Abstract

We report on real-time Raman spectroscopic studies of optically trapped living cells and organelles using an inverted confocal laser-tweezers-Raman-spectroscopy (LTRS) system. The LTRS system was used to hold a single living cell in a physiological solution or to hold a functional organelle within a living cell and consequently measured its Raman spectra. We have measured the changes in Raman spectra of a trapped yeast cell as the function of the temperature of the bathing solution and studied the irreversible cell degeneration during the heat denaturation. In addition, we measured the in-vitro Raman spectra of the nuclei within living pine cells and B. sporeformer, Strep. salivarius, and E. coli bacteria suspended in solution and showed the possibility of using LTRS system as a sensor for rapid identification of microbes in a fluid.

© 2004 Optical Society of America

Full Article  |  PDF Article

References

  • View by:
  • |

  1. Ashkin, J. M. Dziedzic, J. E. Bjorkholm, & S. Chu, ???Observation of a single-beam gradient force optical trap for dielectric particles,??? Opt. Lett. 11, 288-300 (1986).
    [CrossRef] [PubMed]
  2. A. Ashkin, K. M. Dziedzic, & T. Yamane, ???Optical trapping and manipulation of single cells using infrared laser beams,??? Nature 330, 769-771 (1987).
    [CrossRef] [PubMed]
  3. M. P. Sheetz ed., Methods in Cell Biology, Vol. 55, (Academic Press, San Diego, Calif., 1998).
  4. A. D. Mehta, M. Rief, J. A. Spudich, D. A. Smith, & R. M. Simmons, ???Single-molecule biomechanics with optical methods,??? Science 283, 1689-1695 (1999).
    [CrossRef] [PubMed]
  5. K. Visscher, M. J. Schnitzer, & S. M. Block, ???Single kinesin molecules studied with a molecular force clamp,??? Nature 400, 184-189 (1999).
    [CrossRef] [PubMed]
  6. J. Guck, R. Ananthakrishnan, T. J.Moon, C. C. Cunninggham, & J. Kas, ???Optical deformability of soft biological dielectrics,??? Phys. Rev. Lett. 84, 5451-5454 (2000).
    [CrossRef] [PubMed]
  7. M. E. J. Friese, T. A. Nieminen, N. R. Heckenberg, & H. Rubinsztein-Dunlop, "Optical alignment and spinning of laser-trapped microscopic particles,??? Nature 394, 348-350 (1998).
    [CrossRef]
  8. L. Paterson, M. P. MacDonald, J. Arlt, W. Sibbett, P. E. Bryant, & K. Dholakia, "Controlled rotation of optically trapped microscopic particles,??? Science 292, 912-914 (2001).
    [CrossRef] [PubMed]
  9. C. A. Xie, M. A. Dinno, & Y. Q. Li, ???Near-infrared Raman spectroscopy of single optically trapped biological cells,??? Opt. Lett. 27, 249-251 (2002).
    [CrossRef]
  10. C. A. Xie, & Y. Q. Li, ???Raman spectra and optical trapping of highly refractive and nontransparent particles,??? Appl. Phys. Lett. 81, 951-953 (2002).
    [CrossRef]
  11. K. Ajito, & K. Torimitsu, ???Near-infrared Raman spectroscopy of single particles,??? Trends Anal. Chem. 20 (5), 255-262 (2001).
    [CrossRef]
  12. M. P. Houlne, C. M. Sjostrom, R. H. Uibel, J. A. Kleimeyer, & J. M. Harris, ???Confocal Raman microscopy for monitoring chemical reactions on single optically trapped, solid-phase support particles,??? Anal. Chem. 74, 4311-4319 (2002).
    [CrossRef] [PubMed]
  13. G. J. Puppels, F. F. M. de Mul, C. Otto, J. Greve, M. Robert-Nicoud, D. J. Arndt-Jovin, & T. M. Jovin, ???Studying single living cells and chromosomes by confocal Raman microscopy,??? Nature 347, 301-303 (1990).
    [CrossRef] [PubMed]
  14. W. H. Nelson, R. Manoharan, & J. F. Sperry, ???UV resonance Raman studies of bacteria,??? Appl. Spectrosc. Rev. 27, 67-124 (1992).
    [CrossRef]
  15. C. Otto, N. M. Sijtsema, & J. Greve, ???Confocal Raman microspectroscopy of the activation of single neutrophilic granulocytes,??? Eur. Biophys. J. 27, 582-589 (1998).
    [CrossRef] [PubMed]
  16. K. C. Schuster, E. Urlaub, & J. R. Gapes, ???Single-cell analysis of bacteria by Raman microscopy: spectral information on the chemical composition of cells and on the heterogeneity in a culture,??? J. Microbiol. Meth. 42, 29-38 (2000).
    [CrossRef]
  17. B. R. Wood, B. Tait, & D. McNaughton, ???Micro-Raman characterisation of the R to T state transition of haemoglobin within a single living erythrocyte,??? Biochem. Biophys. Acta. 1539, 58-70 (2001).
    [CrossRef] [PubMed]
  18. K. Maquelin, L. P. Choo-Smith, T. van Vreeswijk, B. Smith, H. A. Bruining, H. P. Endtz, & G. J. Puppels, ???Raman spectroscopic method for identification of clinically relevant microorganisms growing on solid culture medium,??? Anal. Chem. 72, 12-19 (2000).
    [CrossRef] [PubMed]
  19. W. H. Nelson, & J. F. Sperry, Modern techniques for rapid microbiological analysis, W. Nelson ed., (VCH Publishers, New York, N.Y. 1991), pp.97-143.
  20. A. T. Tu, Raman spectroscopy in biology: principle and applications, (Wiely, New York, 1982).
  21. Y. Huang, T. Karashima, M. Yamanoto and H. Hamaguchi, ???Molecular-level pursuit of yeast mitosis by time- and space-resolved Raman spectroscopy,??? J. Raman Spectrosc. 34, 1-3 (2003).
    [CrossRef]
  22. Y. Huang, T. Karashima, M. Yamanoto, T. Ogura and H. Hamaguhci, ???Raman spectroscopic signature of life in a living yeast cell,??? J. Raman Spectrosc. 35, 525-526 (2004).
    [CrossRef]

Anal. Chem. (2)

M. P. Houlne, C. M. Sjostrom, R. H. Uibel, J. A. Kleimeyer, & J. M. Harris, ???Confocal Raman microscopy for monitoring chemical reactions on single optically trapped, solid-phase support particles,??? Anal. Chem. 74, 4311-4319 (2002).
[CrossRef] [PubMed]

K. Maquelin, L. P. Choo-Smith, T. van Vreeswijk, B. Smith, H. A. Bruining, H. P. Endtz, & G. J. Puppels, ???Raman spectroscopic method for identification of clinically relevant microorganisms growing on solid culture medium,??? Anal. Chem. 72, 12-19 (2000).
[CrossRef] [PubMed]

Appl. Phys. Lett. (1)

C. A. Xie, & Y. Q. Li, ???Raman spectra and optical trapping of highly refractive and nontransparent particles,??? Appl. Phys. Lett. 81, 951-953 (2002).
[CrossRef]

Appl. Spectrosc. Rev. (1)

W. H. Nelson, R. Manoharan, & J. F. Sperry, ???UV resonance Raman studies of bacteria,??? Appl. Spectrosc. Rev. 27, 67-124 (1992).
[CrossRef]

Biochem. Biophys. Acta. (1)

B. R. Wood, B. Tait, & D. McNaughton, ???Micro-Raman characterisation of the R to T state transition of haemoglobin within a single living erythrocyte,??? Biochem. Biophys. Acta. 1539, 58-70 (2001).
[CrossRef] [PubMed]

Eur. Biophys. J. (1)

C. Otto, N. M. Sijtsema, & J. Greve, ???Confocal Raman microspectroscopy of the activation of single neutrophilic granulocytes,??? Eur. Biophys. J. 27, 582-589 (1998).
[CrossRef] [PubMed]

J. Microbiol. Meth. (1)

K. C. Schuster, E. Urlaub, & J. R. Gapes, ???Single-cell analysis of bacteria by Raman microscopy: spectral information on the chemical composition of cells and on the heterogeneity in a culture,??? J. Microbiol. Meth. 42, 29-38 (2000).
[CrossRef]

J. Raman Spectrosc. (2)

Y. Huang, T. Karashima, M. Yamanoto and H. Hamaguchi, ???Molecular-level pursuit of yeast mitosis by time- and space-resolved Raman spectroscopy,??? J. Raman Spectrosc. 34, 1-3 (2003).
[CrossRef]

Y. Huang, T. Karashima, M. Yamanoto, T. Ogura and H. Hamaguhci, ???Raman spectroscopic signature of life in a living yeast cell,??? J. Raman Spectrosc. 35, 525-526 (2004).
[CrossRef]

Nature (4)

G. J. Puppels, F. F. M. de Mul, C. Otto, J. Greve, M. Robert-Nicoud, D. J. Arndt-Jovin, & T. M. Jovin, ???Studying single living cells and chromosomes by confocal Raman microscopy,??? Nature 347, 301-303 (1990).
[CrossRef] [PubMed]

A. Ashkin, K. M. Dziedzic, & T. Yamane, ???Optical trapping and manipulation of single cells using infrared laser beams,??? Nature 330, 769-771 (1987).
[CrossRef] [PubMed]

K. Visscher, M. J. Schnitzer, & S. M. Block, ???Single kinesin molecules studied with a molecular force clamp,??? Nature 400, 184-189 (1999).
[CrossRef] [PubMed]

M. E. J. Friese, T. A. Nieminen, N. R. Heckenberg, & H. Rubinsztein-Dunlop, "Optical alignment and spinning of laser-trapped microscopic particles,??? Nature 394, 348-350 (1998).
[CrossRef]

Opt. Lett. (2)

Phys. Rev. Lett. (1)

J. Guck, R. Ananthakrishnan, T. J.Moon, C. C. Cunninggham, & J. Kas, ???Optical deformability of soft biological dielectrics,??? Phys. Rev. Lett. 84, 5451-5454 (2000).
[CrossRef] [PubMed]

Science (2)

L. Paterson, M. P. MacDonald, J. Arlt, W. Sibbett, P. E. Bryant, & K. Dholakia, "Controlled rotation of optically trapped microscopic particles,??? Science 292, 912-914 (2001).
[CrossRef] [PubMed]

A. D. Mehta, M. Rief, J. A. Spudich, D. A. Smith, & R. M. Simmons, ???Single-molecule biomechanics with optical methods,??? Science 283, 1689-1695 (1999).
[CrossRef] [PubMed]

Trends Anal. Chem. (1)

K. Ajito, & K. Torimitsu, ???Near-infrared Raman spectroscopy of single particles,??? Trends Anal. Chem. 20 (5), 255-262 (2001).
[CrossRef]

Other (3)

M. P. Sheetz ed., Methods in Cell Biology, Vol. 55, (Academic Press, San Diego, Calif., 1998).

W. H. Nelson, & J. F. Sperry, Modern techniques for rapid microbiological analysis, W. Nelson ed., (VCH Publishers, New York, N.Y. 1991), pp.97-143.

A. T. Tu, Raman spectroscopy in biology: principle and applications, (Wiely, New York, 1982).

Cited By

OSA participates in CrossRef's Cited-By Linking service. Citing articles from OSA journals and other participating publishers are listed here.

Alert me when this article is cited.


Figures (4)

Fig. 1.
Fig. 1.

Experimental setup of the combined laser tweezers and Raman spectroscopy system. Abbreviations: M, mirror; L, lens; DM, dichroic mirror; PH, pinhole; HNF, holograph notch filter.

Fig. 2.
Fig. 2.

NIR Raman spectra of (a) a single trapped nucleus of a pine cell, (b) Bacillus sporeformer, (c) Streptococcus salivarius, and (d) E. coli bacterium suspended in a liquid medium. The pine nucleus is moved to the left from near the center of the cell, as indicated by the arrow in the image (a). The background spectrum measured without the cell in the trap has been subtracted for each spectrum. Spectrum (a) is average result of eight pine cells and spectrum (b–d) is the average result of 20 bacterial cells, respectively. Experimental conditions: trapping power 5 mW, Raman excitation power 15 mW; acquisition time 30 s for curve (a) and 60 s for curves (b, c, d). Raman line assignments are based on assignments in [13, 15, 16, 18]. Abbreviations: Phe, phenylalanine; Tyr, tyrosine; A, adenine; C, cytosine; G, guanine; BK, backbone; p, protein; def, deformation; str, stretching.

Fig. 3.
Fig. 3.

Raman spectra of a single yeast cell at different temperatures. A living yeast cell is held in the beam focus while the medium’s temperature is changed slowly. Experimental conditions: trapping power 2 mW, Raman excitation power 16 mW; acquisition time 20 s. Raman band assignments are based on the assignments in [13, 15, 16, 18].

Fig. 4.
Fig. 4.

Signal intensity of the 1004 cm-1 band as the function of temperature. The smooth curve is an exponential function fit.

Metrics