Plasmonic-based instrument response function for time-resolved fluorescence: toward proper lifetime analysis

  • Radoslaw Szlazak
  • Krzysztof Tutaj
  • Wojciech Grudzinski
  • Wieslaw I. Gruszecki
  • Rafal Luchowski
Research Paper


In this report, we investigated the so-called plasmonic platforms prepared to target ultra-short fluorescence and accurate instrumental response function in a time-domain spectroscopy and microscopy. The interaction of metallic nanoparticles with nearby fluorophores results in the increase of the dye fluorescence quantum yield, photostability and decrease of the lifetime parameter. The mentioned properties of platforms were applied to achieve a picosecond fluorescence lifetime (21 ps) of erythrosin B, used later as a better choice for deconvolution of fluorescence decays measured with “color” sensitive photo-detectors. The ultra-short fluorescence standard based on combination of thin layers of silver film, silver colloidal nanoparticles (about 60 nm in diameter), and top layer of erythrosin B embedded in 0.2 % poly(vinyl) alcohol. The response functions were monitored on two photo-detectors; microchannel plate photomultiplier and single photon avalanche photodiode as a Rayleigh scattering and ultra-short fluorescence. We demonstrated that use of the plasmonic base fluorescence standard as an instrumental response function results in the absence of systematic error in lifetime measurements and analysis.


Instrument response function Time-resolved spectroscopy Fluorescence lifetime Surface plasmons Nanoparticles 



Time correlated single photon counting


Instrument response function


Microchannel plate photomultiplier tube


Single photon avalanche photodiodes


Erythrosin B


Poly(vinyl) alcohol


Surface plasmon


Plasmonic platform





This work was supported by National Science Centre (Grant # N N202 112340) and Foundation for Polish Science within the project of “Molecular Spectroscopy for BioMedical Studies.” The authors kindly acknowledge prof. Marek Tchorzewski and the “NanoFun” network for access to the fluorescence microscopy facility.


  1. Bastiaens PI, van Hoek A, Wolkers WF, Brochon JC, Visser AJ (1992) Comparison of the dynamical structures of lipoamide dehydrogenase and glutathione reductase by time-resolved polarized flavin fluorescence. Biochemistry 31:7050–7060CrossRefGoogle Scholar
  2. Boens N, Qin W, Basarić N, Hofkens J, Ameloot M, Pouget J, Lefèvre J-P, Valeur B, Gratton E, vande Ven M, Silva ND Jr, Engelborghs Y, Willaert K, Sillen A, Rumbles G, Phillips D, Visser AJWG, van Hoek A, Lakowicz JR, Malak H, Gryczynski I, Szabo AG, Krajcarski DT, Tamai N, Miura A (2007) Fluorescence lifetime standards for time and frequency domain fluorescence spectroscopy. Anal Chem 79:2137–2149. doi: 10.1021/ac062160k CrossRefGoogle Scholar
  3. Camden JP, Dieringer JA, Wang Y, Masiello DJ, Marks LD, Schatz GC, Van Duyne RP (2008) Probing the structure of single-molecule surface-enhanced Raman scattering hot spots. J Am Chem Soc 130:12616–12617. doi: 10.1021/ja8051427 CrossRefGoogle Scholar
  4. Carminati R, Greffet JJ, Henkel C, Vigoureux JM (2006) Radiative and non-radiative decay of a single molecule close to a metallic nanoparticle. Opt Commun 261:368–375. doi: 10.1016/j.optcom.2005.12.009 CrossRefGoogle Scholar
  5. Chizhik A, Schleifenbaum F, Gutbrod R, Chizhik A, Khoptyar D, Meixner AJ, Enderlein J (2009) Tuning the fluorescence emission spectra of a single molecule with a variable optical subwavelength metal microcavity. Phys Rev Lett 102:073002CrossRefGoogle Scholar
  6. Conn PM (2012) Imaging and spectroscopic analysis of living cells: optical and spectroscopic techniques. Academic Press, San DiegoGoogle Scholar
  7. Gersten J, Nitzan A (1981) Spectroscopic properties of molecules interacting with small dielectric particles. J Chem Phys 75:1139–1152. doi: 10.1063/1.442161 CrossRefGoogle Scholar
  8. Gryczynski Z, Bucci E (1993) A new front-face optical cell for measuring weak fluorescent emissions with time resolution in the picosecond time scale. Biophys Chem 48:31–38CrossRefGoogle Scholar
  9. Gryczynski I, Kusba J, Lakowicz JR (1994) Light quenching of fluorescence using time-delayed laser pulses as observed by frequency-domain fluorometry. J Phys Chem 98:8886–8895. doi: 10.1021/j100087a012 CrossRefGoogle Scholar
  10. Gryczynski I, Hell SW, Lakowicz JR (1997) Light quenching of pyridine2 fluorescence with time-delayed pulses. Biophys Chem 66:13–24. doi: 10.1016/S0301-4622(96)02264-8 CrossRefGoogle Scholar
  11. Gryczynski I, Malicka J, Nowaczyk K, Gryczynski Z, Lakowicz JR (2004) Effects of sample thickness on the optical properties of surface plasmon-coupled emission. J Phys Chem B 108:12073–12083. doi: 10.1021/jp0312619 CrossRefGoogle Scholar
  12. Kawski A, Gryczynski I, Nowaczyk K, Bojarski P, Lichacz J (1991) Deactivation of the S1-state of ω-substituted 4-dimethylamino-trans-styrenes in alkane solutions. J Phys Sci 46:1043–1048Google Scholar
  13. Kawski A, Gryczynski J, Gryczynski Z (1993) Fluorescence anisotropies of 4-dimethylamino-ω-diphenylphosphinyl-trans-styrene in isotropic media in the case of one- and two-photon excitation. J Phys Sci 48:551–556Google Scholar
  14. Lakowicz JR (2001) Radiative decay engineering: biophysical and biomedical applications. Anal Biochem 298:1–24. doi: 10.1006/abio.2001.5377 CrossRefGoogle Scholar
  15. Lakowicz JR (2006) Radiative decay engineering: metal-enhanced fluorescence. In: Geddes CD (ed) Principles of fluorescence spectroscopy. Springer, Boston, pp 841–859CrossRefGoogle Scholar
  16. Lakowicz JR, Gryczynski I, Laczko G, Gloyna D (1991) Picosescond fluorescence lifetime standards for frequency- and time-domain fluorescence. J Fluoresc 1:87–93. doi: 10.1007/BF00865204 CrossRefGoogle Scholar
  17. Lakowicz JR, Shen Y, D’Auria S, Malicka J, Fang J, Gryczynski Z, Gryczynski I (2002) Radiative decay engineering. 2. Effects of silver island films on fluorescence intensity, lifetimes, and resonance energy transfer. Anal Biochem 301:261–277. doi: 10.1006/abio.2001.5503 CrossRefGoogle Scholar
  18. Luchowski R, Gryczynski Z, Sarkar P, Borejdo J, Szabelski M, Kapusta P, Gryczynski I (2009a) Instrument response standard in time-resolved fluorescence. Rev Sci Instrum 80(3):033109. doi: 10.1063/1.3095677 CrossRefGoogle Scholar
  19. Luchowski R, Kapusta P, Szabelski M, Sarkar P, Borejdo J, Gryczynski Z, Gryczynski I (2009b) Forster resonance energy transfer (FRET)-based picosecond lifetime reference for instrument response evaluation. Meas Sci Technol 20:095601. doi: 10.1088/0957-0233/20/9/095601 CrossRefGoogle Scholar
  20. Luchowski R, Calander N, Shtoyko T, Apicella E, Borejdo J, Gryczynski Z, Gryczynski I (2010a) Plasmonic platforms of self-assembled silver nanostructures in application to fluorescence. J Nanophotonics. doi: 10.1117/1.3500463
  21. Luchowski R, Sabnis S, Szabelski M, Sarkar P, Raut S, Gryczynski Z, Borejdo J, Bojarski P, Gryczynski I (2010b) Self-quenching of uranin: instrument response function for color sensitive photo-detectors. J Lumin 130:2446–2451. doi: 10.1016/j.jlumin.2010.07.027 CrossRefGoogle Scholar
  22. Mukerjee A, Luchowski R, Ranjan AP, Raut S, Vishwanatha JK, Gryczynski Z, Gryczynski I (2010) Enhanced fluorescence of curcumin on plasmonic platforms. Curr Pharm Biotechnol 11:223–228CrossRefGoogle Scholar
  23. Pyatenko A, Yamaguchi M, Suzuki M (2007) Synthesis of spherical silver nanoparticles with controllable sizes in aqueous solutions. J Phys Chem C 111:7910–7917. doi: 10.1021/jp071080x CrossRefGoogle Scholar
  24. Sorensen TJ, Laursen BW, Luchowski R, Shtoyko T, Akopova I, Gryczynski Z, Gryczynski I (2009) Enhanced fluorescence emission of Me-ADOTA(+) by self-assembled silver nanoparticles on a gold film. Chem Phys Lett 476:46–50. doi: 10.1016/j.cplett.2009.05.064 CrossRefGoogle Scholar
  25. Szabelski M, Ilijev D, Sarkar P, Luchowski R, Gryczynski Z, Kapusta P, Erdmann R, Gryczynski I (2009a) Collisional quenching of erythrosine b as a potential reference dye for impulse response function evaluation. Appl Spectrosc 63:363–368CrossRefGoogle Scholar
  26. Szabelski M, Luchowski R, Gryczynski Z, Kapusta P, Ortmann U, Gryczynski I (2009b) Evaluation of instrument response functions for lifetime imaging detectors using quenched Rose Bengal solutions. Chem Phys Lett 471:153–159. doi: 10.1016/j.cplett.2009.02.001 CrossRefGoogle Scholar
  27. Van den Berg PA, Mulrooney SB, Gobets B, van Stokkum IH, van Hoek A, Williams CH Jr, Visser AJ (2001) Exploring the conformational equilibrium of E. coli thioredoxin reductase: characterization of two catalytically important states by ultrafast flavin fluorescence spectroscopy. Protein Sci 10:2037–2049. doi: 10.1110/ps.06701 CrossRefGoogle Scholar
  28. Van Den Zegel M, Boens N, Daems D, De Schryver FC (1986) Possibilities and limitations of the time-correlated single photon counting technique: a comparative study of correction methods for the wavelength dependence of the instrument response function. Chem Phys 101:311–335. doi: 10.1016/0301-0104(86)85096-0 CrossRefGoogle Scholar
  29. Weitz DA, Garoff S, Gersten JI, Nitzan A (1983) The enhancement of Raman scattering, resonance Raman scattering, and fluorescence from molecules adsorbed on a rough silver surface. J Chem Phys 78:5324–5338. doi: 10.1063/1.445486 CrossRefGoogle Scholar
  30. Zuker M, Szabo A, Bramall L, Krajcarski D, Selinger B (1985) Delta-function convolution method (DFCM) for fluorescence decay experiments. Rev Sci Instrum 56:14–22. doi: 10.1063/1.1138457 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Radoslaw Szlazak
    • 1
  • Krzysztof Tutaj
    • 1
  • Wojciech Grudzinski
    • 1
  • Wieslaw I. Gruszecki
    • 1
  • Rafal Luchowski
    • 1
  1. 1.Department of Biophysics, Institute of PhysicsMaria Curie-Sklodowska UniversityLublinPoland

Personalised recommendations