Compared to the traditional heterogeneous assays, a homogeneous immunoassay is a preferred format for its simplicity. By cloning and isolating luminescent proteins from bioluminescent organisms, bioluminescence has been widely used for various biological applications. In this study, we present the development of a homogeneous luminescence immunoassay (FNanoBiT assay) for detection of fumonisin B1 (FB1), based on the binding of two subunits of an engineered luminescent protein (NanoLuc). For the detection of the mycotoxin FB1 in foods, the anti-fumonisin antibody was conjugated to the large subunit of NanoLuc (FLgBiT), and the FB1 was conjugated to the small subunit (FSmBiT). The conjugates were used for the detection of FB1 in a competitive immunoassay format without the need of a secondary antibody, or washing steps. The developed FNanoBiT assay revealed high specificity toward FB1 with no cross-reactivity with other mycotoxins, and it demonstrated acceptable recovery (higher than 94%) and relative standard deviation from spiked maize samples. Further, the assay was successfully applied for the detection of FB1 in naturally contaminated maize, with a dynamic range of 0.533–6.81 ng mL-1 and a detection limit of 0.079 ng mL-1. The results derived with FNanoBiT assay of all spiked samples showed a strong correlation to those obtained by the High-performance liquid chromatography method. Thus, the FNanoBiT based homogeneous immunoassay could be used as a rapid, and simple tool for the analysis of mycotoxin-contaminated foods.

与传统的异质分析相比，同质免疫分析是一种简便易行的首选格式。通过从生物发光生物中克隆和分离发光蛋白，生物发光已被广泛用于各种生物学应用。在这项研究中，我们基于工程发光蛋白（NanoLuc）的两个亚基的结合，提出了用于检测伏马菌素B1（FB1）的均相发光免疫测定法（FNanoBiT测定）。为了检测食品中的霉菌毒素FB1，将抗伏马菌素抗体与NanoLuc的大亚基（FLgBiT）偶联，将FB1与小亚基（FSmBiT）偶联。所述缀合物以竞争性免疫测定形式用于检测FB1，而无需二抗或洗涤步骤。发达的FNanoBiT分析显示出对FB1的高特异性，与其他霉菌毒素没有交叉反应，并且显示出可接受的回收率（高于94％）和加标玉米样品的相对标准偏差。此外，该测定法成功地用于自然污染玉米中FB1的检测，其动态范围为0.533–6.81 ng mL-1，检出限为0.079 ng mL-1。FNanoBiT分析得出的所有加标样品的结果均与通过高效液相色谱法获得的结果具有很强的相关性。因此，基于FNanoBiT的均相免疫测定可以用作分析真菌毒素污染食品的快速，简单的工具。结果表明加标玉米样品的回收率（高于94％）和相对标准偏差均可接受。此外，该测定法成功地用于自然污染玉米中FB1的检测，其动态范围为0.533–6.81 ng mL-1，检出限为0.079 ng mL-1。FNanoBiT分析得出的所有加标样品的结果均与通过高效液相色谱法获得的结果具有很强的相关性。因此，基于FNanoBiT的均相免疫测定可以用作分析真菌毒素污染食品的快速，简单的工具。结果表明加标玉米样品的回收率（高于94％）和相对标准偏差均可接受。此外，该测定法成功地用于自然污染玉米中FB1的检测，其动态范围为0.533–6.81 ng mL-1，检出限为0.079 ng mL-1。FNanoBiT分析得出的所有加标样品的结果均与通过高效液相色谱法获得的结果具有很强的相关性。因此，基于FNanoBiT的均相免疫测定可以用作分析真菌毒素污染食品的快速，简单的工具。FNanoBiT分析得出的所有加标样品的结果均与通过高效液相色谱法获得的结果具有很强的相关性。因此，基于FNanoBiT的均相免疫测定可以用作分析真菌毒素污染食品的快速，简单的工具。FNanoBiT分析得出的所有加标样品的结果均与通过高效液相色谱法获得的结果具有很强的相关性。因此，基于FNanoBiT的均相免疫测定可以用作分析真菌毒素污染食品的快速，简单的工具。