore

20.80

自引率

7.10%

发文量

1006

审稿时长

29 days

期刊介绍： Biosensors & Bioelectronics, along with its open access companion journal Biosensors & Bioelectronics: X, is the leading international publication in the field of biosensors and bioelectronics. It covers research, design, development, and application of biosensors, which are analytical devices incorporating biological materials with physicochemical transducers. These devices, including sensors, DNA chips, electronic noses, and lab-on-a-chip, produce digital signals proportional to specific analytes. Examples include immunosensors and enzyme-based biosensors, applied in various fields such as medicine, environmental monitoring, and food industry. The journal also focuses on molecular and supramolecular structures for enhancing device performance.