基于纳米金、量子点的均相可视化技术在多组分同步检测中的应用研究

Recently, the simultaneous determination of multiresidues in food and feed is the research hot point. However, these methods are mainly based on the expensitive instruments such as high performance liquid chromatography or liquid chromatography tandem mass spectrometry, which is difficult and expensive. The rapid dectection based on the chemistry or immunology need not large apparatus and equipment, and is easy to operate. But the shortcoming of rapid detection is only dectect a analyte one time. Thus, this project is that the multicomponent simultaneous detect using gold nanoparticle and quantum dots based homogeneous colorimetric assays with the naked eye. For example, beta-agonists showed the capability of directly reducing HAuCl4 into atomic gold, which involved oxidation of the amine or phenol group on the benzene ring of the beta-agonists. The resulted atomic gold formed AuNPs spontaneously, which is red with naked eye and had strong plasmon absorption at 528 nm. The linear relationship between the concentrations of beta-agonists and the AuNPs plasmon absorbance granted quantitative determination of beta-agonists in solutions. In addtion, QDs are robust inorganic fluorophores that exhibit size-dependent fluorescence emission spectra that span the visible spectrum. Their broad emission spectra allow simultaneous excitation of different particle sizes at a single wavelength with emission at multiple wavelengths. Thus, Quantum dots (QDs) have the potential to simplify the performance of multiplexed analysis. Sandwich immunoassays for the detection of multicompent were performed simultaneously in a homogeneous immunosaay using magnetic beads as support. The developed colorimetric assay can be used for the detection of beta-agonists, terracyline antibiotics, bacterial toxins, soybean proteins from feed, serum, urine and other liquid samples in the presence of interference from complex matix, which is a real high throughput rapid method.

多组分残留检测以其高效快速等优势迅速成为食品检测技术发展的热点。目前多残留检测方法主要是利用大型仪器如色谱质谱仪，测试成本和难度都比较大。而基于化学、免疫学等的快速检测技术虽不需要大型仪器，操作也简单，但只能实现单一组分的检测。因此，本项目提出基于纳米金、量子点的高灵敏度的均相可视化多组分快速检测技术研究，即将纳米金、量子点在不同的尺寸、不同聚集状态所表现出的颜色不同和相应紫外、荧光光谱的差异，与均相反应的简单快速相结合，采用化学比色反应或荧光均相免疫的方法，不需要仪器，只需通过检测样品的溶液颜色变化实现在复杂基质如饲料、牛奶、血液、尿液等中的beta-受体激动剂、四环素类药物、霉菌毒素、大豆抗原蛋白等多组分残留检测。同时借助一些提高灵敏度的检测技术如纳米种子的催化反应等，使灵敏度也能达到甚至超过仪器检测的水平。该技术能够在短时间内分析多个样本的多种成分，真正实现在种类和数量上的高通量。

多组分残留检测以其高效快速等优势迅速成为食品检测技术发展的热点。目前多残留检测方法主要是利用大型仪器如色谱质谱仪，测试成本和难度都比较大。因此，本项目提出基于纳米金、量子点的高灵敏度的均相可视化多组分快速检测技术研究，主要研究工作如下：（1）建立了基于纳米金的均相可视化技术检测畜禽产品中的多组分残留。分别以13种-受体激动剂类药物和4种四环素类药物为检测对象，在一定的化学反应环境下，利用其与氯金酸反应生成的纳米金的颜色变化肉眼筛选是否含有该类药物，并同时用紫外可见光谱监测纳米金在520 nm左右的特征吸收峰，实现这些物质在畜禽产品的高灵敏度可视化检测。（2）建立一种快速检测动物体液中β-激动剂、四环素类和磺胺类药物的纳米金试纸条免疫层析技术。该方法选用高灵敏度的抗体，采用胶体金微孔冻存技术，检测动物体液中的β-激动剂、四环素类和磺胺类药物等3类共26种兽药，具有简单快速等特点。各药物的消线浓度在0.2-10 ppb之间，检测灵敏度远低于我国及欧盟等对这些药物的最高限量要求，且不需要仪器设备。（3）建立了基于荧光量子点的可视化技术检测饲料中的多组分残留。项目以玉米赤霉烯酮和呕吐毒素2种霉菌毒素为研究对象，利用CdSe作为荧光探针标记抗不同目标分析物的抗体，利用荧光量子点的荧光信号建立一种特异性强、简便高效的多种目标分析物快速检测的免疫试纸条分析技术。（4）实际样品的测定。通过动物饲喂试验或市场收集，得到含有不同浓度的-受体激动剂类违禁药物、四环素类抗生素药和霉菌毒素的实际阳性样品，利用上述所建立的可视化技术进行快速筛选，同时样品用液相色谱-串联质谱进行比较和验证，结果表明建立的快速多组分检测技术的灵敏度和准确度均到达了液相色谱-串联质谱的水平。该技术真正实现在种类和数量上的高通量，必将极大加快检测速度和降低检测成本，从而能够大大加强执法机构的执法力度，确保畜禽产品的安全，创造良好的社会和经济效益。