Rapid identification of differentiation markers from whole epithelial cells by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry and statistical analysis.
Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) was applied to identify markers for cellular differentiation. The differentiation of a human colon epithelial carcinoma T84 cell line was monitored over a period of 28 days by transepithelial electrical resistance (TER) measurements, alkaline phosphatase (AP) assay, and MALDI-TOF mass spectral fingerprints combined with statistical analysis. MALDI-MS generated specific mass spectral fingerprints characteristic of cell differentiation. Twenty-two ions were selected as diagnostic signals of fully differentiated T84 cells. Ten protein ion signals, detected by MALDI-MS and validated by statistical analysis, were proposed as T84 cell differentiation markers. Among these signals, ubiquitin was identified as a T84 cell differentiation marker by nanospray liquid chromatography/tandem mass spectrometry (nanoLC/MS/MS). Moreover, depending on the concentration of the cells seeded on the growth support, it was possible to predict the timing of the exponential phase and of cellular differentiation by MALDI-MS-derived marker ions. MALDI-TOFMS was compared to other methods for the determination of cellular differentiation: TER measurements are rapid but yield limited information as to the cellular differentiation state. AP assays are more specific for the differentiation state but take more time. By contrast, MALDI-MS has been found to be a fast, sensitive and precise method for cell differentiation assessment and provides the opportunity for multiplexing and high throughput. Moreover, the consumable costs per assay are very low.
Marvin-Guy
Laure F
LF
Bioanalytical Sciences Department, Nestlé Research Center, Lausanne, Switzerland. laure.marvin-guy@rdls.nestle.com
Duncan
Peter
P
Wagnière
Sandrine
S
Antille
Nicolas
N
Porta
Nadine
N
Affolter
Michael
M
Kussmann
Martin
M
D000469:Alkaline Phosphatase / Q000032:analysis / Q000378:metabolism; D015415:Biomarkers / Q000032:analysis / Q000378:metabolism*; D002454:Cell Differentiation / Q000502:physiology*; D045744:Cell Line, Tumor; D002851:Chromatography, High Pressure Liquid; D003110:Colonic Neoplasms; D017097:Electric Impedance; D020895:Enterocytes / Q000737:chemistry / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D006801:Humans; D036103:Nanotechnology; D010449:Peptide Mapping / Q000706:statistics & numerical data; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry; D025801:Ubiquitin / Q000032:analysis / Q000378:metabolism
D015415:Biomarkers; D025801:Ubiquitin; D000469:Alkaline Phosphatase