Chapter 4: NEMS Mass Spectrometry and Inertial Imaging
4.1 Overview and Motivation
In this section, we discuss some of the more common methods of mass spectrometry including time-of-flight measurement, magnetic sector mass analyzer, and a quadrupole [15], and the advantages and disadvantages of each.
One of the conceptually simplest methods for mass spectrometry is a time- of-flight measurement (Figure 4-1). The analyte is injected into a vacuum chamber from the fluid phase and ionized, typically using electrospray ionization [16, 17]. It then travels through a discrete acceleration region which results in different particles having different final velocities depending on the field and the charge-to-mass ratios of the various analytes. The separate species then travel at different velocities in the drift tube and arrive at the detector at different times [18, 19]. Given the duration of the particle in the tube and the length of the tube, the mass-to-charge ratio can be
deduced.
Figure 4-1 Time-of-flight measurement to perform mass spectrometry. The particles are charged and start on the left of the figure where they are accelerated through an electric field created by a DC voltage. The particle travels to the right and the duration of the particle in the tube is used to calculate the mass-to-charge ratio. Figure adapted from Glish, G.L. and R.W. Vachet, The basics of mass spectrometry in the twenty-first century. Nat Rev Drug Discov, 2003. 2(2): p. 140-150. [15].
The magnetic sector mass analyzer provides another method of performing mass spectrometry (Figure 4-2). In this case the sample is also typically introduced by electrospray ionization and accelerated. In this case, the particle is accelerated in an electric field and a magnetic field is applied perpendicular to both the plane of analyte motion and the direction of the electric field. The Lorentz force, induced by analyte motion in the applied magnetic field, deflects the analyte according to its mass-to-charge ratio. The trajectories of the different analytes have different radii, depending upon the Lorentz force they experience. With a detector at a fixed position (Fig. 4.2), the magnetic field is swept to permit detection of species with different mass-to-charge ratios.
Figure 4-2 Magnetic sector mass analyzer. The particles are charged and start at the top of the figure where they are accelerated through an electric field created by a DC voltage. Due to the applied magnetic field, particles with different masses travel at a different velocity and thus have a different turn radius. By varying the strength of the magnetic field, samples of different mass-to-charge ratios are detected. Figure adapted from Glish, G.L. and R.W.
Vachet, The basics of mass spectrometry in the twenty-first century.
Nat Rev Drug Discov, 2003. 2(2): p. 140-150. [15].
Typically, both the time-of-flight and the magnetic sector mass analyzers are large instruments, with dimensions usually between 0.5 𝑚 and 2 𝑚. This permits sufficient trajectory lengths to permit separation of species with sufficiently high resolution. To reduce the size of mass spectrometers, quadrupole spectrometry has been developed (Figure 4-3). A quadrupole consists of four parallel cylindrical metal electrodes. Each opposing electrode pair is connected electrically, and a radio frequency voltage with a DC offset is applied between each pair of rods; the rods being driven in antiphase. Ions travel down the quadrupole between the rods. Only
ions of a certain mass-to-charge ratio reach the detector for a given ratio of voltages; other ions have unstable trajectories and collide with the rods or are ejected from the interstitial space between them.
Figure 4-3 Diagram of quadrupole used in traditional mass spectrometry. The quadrupole is a set of four metal rods in parallel.
In this figure, the fourth and the closest rod is not drawn so the ions are more easily seen. Opposing rods are connected electrically and the two pairs of rods have an RF and DC voltage offset. By tuning the frequency and amplitude of the voltage on the rods, only charged particles of a certain mass-to-charge ratio arrive at the detector. Charged particles of other mass-to-charge ratios collide with the rod or spin out of orbit and thus are eliminated. Figure adapted from Glish, G.L. and R.W. Vachet, The basics of mass spectrometry in the twenty-first century. Nat Rev Drug Discov, 2003. 2(2): p. 140-150. [15].
These three mass spectrometry methods can provide excellent mass resolution; the best provide the ability to measure down to a small fraction of the
proton mass (1.007 𝑎𝑚𝑢 ~ 1.007 𝐷𝑎). However, these methods typically lack sufficient dynamic range to measure large samples, exceeding several hundred kDa.
Examples of species in this range are large protein complexes (membrane proteins, antibody isoforms, organelles, and viruses), which can range from . 05 − 100′𝑠 of MDa. Existing forms of mass spectrometry also require charging (ionizing) the sample; for non-covalently bonded complexes this can compromise the integrity of the analytes and potentially induce their fragmentation.
NEMS-MS offers unique opportunities to overcome these limitations.
NEMS-MS offers the ability of single molecule detection, and the analytes need not be ionized for analysis and detection. While there exists single ion detectors, the convolution of measuring the mass-to-charge ratio instead of a direct mass measurement makes it challenging to know accurately the mass of the single molecule for very large species that are in heterogeneous mixtures. NEMS-MS offers a large dynamic range (perhaps ~7 orders of magnitude or more), offering the ability to measure very large, as well as small, molecules. Current implementations of NEMS-MS permit measurement of molecules ranging in mass from 104 𝐷𝑎 to 109 𝐷𝑎. In the next section, we discuss the operating principal of how NEMS are employed for mass spectrometry.