Investigating the quantitative nature of MALDI-TOF MS.
MALDI-TOF MS has been applied by several groups to relative quantitative measurements. At the same time, the non-quantitative character of this method has been widely reported. We conducted experiments to test the reliability of this technique for quantitation using the statistical method of inverse confidence limit calculation for the first time in this context. The relationship between relative intensities of known amounts of standard peptides and their concentration ratios was investigated. We found that the concentration ratios determined by relative intensity measurements were highly inaccurate and strongly influenced by the molecular milieu of the sample analyzed. Thus, we emphasize the necessity of using the sample itself for calibration. We also performed experiments using an isotope-labeled derivative of the analyte as an internal standard for calibration line generation. As expected, the use of such standard led to a dramatic increase in precision and a less pronounced improvement in accuracy. We recommend performing a similar statistical analysis as a demonstration of reliability for every system where MALDI-TOF MS is used for quantitative measurements.
Szájli
Emília
E
Proteomics Research Group, Biological Research Center, Hungarian Academy of Sciences, Temesvári körút 62, H-6726 Szeged, Hungary.
Fehér
Tamás
T
Medzihradszky
Katalin F
KF
D000595:Amino Acid Sequence; D000818:Animals; D002138:Calibration; D002417:Cattle; D045304:Cytochromes c / Q000032:analysis / Q000737:chemistry; D004926:Escherichia coli; D007553:Isotope Labeling; D008969:Molecular Sequence Data; D010455:Peptides / Q000032:analysis / Q000737:chemistry; D012015:Reference Standards; D012044:Regression Analysis; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D000509:alpha-Fetoproteins / Q000032:analysis
D010455:Peptides; D000509:alpha-Fetoproteins; D045304:Cytochromes c