摘要
将多光子激发荧光探测与毛细管电泳技术相结合,研制了多光子激发荧光 毛细管电泳联用装置。这种方法可以高效快速的分离检测复杂样品中多种不同的荧光分子。作者对5HT ,FAD ,NADH这三种重要的生物分子,不用染料标记,分别用双光子激发和三光子激发,进行了直接的分离、识别和检测。得到的检测限分别是5HT 1. 0×10 -6mol·L-1,FAD 7. 4×10 -7mol·L-1,NADH 9. 8×10 -7mol·L-1。5HT的检测限比紫外吸收低2个数量级;FAD和NADH的检测限比紫外吸收低1个数量级。
This paper describes a new method, multiphoton excitation fluorescence detection combined with capillary electrophoresis separation. The excitation source was a self mode-locked femtosecond titanium-sapphire laser (Spectra-physics Inc.), producing a stream of pulses with a pulse duration of about 100 fs at 82 MHz repetition rate. Its average power was about 200 mW at 750 nm. The laser beam was focused into a thin wall flow cell of capillary electrophoresis. A high numerical aperture objective (100 X NA 1.25) was chosen to focus the laser beam and to collect the fluorescence emitted by detecting molecules. Then the fluorescence was detected by a fast response PMT. All data were acquired and processed by a microcomputer. For three biological molecules, 5HT, FAD and NADH, it was demonstrated that they can be separated and detected efficiently by this method with three-photon and two-photon excitation respectively using only one 750 nm laser beam. The detection limits were 1.0 x 10(-6) mol (.) L-1 for 5HT, 7.4 x 10(-7) mol (.) L-1 for FAD and 9.8 x 10(-7) mol (.) L-1 for NADH using the criterion of three standard deviations above background. The results are much better than those with UV absorption detection by one or two magnitudes.
出处
《光谱学与光谱分析》
SCIE
EI
CAS
CSCD
北大核心
2005年第4期502-505,共4页
Spectroscopy and Spectral Analysis
基金
国家自然科学基金 (30 0 70 2 0 9
1 0 2 740 32 )
教育部博士点基金 (2 0 0 0 0 0 0 352 )资助项目
