Bimodal size distribution immuno-quantum dots for fluorescent western blotting assay with high sensitivity and extended
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ORIGINAL PAPER
Bimodal size distribution immuno-quantum dots for fluorescent western blotting assay with high sensitivity and extended dynamic range Wannian Yan 1 & Lingzhi Fan 1 & Jin Li 2 & Yijiang Wang 3 & Huanxing Han 4,5 & Fei Tan 1 & Pengfei Zhang 1 Received: 2 June 2020 / Accepted: 29 September 2020 # Springer-Verlag GmbH Austria, part of Springer Nature 2020
Abstract 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.
Keywords Quantum dots . Nanobeads . Western blotting . Highly sensitive immunoassay . Extended dynamic range
Wannian Yan, Lingzhi Fan and Jin Li contributed equally to this work. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00604-020-04578-z) contains supplementary material, which is available to authorized users. * Fei Tan [email protected] * Pengfei Zhang [email protected] 1
Department of Central Laboratory, Shanghai Skin Disease Hospital, Tongji University School of Medicine, Shanghai 200443, China
2
Shandong Zhifu Hospital, Yantai 26400, Shandong, China
3
Department of Periodontology, School & Hospital of Stomatology, Shanghai Engineering Research Center of Tooth Restoration and Regeneration, Tongji University, Shanghai 200072, China
4
Department of Pharmacy, Changzheng Hospital, The Second Military Medical University, Shanghai 200433, China
5
Aliex Technology Group Co., Ltd, No. 152, Lane 468, North Hengshahe Road, Shanghai, China
Chemiluminescent western blotting assay offers high sensitivity and easy operation for specific protein detection, while it still suffers from poor reproducibility and less precisely quantitative results [1–3]. Fluorescent western blot assay is emerging as a new trend due to its improved repr
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