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Spectral imaging of single molecules by transmission grating-based epi-fluorescence microscopy |
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| Authors: | Han Rui Zhang YeWang Dong Xiuling Gai Hongwei Yeung Edward S |
| Institution: | a Biomedical Engineering Center, Hunan University, Changsha, Hunan 410082, PR China b School of Pharmaceutics, Jiangsu University, Xuefu Road 301, Zhenjiang 212013, PR China c Dalian the Third Municipal Hospital, Dalian 116033, PR China d State Key Laboratory of Chemo/Biosensing and Chemometrics, Hunan University, Changsha 410082, PR China e Department of Chemistry and Ames Laboratory, Iowa State University, Ames, IA 50011, USA |
| Abstract: | A spectral imaging method of single protein molecules labeled with a single fluorophore is presented. The method is based on a transmission grating and a routine fluorescence microscope. The bovine serum alubmin (BSA) and antiBSA molecules labeled with Alexa Fluor 488 and Alexa Fluor 594, respectively, are used as the model proteins. The fluorescence of single molecules is dispersed into zeroth-order spectrum and first-order spectrum by the transmission grating. Results show that the fluorescence emission spectrum of single molecule converted from the first-order spectral imaging is in good agreement with the bulk fluorescence spectrum. The spectral resolution of 2.4 nm/pixel is obtained, which is sufficient for identifying the molecular species in a multicomponent system. |
| Keywords: | Single protein molecule First-order spectrum Transmission grating Fluorescence microscopy |
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