Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy

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Abstract

We have combined nonresonant Raman microspectroscopy and spectral imaging with stable isotope labeling by amino acids in cell culture (SILAC) to selectively detect the incorporation of deuterium-labeled phenylalanine, tyrosine, and methionine into proteins in intact, single HeLa cells. The C−D stretching vibrational bands in these amino acids are observed in the 2100−2300 cm−1 spectral region that is devoid of vibrational contributions from other, nondeuterated intracellular constituents. We found that incubation with deuterated amino acids for 8 h in cell culture already led to clearly detectable isotope-related signals in Raman spectra of HeLa cells. As expected, the level of isotope incorporation into proteins increased with incubation time, reaching 55% for deuterated phenylalanine after 28 h. Raman spectral imaging of HeLa cells incubated with deuterium-labeled amino acids showed similar spatial distributions for both isotope-labeled and unlabeled proteins, as evidenced by Raman ratio imaging. The SILAC−Raman methodology presented here combines the strengths of stable isotopic labeling of cells with the nondestructive and quantitative nature of Raman chemical imaging and is likely to become a powerful tool in both cell biology applications and research on tissues or whole organisms
Original languageUndefined
Pages (from-to)9576-9582
Number of pages7
JournalAnalytical chemistry
Volume80
Issue number24
DOIs
Publication statusPublished - 2008

Keywords

  • METIS-254098
  • IR-75806

Cite this

@article{f411ce32e37a41f0989f619812ff6ea1,
title = "Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy",
abstract = "We have combined nonresonant Raman microspectroscopy and spectral imaging with stable isotope labeling by amino acids in cell culture (SILAC) to selectively detect the incorporation of deuterium-labeled phenylalanine, tyrosine, and methionine into proteins in intact, single HeLa cells. The C−D stretching vibrational bands in these amino acids are observed in the 2100−2300 cm−1 spectral region that is devoid of vibrational contributions from other, nondeuterated intracellular constituents. We found that incubation with deuterated amino acids for 8 h in cell culture already led to clearly detectable isotope-related signals in Raman spectra of HeLa cells. As expected, the level of isotope incorporation into proteins increased with incubation time, reaching 55{\%} for deuterated phenylalanine after 28 h. Raman spectral imaging of HeLa cells incubated with deuterium-labeled amino acids showed similar spatial distributions for both isotope-labeled and unlabeled proteins, as evidenced by Raman ratio imaging. The SILAC−Raman methodology presented here combines the strengths of stable isotopic labeling of cells with the nondestructive and quantitative nature of Raman chemical imaging and is likely to become a powerful tool in both cell biology applications and research on tissues or whole organisms",
keywords = "METIS-254098, IR-75806",
author = "{van Manen}, H.J. and Lenferink, {Aufrid T.M.} and Cornelis Otto",
year = "2008",
doi = "10.1021/ac801841y",
language = "Undefined",
volume = "80",
pages = "9576--9582",
journal = "Analytical chemistry",
issn = "0003-2700",
publisher = "American Chemical Society",
number = "24",

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Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy. / van Manen, H.J.; Lenferink, Aufrid T.M.; Otto, Cornelis.

In: Analytical chemistry, Vol. 80, No. 24, 2008, p. 9576-9582.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Noninvasive imaging of protein metabolic labeling in single human cells using stable isotopes and Raman microscopy

AU - van Manen, H.J.

AU - Lenferink, Aufrid T.M.

AU - Otto, Cornelis

PY - 2008

Y1 - 2008

N2 - We have combined nonresonant Raman microspectroscopy and spectral imaging with stable isotope labeling by amino acids in cell culture (SILAC) to selectively detect the incorporation of deuterium-labeled phenylalanine, tyrosine, and methionine into proteins in intact, single HeLa cells. The C−D stretching vibrational bands in these amino acids are observed in the 2100−2300 cm−1 spectral region that is devoid of vibrational contributions from other, nondeuterated intracellular constituents. We found that incubation with deuterated amino acids for 8 h in cell culture already led to clearly detectable isotope-related signals in Raman spectra of HeLa cells. As expected, the level of isotope incorporation into proteins increased with incubation time, reaching 55% for deuterated phenylalanine after 28 h. Raman spectral imaging of HeLa cells incubated with deuterium-labeled amino acids showed similar spatial distributions for both isotope-labeled and unlabeled proteins, as evidenced by Raman ratio imaging. The SILAC−Raman methodology presented here combines the strengths of stable isotopic labeling of cells with the nondestructive and quantitative nature of Raman chemical imaging and is likely to become a powerful tool in both cell biology applications and research on tissues or whole organisms

AB - We have combined nonresonant Raman microspectroscopy and spectral imaging with stable isotope labeling by amino acids in cell culture (SILAC) to selectively detect the incorporation of deuterium-labeled phenylalanine, tyrosine, and methionine into proteins in intact, single HeLa cells. The C−D stretching vibrational bands in these amino acids are observed in the 2100−2300 cm−1 spectral region that is devoid of vibrational contributions from other, nondeuterated intracellular constituents. We found that incubation with deuterated amino acids for 8 h in cell culture already led to clearly detectable isotope-related signals in Raman spectra of HeLa cells. As expected, the level of isotope incorporation into proteins increased with incubation time, reaching 55% for deuterated phenylalanine after 28 h. Raman spectral imaging of HeLa cells incubated with deuterium-labeled amino acids showed similar spatial distributions for both isotope-labeled and unlabeled proteins, as evidenced by Raman ratio imaging. The SILAC−Raman methodology presented here combines the strengths of stable isotopic labeling of cells with the nondestructive and quantitative nature of Raman chemical imaging and is likely to become a powerful tool in both cell biology applications and research on tissues or whole organisms

KW - METIS-254098

KW - IR-75806

U2 - 10.1021/ac801841y

DO - 10.1021/ac801841y

M3 - Article

VL - 80

SP - 9576

EP - 9582

JO - Analytical chemistry

JF - Analytical chemistry

SN - 0003-2700

IS - 24

ER -