Monitoring cell cycle distributions in MCF-7 cells using near-field photothermal microspectroscopy.
Microspectroscopic techniques such as Fourier transform infrared (FTIR) have played an important role in "fingerprinting" the biochemical composition of cellular components. Based on structure and function, complex biomolecules absorb energy in the mid-infrared (lambda = 2-20 microm) yielding characteristic vibrational infrared (IR) spectra. However, optical detection FTIR microspectroscopy may not be suitable for IR-absorbing sample materials. Photothermal microspectroscopy (PTMS) permits the direct measurement of heat generated as a result of sample material absorbing radiation. This approach generates true absorption spectra and is implemented by interfacing a scanning probe microscope and an FTIR spectrometer. Detection is performed using a near-field ultra-miniaturized temperature sensor. Employing PTMS, IR spectra of MCF-7 cells were examined in spectral regions (900-2000 cm(-1)) corresponding to proteins, DNA, RNA, glycoproteins, carbohydrates, lipids, and levels of protein phosphorylation. As a cell passes through the cell cycle, its nuclear material decondenses and condenses and this has led to ambiguity as to whether the intensity of such spectral regions may be associated with the G(1)-, S- or G(2)-phases of the cell cycle. Cultured cells were tracked over a time course known to correspond to marked alterations in cell-cycle distributions, as determined using flow cytometry. Experiments were carried out in the absence or presence of lindane, a pesticide known to induce G(1)-arrest in MCF-7 cells. Significant (P < 0.05) elevations in spectral intensities were associated with exponentially growing cell populations, predominantly in S-phase or G(2)-phase, compared to more quiescent populations predominantly in G(1)-phase. Increases in the absorption band at 970 cm(-1), associated with elevated protein phosphorylation, were observed in vibrational spectra of exponentially growing cell populations compared to those exhibiting a slowing in their growth kinetics. These results seem to suggest that intracellular bulk changes, associated with transit through the cell cycle, can be tracked using PTMS.
Hammiche
Azzedine
A
Department of Physics, Institute of Environmental and Natural Sciences, Lancaster University, Lancaster, United Kingdom.
German
Matthew J
MJ
Hewitt
Rebecca
R
Pollock
Hubert M
HM
Martin
Francis L
FL
D000704:Analysis of Variance; D001703:Biophysics / Q000379:methods*; D002241:Carbohydrates / Q000737:chemistry; D002453:Cell Cycle; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000378:metabolism; D004247:DNA / Q000737:chemistry; D004574:Electromagnetic Fields; D005434:Flow Cytometry; D016193:G1 Phase; D006023:Glycoproteins / Q000737:chemistry; D001556:Hexachlorocyclohexane / Q000737:chemistry; D006801:Humans; D008055:Lipids / Q000737:chemistry; D015591:Microspectrophotometry / Q000379:methods; D011506:Proteins / Q000737:chemistry; D012313:RNA / Q000737:chemistry; D013055:Spectrophotometry, Infrared; D017550:Spectroscopy, Fourier Transform Infrared / Q000295:instrumentation / Q000379:methods*; D013696:Temperature; D013997:Time Factors
D002241:Carbohydrates; D006023:Glycoproteins; D008055:Lipids; D011506:Proteins; D001556:Hexachlorocyclohexane; D012313:RNA; D004247:DNA
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