Quantification of singly charged biomolecules by electrospray ionization fourier transform ion cyclotron resonance mass spectrometry utilizing an internal standard

✍ Scribed by Eric F. Gordon; David C. Muddiman

• Publisher: John Wiley and Sons
• Year: 1999
• Tongue: English
• Weight: 132 KB
• Volume: 13
• Category: Article
• ISSN: 0951-4198
• DOI: 10.1002/(sici)1097-0231(19990215)13:3<164::aid-rcm474>3.0.co;2-l

✦ Synopsis

The quantification of Cyclosporin A (CsA) and related biological compounds by various mass spectrometric techniques is well established. It is precisely this fact that makes the use of CsA particularly appealing as a model system in evaluating the quantitative properties of electrospray ionization coupled to Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS). Utilizing the internal standard methodology, we have generated a linear calibration curve ranging from 5-250 ng/mL CsA described by the equation y = 0.00503x-0.00958, with a %RSD slope of 0.74% and a correlation coefficient (r 0 ) equal to 0.999 2 (N = 30). The internal standard (200 ng/mL Cyclosporin G) was found to be necessary to establish the reported dynamic range, two orders of magnitude based on an experimentally determined detection limit of 2.4 ng/mL CsA, and also significantly improved the overall precision of the method. The data represented in the standard curve were collected as 1.05 s single acquisitions (512 K data points at an ADC rate of 500 kHz) and unapodized prior to the Fourier transformation of the time-domain data. We report that full Hanning apodization has a statistically significant negative effect (F-test at 95% confidence level) on the regression statistics of the curve (i.e. increasing the %RSD slope to 1.33%). Finally, we report that acceptable quantitative properties of an FTICR-MS experiment can be realized as soon as the detection of a second isotopic beat for the targeted species is obtained. This should allow for the practical and effective coupling of on-line separation techniques for quantitative analysis using ESI-FTICR-MS.