Bimodal size distribution immuno-quantum dots for fluorescent western blotting assay with high sensitivity and extended dynamic range,Microchimica Acta

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Bimodal size distribution immuno-quantum dots for fluorescent western blotting assay with high sensitivity and extended dynamic range
Microchimica Acta ( IF 5.3 ) Pub Date : 2020-10-09 , DOI: 10.1007/s00604-020-04578-z
Wannian Yan ₁ , Lingzhi Fan ₁ , Jin Li ₂ , Yijiang Wang ₃ , Huanxing Han _{4,

5} , Fei Tan ₁ , Pengfei Zhang ₁

Affiliation

A highly sensitive quantum dot (QD)–based western blot assay with extended dynamic range was developed. Bimodal size distribution QD (BQ) immunoprobes composed of small size single QD (7.3 nm) and big size QD nanobead (QB) (82.9 nm) were employed for fluorescent western blot immunoassay on a membrane. Small size QD immunoprobes contributed to wider dynamic range of assay, while big size QB immunoprobes provided higher detection sensitivity. This BQ-based western blot assay can achieve a wide dynamic range (from 7.8 to 4000 ng IgG) and is nearly as sensitive as commercial available ultrasensitive chemiluminescent methods, just using a simple gel imager with UV light (365 nm) excitation and red light filter (610 nm). The fluorescent signals of BQ western blot were stable for 10 min, while chemiluminescent signals faded after 1 min. Moreover, this BQ immunoprobe was utilized for the detection of housekeeping protein and specific target proteins in complex cell lysate samples. The limit of detection of housekeeping protein is 0.25 μg of cell lysate, and the signal intensities were proportional to loading protein amount in a wide range from 0.61 to 80 μg. We believe that this new strategy of bimodal size distribution nanoparticles can also be expanded for other functional nanoparticle-based biological assays to improve the sensitivity and extend the dynamic range. Graphical abstract Graphical abstract

中文翻译：

用于荧光蛋白质印迹分析的双峰尺寸分布免疫量子点具有高灵敏度和扩展动态范围

开发了具有扩展动态范围的基于高灵敏度量子点 (QD) 的蛋白质印迹分析。由小尺寸单个 QD (7.3 nm) 和大尺寸 QD 纳米珠 (QB) (82.9 nm) 组成的双峰尺寸分布 QD (BQ) 免疫探针用于膜上的荧光蛋白质印迹免疫测定。小尺寸 QD 免疫探针有助于更宽的检测动态范围，而大尺寸 QB 免疫探针提供更高的检测灵敏度。这种基于 BQ 的蛋白质印迹分析可以实现宽动态范围（从 7.8 到 4000 ng IgG），其灵敏度几乎与市售的超灵敏化学发光方法一样灵敏，只需使用带有紫外光（365 nm）激发和红光的简单凝胶成像仪滤光片（610 纳米）。BQ蛋白质印迹的荧光信号稳定10分钟，而化学发光信号在1分钟后消失。而且，该 BQ 免疫探针用于检测复杂细胞裂解物样品中的管家蛋白和特定靶蛋白。管家蛋白的检出限为0.25 μg细胞裂解液，信号强度与上样蛋白量成正比，范围从0.61到80 μg。我们相信这种双峰尺寸分布纳米粒子的新策略也可以扩展到其他基于功能性纳米粒子的生物测定，以提高灵敏度并扩大动态范围。图形摘要图形摘要在 0.61 到 80 μg 的宽范围内，信号强度与上样蛋白量成正比。我们相信这种双峰尺寸分布纳米粒子的新策略也可以扩展到其他基于功能性纳米粒子的生物测定，以提高灵敏度并扩大动态范围。图形摘要图形摘要在 0.61 到 80 μg 的宽范围内，信号强度与上样蛋白量成正比。我们相信这种双峰尺寸分布纳米粒子的新策略也可以扩展到其他基于功能性纳米粒子的生物测定，以提高灵敏度并扩大动态范围。图形摘要图形摘要

更新日期：2020-10-09

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