Highlighted Text:
Show |
Hide
Full text loading...
Chapter 13 : Raman-Fluorescence In Situ Hybridization
Authors:
Daniel S. Read1,
Andrew S. Whiteley2
VIEW AFFILIATIONS
HIDE AFFILIATIONS
Affiliations:
1: Centre for Ecology and Hydrology, Benson Lane, Wallingford, OX10 8BB, United Kingdom;
2: Centre for Ecology and Hydrology, Benson Lane, Wallingford, OX10 8BB, United Kingdom
Content Type: Monograph
Publication Year:
2011
Category: Environmental Microbiology; Applied and Industrial Microbiology
Book DOI:
10.1128/9781555816896
Chapter DOI:
10.1128/9781555816896.ch13
MyBook is a cheap paperback edition of the original book and will be sold at uniform, low price.
Preview this chapter:
Raman-Fluorescence In Situ Hybridization, Page 1 of 2
< Previous page | Next page > /docserver/preview/fulltext/10.1128/9781555816896/9781555815370_Chap13-1.gif /docserver/preview/fulltext/10.1128/9781555816896/9781555815370_Chap13-2.gif
Abstract:
As with many other molecular microbiology methods, the family of stable isotope probing (SIP) techniques based on the analysis of isotope- labeled nucleic acids, phospholipid fatty acids (PLFAs), or proteins are frequently performed at the level of the community. Single-cell studies encompass a range of techniques, including bacterial bioreporters, flow cytometry, fluorescence in situ hybridization (FISH) in combination with microautoradiography (MAR), nanoscale secondary ion mass spectrometry (nanoSIMS), and Raman spectroscopy. In general, FISH procedures are almost identical to standard epifluorescence microscopic procedures. The ability to analyze microbial communities at the level of the single cell, rather than at the level of the population or community, is one of the main advantages of Raman, and the area where it differs from conventional SIP techniques. Conventional SIP techniques tend to focus on isotope incorporation into one specific biomolecule, such as PLFAs, DNA, RNA, or proteins. Raman has a significant advantage in that it is looking at isotope incorporation by all these biomolecules simultaneously, depending of course on the exact constitution and quality of the collected Raman spectra. Raman is a sensitive technique, with isotope incorporation being detected in microbes cultured in growth media containing as low as 10% 13C. Raman spectroscopy has also been combined with optical trapping and manipulation to achieve cell sorting of isotopically labeled cells.
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
Key Concept Ranking
- Microbial Ecology
- 0.73556197
- 16s rRNA Sequencing
- 0.46080306
- Raman Spectroscopy
- 0.40555432
0.73556197
Figures
Raman-Fluorescence In Situ Hybridization
[com.pub2web.rdf.cci.facet.ContentItem[id=http://asm.metastore.ingenta.com/content/author/10.1128/9781555816896.ch13-1,webId=/content/author/10.1128/9781555816896.ch13-1,properties={foaf_givenname=Daniel S., foaf_name=Daniel S. Read, foaf_surname=Read, pub_isAffiliatedWith=[com.pub2web.rdf.cci.facet.ContentItem[id=http://asm.metastore.ingenta.com/content/affiliation/10.1128/9781555816896.ch13-aff1]]}], com.pub2web.rdf.cci.facet.ContentItem[id=http://asm.metastore.ingenta.com/content/author/10.1128/9781555816896.ch13-2,webId=/content/author/10.1128/9781555816896.ch13-2,properties={foaf_givenname=Andrew S., foaf_name=Andrew S. Whiteley, foaf_surname=Whiteley, pub_isAffiliatedWith=[com.pub2web.rdf.cci.facet.ContentItem[id=http://asm.metastore.ingenta.com/content/affiliation/10.1128/9781555816896.ch13-aff2]]}]]
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig01
FIGURE 1
Jablonski diagram. A Jablonski diagram illustrates the transitions between the electronic states of a molecule and how these relate to infrared (IR) absorption, Rayleigh scattering, and Stokes and anti-Stokes Raman scattering.
10.1128/9781555816896/f0280-01_thmb.gif
10.1128/9781555816896/f0280-01.gif
FIGURE 1
Jablonski diagram. A Jablonski diagram illustrates the transitions between the electronic states of a molecule and how these relate to infrared (IR) absorption, Rayleigh scattering, and Stokes and anti-Stokes Raman scattering.
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig02
FIGURE 2
The main features of a Raman microspectrometer. Transmitted light is used to visualize microbial cells on a Raman-inert slide. Light from a monochromatic laser source is directed down the microscope objective, and Raman scattered light is collected back through the same objective. Unwanted light frequencies are filtered with a notch filter, before the light beam is diffracted using a grating and dispersed onto a cooled CCD camera. This signal can be used to create a Raman spectrum or a Raman chemical image. (
After Notinger and Hench, 2006
.)
10.1128/9781555816896/f0281-01_thmb.gif
10.1128/9781555816896/f0281-01.gif
FIGURE 2
The main features of a Raman microspectrometer. Transmitted light is used to visualize microbial cells on a Raman-inert slide. Light from a monochromatic laser source is directed down the microscope objective, and Raman scattered light is collected back through the same objective. Unwanted light frequencies are filtered with a notch filter, before the light beam is diffracted using a grating and dispersed onto a cooled CCD camera. This signal can be used to create a Raman spectrum or a Raman chemical image. ( After Notinger and Hench, 2006 .)
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig03
FIGURE 3
Raman spectra of microbial cells. Raman spectra collected from a nonpigmented bacterial cell (E. coli) and a pigmented bacterial cell (unidentified environmental isolate) using a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm.
10.1128/9781555816896/f0283-01_thmb.gif
10.1128/9781555816896/f0283-01.gif
FIGURE 3
Raman spectra of microbial cells. Raman spectra collected from a nonpigmented bacterial cell (E. coli) and a pigmented bacterial cell (unidentified environmental isolate) using a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm.
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig04
FIGURE 4
Raman spectra of labeled and unlabeled compounds. Raman spectra of unlabeled (solid line) and labeled [U-13C6] (dashed line) glucose collected on a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm.
10.1128/9781555816896/f0284-01_thmb.gif
10.1128/9781555816896/f0284-01.gif
FIGURE 4
Raman spectra of labeled and unlabeled compounds. Raman spectra of unlabeled (solid line) and labeled [U-13C6] (dashed line) glucose collected on a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm.
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig05
FIGURE 5
Isotope labeling series. Raman spectra collected from E. coli cultured in media containing 0, 50, or 100% [U-13C6]glucose as the sole carbon source. Figures are offset for clarity. Raman spectra collected on a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm. Major peak shifts are highlighted in gray.
10.1128/9781555816896/f0285-01_thmb.gif
10.1128/9781555816896/f0285-01.gif
FIGURE 5
Isotope labeling series. Raman spectra collected from E. coli cultured in media containing 0, 50, or 100% [U-13C6]glucose as the sole carbon source. Figures are offset for clarity. Raman spectra collected on a LabRAM HR800 UV confocal Raman microscope (Horiba Scientific) with an excitation wavelength of 532 nm. Major peak shifts are highlighted in gray.
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
/content/10.1128/9781555816896.ch13.ch13fig06
FIGURE 6
Outline of the major steps involved in Raman-FISH. The stable isotope is added to the environmental sample, either in a microcosm (shown) or in the field. Microbes are permeabilized and labeled with fluorescently tagged FISH probes, and epifluorescence microscopy is used to identify cells of interest. Once identified, Raman optics are switched to Raman mode to analyze uptake of stable isotope at the single-cell level.
After Amann and Fuchs (2008)
.
10.1128/9781555816896/f0286-01_thmb.gif
10.1128/9781555816896/f0286-01.gif
FIGURE 6
Outline of the major steps involved in Raman-FISH. The stable isotope is added to the environmental sample, either in a microcosm (shown) or in the field. Microbes are permeabilized and labeled with fluorescently tagged FISH probes, and epifluorescence microscopy is used to identify cells of interest. Once identified, Raman optics are switched to Raman mode to analyze uptake of stable isotope at the single-cell level. After Amann and Fuchs (2008) .
Citation:
Read D, Whiteley A.
2011.
Raman-Fluorescence In Situ Hybridization,
p 279-294.
In
Murrell J, Whiteley A (ed),
Stable Isotope Probing and Related Technologies.
ASM Press, Washington, DC.
doi: 10.1128/9781555816896.ch13
References
/content/book/10.1128/9781555816896.ch13
1.
Amann, R., and, B. M. Fuchs. 2008. Single-cell Identification in microbial communities by improved fluorescence in situ hybridization techniques. Nat. Rev. Microbiol. 6: 339– 348.
2.
Basar, G., and, S. Kin. 2008. Monitoring of spectroscopic changes of a single trapped fission yeast cell by using a Raman tweezers set-up. Opt. Commun. 281: 4998– 5003.
3.
Behrens, S.,, T. Losekann,, J. Pett-Ridge,, P. K. Weber,, W. O. Ng,, B. S. Stevenson,, I. D. Hutcheon,, D. A. Relman, and, A. M. Spormann. 2008. Linking microbial phylogeny to metabolic activity at the single-cell level by using enhanced element labeling-catalyzed reporter deposition fluorescence in situ hybridization (EL-FISH) and NanoSIMS. Appl. Environ. Microbiol. 74: 3143– 3150.
4.
Bertaux, J.,, U. Gloger,, M. Schmid,, A. Hartmann, and, S. Scheu. 2007. Routine fluorescence in situ hybridization in soil. J. Microbiol. Methods 69: 451– 460.
5.
Camp, C. H.,, S. Yegnanarayanan,, A. A. Eftekhar,, H. Sridhar, and, A. Adibi. 2009. Multiplex coherent anti-Stokes Raman scattering (MCARS) for chemically sensitive, label-free flow cytometry. Opt. Express 17: 22879– 22889.
6.
Chan, J. W.,, H. Winhold,, M. H. Corzett,, J. M. Ulloa,, M. Cosman,, R. Balhorn, and, T. Huser. 2007. Monitoring dynamic protein expression in living E-coli bacterial cells by laser tweezers Raman spectroscopy. Cytom. Part A 71A: 468– 474.
7.
Curtis, T. P.,, W. T. Sloan, and, J. W. Scannell. 2002. Estimating prokaryotic diversity and its limits. Proc. Natl. Acad. Sci. USA 99: 10494– 10499.
8.
Davidson, C. J., and, M. G. Surette. 2008. Individuality in bacteria. Annu. Rev. Genet. 42: 253– 268.
9.
de Oliveira, V.E.,, Castro, H.V.,, Edwards, H.G.M.,, De Oliveira, L.F.C. 2010. Carotenes and carotenoids in natural biological samples: a Raman spectroscopic analysis. J. Raman Spectrosc 41: 642– 650.
10.
Fuhrman, J. A. 2009. Microbial community structure and its functional implications. Nature 459: 193– 199.
11.
Huang, W. E.,, A. Ferguson,, A. C. Singer,, K. Law-Son,, I. P. Thompson,, R. M. Kalin,, M. J. Larkin,, M. J. Bailey, and, A. S. Whiteley. 2009. Resolving genetic functions within microbial populations: in situ analyses using rRNA and mRNA stable isotope probing coupled with single-cell Raman-fluorescence in situ hybridization. Appl. Environ. Microbiol. 75: 234– 241.
12.
Huang, W. E.,, R. I. Griffiths,, I. P. Thompson,, M. J. Bailey, and, A. S. Whiteley. 2004. Raman microscopic analysis of single microbial cells. Anal. Chem. 76: 4452– 4458.
13.
Huang, W. E.,, K. Stoecker,, R. Griffiths,, L. New-Bold,, H. Daims,, A. S. Whiteley, and, M. Wagner. 2007. Raman-FISH: combining stable-isotope Raman spectroscopy and fluorescence in situ hybridization for the single cell analysis of identity and function. Environ. Microbiol. 9: 1878– 1889.
14.
Huang, W. E.,, A. D. Ward, and, A. S. Whiteley. 2009. Raman tweezers sorting of single microbial cells. Environ. Microbiol. Rep. 1: 44– 49.
15.
Hutsebaut, D.,, P. Vandenabeele, and, L. Moens. 2005. Evaluation of an accurate calibration and spectral standardization procedure for Raman spectros-copy. Analyst 130: 1204– 1214.
16.
Jaspers, E., and, J. Overmann. 2004. Ecological significance of microdiversity: identical 16S rRNA gene sequences can be found in bacteria with highly divergent genomes and ecophysiologies. Appl. Environ. Microbiol. 70: 4831– 4839.
17.
Montoya, J. P.,, C. M. Holl,, J. P. Zehr,, A. Hansen,, T. A. Villareal, and, D. G. Capone. 2004. High rates of N2 fixation by unicellular diazotrophs in the oligotrophic Pacific Ocean. Nature 430: 1027– 1031.
18.
Nile, S. M., and, S. R. Emery. 1997. Probing single molecules and single nanoparticles by surface-enhanced Raman scattering. Science 275: 1102– 1106.
19.
Notinger, I., and, L. L. Hench. 2006. Raman microspectroscopy: a noninvasive tool for studies of individual living cells in vitro. Expert Rev. Med. Dev. 3: 215– 234.
20.
Orphan, V. J.,, K. U. Hinrichs,, W. Ussler,, C. K. Paull,, L. T. Taylor,, S. P. Sylva,, J. M. Hayes, and, E. F. DeLong. 2001. Comparative analysis of methane-oxidizing archaea and sulfate-reducing bacteria in anoxic marine sediments. Appl. Environ. Microbiol. 67: 1922– 1934.
21.
PedróS-Alió, C. 2006. Marine microbial diversity: can it be determined? Trends Microbiol. 14: 257– 263.
22.
Raman, C. V., and, K. S. Krishnan. 1928. A new type of secondary radiation. Nature 121: 501.
23.
Tringe, S. G.,, C. von Mering,, A. Kobayashi,, A. A. Salamov,, K. Chen,, H. W. Chang,, M. Podar,, J. M. Short,, E. J. Mathur,, J. C. Detter,, P. Bork,, P. Hugenholtz, and, E. M. Rubin. 2005. Comparative metagenomics of microbial communities. Science 308: 554– 557.
24.
van Manen, H. J.,, Y. M. Kraan,, D. Roos, and, C. Otto. 2005. Single-cell Raman and fluorescence microscopy reveal the association of lipid bodies with phagosomes in leukocytes. P. Natl. Acad. Sci. USA 102: 10159– 10164.
25.
van Manen, H. J.,, A. Lenferink, and, C. Otto. 2008. Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy. Anal. Chem. 80: 9576– 9582.
26.
Wachsmann-Hogiu, S.,, T. Weeks, and, T. Huser. 2009. Chemical analysis in vivo and in vitro by Raman spectroscopy-from single cells to humans. Curr. Opin. Biotechnol. 20: 63– 73.
27.
Watson, D. A.,, L. O. Brown,, D. R. Gaskill,, M. Naivar,, S. W. Graves,, S. K. Doorn, and, J. P. Nolan. 2008. A flow cytometer for the measurement of Raman spectra. Cytom. Part A 73A: 119– 128.
28.
Xie, C. G.,, C. Goodman,, M. A. Dinno, and, Y. Q. Li. 2004. Real-time Raman spectroscopy of optically trapped living cells and organelles. Opt. Express 12: 6208– 6214.
29.
Zwirglmaier, K. 2005. Fluorescence in situ hybridisation (FISH)—the next generation. FEMS Microbiol. Lett. 246: 151– 158.