Understanding the influence of post‐excite radius and axial confinement on quantitative proteomic measurements using Fourier transform ion cyclotron resonance mass spectrometry

Abstract

Early studies of Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometry (MS) explored many of the fundamental issues surrounding the potential of the technique to provide quantitative data. Improvements in instrument technology and the analysis of larger molecules in increasingly complex mixtures warrant not only a revisit to some of these earlier studies, but a more comprehensive examination of the influence of various instrument parameters on quantitative (absolute and relative) measurements in proteomics. We present a detailed examination of the role that acquisition time, excite voltage (i.e. excite radius), trapping voltage, and the type of excitation waveform have on the ability of FT-ICR to accurately quantify biological molecules. The use of a stable-isotope-labeled and unlabeled phenyl isocyanate derivatized peptide allows us to ascribe the effects of FT-ICR-MS on quantification, thus eliminating the contribution of ionization differences to ion abundance. To adequately assess the multiple parameters in the large dataset, we develop a multiplicative quality factor that encompasses the total ion abundance, as well as the accuracy and the precision of abundance ratios. This assessment allows facile determination of optimal instrument parameters for quantitative measurements.

Trapping Radial Electric Field Optimization in Compensated FTICR Cells

“…The frequency shifts in Figure 1a and b increase at low postexcitation radii and higher ion populations (i.e., longer external accumulation times), which can be attributed to Columbic interactions also termed space charge effects. In this region, the classical calibration functions are not accurate, and various corrections can be used [13,15,18,[21][22][23]. These results also demonstrate the benefit of using increased ion excitation to achieve higher post-excitation cyclotron radii to reduce columbic interactions and improve MMA, provided the variations of E r /r remain sufficiently small.…”

“…It is convenient to use the Francl's form of the calibration function for characterization of frequency shifts, using the simple Equation 13. Because of extremely high accuracy typical for FTICR mass calibration, this method is very sensitive to small variations of the radial electric field.…”

“…[12][13][14][15][16]. Ideally, if there is no electric field, the cyclotron frequency of an analyte ion, ω c , is readily determined, as shown in Equation (1), where m/z is the mass to charge ratio of the analyte ion and B z is the magnetic field inside of the ion trapping cell:…”

A signal filtering method for improved quantification and noise discrimination in fourier transform ion cyclotron resonance mass spectrometry-based metabolomics data

“…When "drop-out" occurs, the total ion current (TIC) of the signal drops significantly (Ͻ50% of typical values), and the resulting spectrum shows elevated noise levels and is not representative of the sample composition [18]. Some of this noise can be reduced before the measurement, for example by optimization of the ICR detector cell parameters [15,19], or after the measurement, during the preprocessing stages [20]. The overall signal-to-noise ratio (SNR) in a mass spectrum can be improved by acquiring more scans [21]: the magnitude of additive white Gaussian noise (AWGN) increases as n 1⁄2 (where n is the number of scans acquired) and the signal increases as n, and therefore the overall SNR is in principle proportional to n 1⁄2 [22].…”

Trapped-ion cell with improved dc potential harmonicity for FT-ICR MS

“…Although the main components of FT-ICR MS are well established, the growing complexity of modern applications creates a driving force for further optimizations. Fine-tuning acquisition time, excitation power (i.e., excitation radius), trapping voltage, and the type of excitation waveform can significantly improve both mass measurement accuracy and quantitation accuracy of FT-ICR measurements [9,10].…”