title: Comparison of vibrational spectroscopy to biochemical and flow cytometry methods for analysis of the basic biochemical composition of mammalian cells.
abstract: We have conducted an extensive comparison of cellular biochemical composition obtained from infrared and Raman spectra of intact cells with measurements using standard extraction and chemical analysis (including NMR), and flow cytometric assay on fixed cells. Measurements were conducted on a rat fibroblast carcinogenesis model consisting of normal and tumorigenic cells assayed as exponentially growing and plateau-phase cultures. Estimates of protein, DNA, RNA, lipids, and glycogen amounts were obtained from a previous publication in which vibrational spectra were fit to a set of basis spectra representing protein, DNA, RNA, lipids, and glycogen. The Raman spectral estimates of absolute cellular composition were quite similar to the independent biochemical and flow cytometric assays. The infrared spectra gave similar results for protein, lipid, and glycogen but underestimated the DNA content while overestimating the RNA level. When ratios of biochemical concentrations in exponential and plateau-phase cultures were examined, the Raman spectroscopic results were the same, within errors, as the independent methods, in all cases. Several changes in relative biochemical composition due to tumorigenic and proliferative status previously reported using vibrational spectroscopy were confirmed by the independent methods. These results demonstrate that vibrational spectroscopy can provide reliable estimates of the biochemical composition of mammalian cells.
authors.affiliation: Los Alamos National Laboratory, Los Alamos, New Mexico 87545, USA. jmourant@lanl.gov
authors.forename: Judith R
authors.initials: JR
authors.lastname: Mourant
authors.forename: Jorge
authors.initials: J
authors.lastname: Dominguez
authors.forename: Susan
authors.initials: S
authors.lastname: Carpenter
authors.forename: Kurt W
authors.initials: KW
authors.lastname: Short
authors.forename: Tamara M
authors.initials: TM
authors.lastname: Powers
authors.forename: Ryszard
authors.initials: R
authors.lastname: Michalczyk
authors.forename: Nagapratima
authors.initials: N
authors.lastname: Kunapareddy
authors.forename: Anabel
authors.initials: A
authors.lastname: Guerra
authors.forename: James P
authors.initials: JP
authors.lastname: Freyer
mesh_terms: D000818:Animals; D001671:Biochemistry / Q000379:methods*; D001685:Biological Factors / Q000032:analysis*; D002452:Cell Count / Q000379:methods*; D002460:Cell Line; D005347:Fibroblasts / Q000737:chemistry*; D005434:Flow Cytometry / Q000379:methods*; D051381:Rats; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013055:Spectrophotometry, Infrared / Q000379:methods*; D013059:Spectrum Analysis, Raman / Q000379:methods*
chemical_list: D001685:Biological Factors