Fluorescence cross-correlation spectroscopy using single wavelength laser

doi:10.1007/s11458-009-0036-5

Front Chem Chin

2009, Vol. 4

Issue (2) : 191-195 https://doi.org/10.1007/s11458-009-0036-5

RESEARCH ARTICLE

Fluorescence cross-correlation spectroscopy using single wavelength laser

Chao XIE, Chaoqing DONG, Jicun REN(

)

College of Chemistry and Chemical Engineering, Shanghai Jiao Tong University, Shanghai 200240, china

Download: PDF(163 KB) HTML
Export: BibTeX | EndNote | Reference Manager | ProCite | RefWorks

Abstract

In this paper, we first introduced the basic principle of fluorescence cross-correlation spectroscopy (FCCS) and then established an FCCS setup using a single wavelength laser. We systematically optimized the setup, and the detection volume reached about 0.7 fL. The home-built setup was successfully applied for the study of the binding reaction of human immunoglobulin G with goat antihuman immunoglobulin G. Using quantum dots (745 nm emission wavelength) and Rhodamine B (580 nm emission wavelength) as labeling probes and 532 nm laser beam as an excitation source, the cross-talk effect was almost completely suppressed. The molecule numbers in a highly focused volume, the concentration, and the diffusion time and hydrodynamic radii of the reaction products can be determined by FCCS system.

Keywords fluorescence cross-correlation spectroscopy single-molecule detection single laser excitation quantum dot

Corresponding Author(s): REN Jicun,Email:jicunren@sjtu.edu.cn

Issue Date: 05 June 2009

Cite this article:

Jicun REN,Chao XIE,Chaoqing DONG. Fluorescence cross-correlation spectroscopy using single wavelength laser[J]. Front Chem Chin, 2009, 4(2): 191-195.

URL:

https://academic.hep.com.cn/fcc/EN/10.1007/s11458-009-0036-5
https://academic.hep.com.cn/fcc/EN/Y2009/V4/I2/191

Fig.1 FCCS setup with single wavelength laser

Fig.2 Emission spectra of Rhodamine B and QD745
(1) Emission spectra of Rhodamine B, (2) Emission spectra of QD745, (3) Emission filter for channel 1,(4) Dichroic mirror, (5) Emission filter for channel 2

Fig.3 FCS curves in channel 1, 2 and FCCS curve

Fig.4 FCCS curves of reaction products (A) and relation ship between molecular number and diluting times of the solution after the binding reaction (B).
Concentration of reaction product/ (mol·L ) : (a) 1.14×10 ; (b) 2.07×10 ; (c) 4.27×10 ; (d) 1.02×10 ; e) 2.21×10

Fig.5 Normalized FCCS curves of the immune complex

1	Shao C, Hu D H, Sun H Z, Yan L K, Su Z M, Wang R S, Zhu W S, Guo J H, Sun N Z, Sun H, Li Z S, Sun C C. Structure exploration and function prediction of SARS coronavirus E protein. Chem Res Chinese U , 2005, 26(8): 1512-1516 (in Chinese)
2	Zhao J W, Wang N. Electricity characterization of metalloprotein at molecular level by conducting atomic force microscopy. Chem Res Chinese U , 2005, 26(4): 745-753 (in Chinese)
3	Nie S, Chen P, Liang S P. Identification of interacting molecules by biomolecular interaction analysis combined with surface plasmon resonance-mass spectrometry at 10-15 mol Level. Chem Res Chinese U , 2005, 26(1): 68-72 (in Chinese)
4	Heikai A A, Heikal S T, Baird G S, Tsien R Y, Webb W W. Molecular spectroscopy and dynamics of intrinsically fluorescent proteins: Coral Red (dsRed) and Yellow (Citrine). Proc Natl Acad Sci , 2000, 97(22): 11996-12001 doi: 10.1073/pnas.97.22.11996
5	Torres T, Levitus M. Measuring conformational dynamics: A new FCS-FRET approach. J Phys Chem B , 2007, 111(25): 7392-7400 doi: 10.1021/jp070659s
6	Wennmalm S, Edman L, Rigler R. Conformational fluctuations in single DNA molecules. Proc Natl Acad Sci , 1997, 94(20): 10641-10646 doi: 10.1073/pnas.94.20.10641
7	Schwille P, Meyer-Almes F J, Rigler R. Dual-color fluorescence cross-correlation spectroscopy for multicomponent diffusional analysis in solution. J Biophys , 1997, 72(4): 1878-1886 doi: 10.1016/S0006-3495(97)78833-7
8	Larson D R, Gosse J A, Holowka D A, Baird B A, Webb W W. Temporally resolved interactions between antigen-stimulated IgE receptors and lyn kinase on living cells. J Cell Biol , 2005, 171(3): 527-536 doi: 10.1083/jcb.200503110
9	Oyama R,Takashima H, Yonezawa M, Doi N, Miyamoto-Sato E, Kinjo M, Yanagawa H. Protein-protein interaction analysis by c-terminally specific fluorescence labeling and fluorescence cross-correlation spectroscopy. Nucleic Acids Res , 2006, 34(14): e102 doi: 10.1093/nar/gkl477
10	Kettling U, Koltermann A, SchwilleP, Eigen M. Real-time enzyme kinetics monitored by dual-color fluorescence cross-correlation spectroscopy. Proc Natl Acad Sci , 1998, 95(4): 1416-1420 doi: 10.1073/pnas.95.4.1416
11	Rarbach M, Kettling U, Koltermann A, Eigen M. Dual-color fluorescence cross-correlation spectroscopy for monitoring the kinetics of enzyme-catalyzed reactions. Methods , 2001, 24(2): 104-116 doi: 10.1006/meth.2001.1172
12	Bacia K, Schwille P. Dynamic view of cellular processes by in vivo fluorescence auto- and cross-correlation spectroscopy. Methods , 2003, 29 (1): 74-85 doi: 10.1016/S1046-2023(02)00291-8
13	Bacia K, Kim S A, Schwille P. Fluorescence cross-correlation spectroscopy in living cells. Nat Methods , 2006, 3(2): 83-89 doi: 10.1038/nmeth822
14	Kim S A, Heinze K G, Waxham M N, Schwille P. Intracellular calmodulin availability accessed with two-photon cross-correlation. Proc Natl Acad Sci , 2004, 101(1): 105-110 doi: 10.1073/pnas.2436461100
15	Wohland T, Hwang L C. Recent advances in fluorescence cross-correlation spectroscopy. Cell Biochem Biophys , 2007, 49(1): 1-13 doi: 10.1007/s12013-007-0042-5
16	Magde D, Elson E L, Webb W W. Thermodynamic fluctuations in a reacting system: measurements by fluorescence correlation spectroscopy. Phys Rev Lett , 1972, 29(11): 705-708 doi: 10.1103/PhysRevLett.29.705
17	Rigler R, U Mets. Diffusion of single molecules through a gaussian laser beam. Soc Photo-Opt Instrum Eng , 1992, 1921 (1): 239-248
18	Rigler R, Mets U, Widengren J, Kask P. Fluorescence correlation spectroscopy with high count rates and low background: analysis of translational diffusion. Eur Biophys J , 1993, 22(3): 169-175 doi: 10.1007/BF00185777
19	Eigen M, Rigler R. Sorting Single Molecules: Application to diagnostics and evolutionary biotechnology. Proc Natl Acad Sci , 1994, 91(13): 5740-5747 doi: 10.1073/pnas.91.13.5740
20	Zhang P D, Ren J C. Advances in fluorescence correlation spectroscopy and its applications in single molecule detection. Chinese J Anal Chem , 2005, 33(6): 875-880 (in Chinese)
21	Dong C Q, Bi R, Qian H F, Li L, RenJ C. Coupling fluorescence correlation spectroscopy with microchip electrophoresis to determine the effective surface charge of water-soluble quantum dots. Small , 2006, 2(4): 534-538 doi: 10.1002/smll.200500456
22	Dong C Q, Qian H F, Fang N F, Ren J C. Study of fluorescence quenching and dialysis process of cdte quantum dots, using ensemble techniques and fluorescence correlation spectroscopy. J Phys Chem B , 2006, 110(23): 11069-11075 doi: 10.1021/jp060279r
23	Dong C Q, Zhang P D, Bi R, Ren J C. Characterization of solution-phase DNA hybridization by fluorescence autocorrelation spectroscopy: genotyping of C677T from methylene tetrahydrofolate reductase gene. Talanta , 2007, 70(71): 1192-1197 doi: 10.1016/j.talanta.2006.06.019
24	Dong C Q, Ren J C. On-line investigation of laser-induced aggregation and photo-activation of cdte quantum dots by fluorescence correlation spectroscopy. J Phys Chem C , 2007, 111(22): 7918-7923 doi: 10.1021/jp071082h
25	Qian H F, Dong C Q, Peng J L, Qiu X, Xu Y H, Ren J C. High-quality and water-soluble near-infrared photoluminescent CdHgTe/CdS quantum dots prepared by adjusting size and composition. J Phys Chem C , 2007, 111 (45): 16852-16857 doi: 10.1021/jp074961c

[1]	Guanjiao CHEN, Wenjin ZHANG, Xinhua ZHONG, . Single-source precursor route for overcoating CdS and ZnS shells around CdSe core nanocrystals[J]. Front. Chem. China, 2010, 5(2): 214-220.
[2]	He Rong, You Xiaogang, Tian Hongye, Gao Feng, Cui Daxiang, Gu Hongchen. Synthesis and characterization of monodisperse CdSe quantum dots different organic solvents[J]. Front. Chem. China, 2006, 1(4): 378-383.
[3]	Tian Hongye, Shao Jun, He Rong, Gao Feng, Cui Daxiang, Gu Hongchen. Preparation and properties of polymer and quantum dot composites[J]. Front. Chem. China, 2006, 1(4): 474-478.

Viewed

Full text

Abstract

Cited

Shared

Discussed