MALDI ion trap mass spectrometer with charge detector for large biomolecule detection

Up to now, all commercial matrix-assisted laser desorption/ionization (MALDI) mass spectrometers still can not efficiently analyze very large biomolecules. In this work, we report the development of a novel MALDI ion trap mass spectrometer which can enrich biomolecular ions to enhance the detection sensitivity. A charge detector was installed to measure the large ions directly. With this design, we report the first measurement of IgM with the mass-to-charge ratio (m/z) at 980?000. In addition, quantitative measurements of the number of ions can be obtained. A step function frequency scan was first developed to get a clear signal in the m/z range from 200,000 to 1,000,000.     

Analysis of intact tissue by intermediate-pressure MALDI on a linear ion trap mass spectrometer

The use of an intermediate-pressure matrix-assisted laser desorption/ionization (IP-MALDI) source working at 0.17 Torr on a linear ion trap (LIT) was investigated for the analysis of tissue specimens, in particular, spinal cord sections. MALDI, with 2,5-dihydroxybenzoic acid (DHB) as the matrix, was employed for the detection of phospholipids. The matrix was applied to the tissue using electrospray to avoid analyte migration. The results indicate that analyzing tissue specimens at nontraditional MALDI vacuum pressures is possible. Coupling MALDI to an LIT permits the use of MSn, which is critical for the ability to identify compounds desorbed directly from tissue specimens. Using MSn, ions detected from m/z 600-1000 were characterized as phosphatidlycholines, PC. Specifically, using tandem MS, PC ions could be classified as either [M + H]+ or [M + Na]+ because the fragmentation patterns of protonated and sodiated phosphatidlycholines follow different pathways.     

MALDI of individual biomolecule-containing airborne particles in an ion trap mass spectrometer

Individual airborne biomolecule-containing particles were detected and characterized in near real-time by matrix-assisted laser desorption/ionization (MALDI) with an ion trap mass spectrometer. Biomolecule-containing particles were laboratory-generated and passed through a heated region containing a solution of matrix in equilibrium with the gas phase. Passage into a cooler region created a supersaturation, resulting in rapid deposition of the matrix vapor onto the biomolecule-containing particles, whereupon they were sampled into the inlet of our spectrometer. The coated particles were collimated and individually sized by light-scattering-based time-of-flight. When the sized particle reached the center of the ion trap, it was irradiated with a focused 266-nm laser, and the resulting ions were mass-analyzed. Mass spectra of leucine enkephalin, bradykinin, substance P, and polylysine-containing particles were determined with attomole sensitivity. Structural information of the peptides contained in an individual particle was obtained by tandem mass spectrometry. Analysis of the results yields insights into the aerosol laser ablation ionization process that suggests an optically limited mechanism for ion production that has interesting ramifications on the utility of aerosol-based MALDI as an analytical technique.     

Halo ion trap mass spectrometer

We describe a novel radio frequency ion trap mass analyzer based on toroidal trapping geometry and microfabrication technology. The device, called the halo ion trap, consists of two parallel ceramic plates, the facing surfaces of which are imprinted with sets of concentric ring electrodes. Radii of the imprinted rings range from 5 to 12 mm, and the spacing between the plates is 4 mm. Unlike conventional ion traps, in which hyperbolic metal electrodes establish equipotential boundary conditions, electric fields in the halo ion trap are established by applying different radio frequency potentials to each ring. The potential on each ring can be independently optimized to provide the best trapping field. The halo ion trap features an open structure, allowing easy access for in situ ionization. The toroidal geometry provides a large trapping and analyzing volume, increasing the number of ions that can be stored and reducing the effects of space-charge on mass analysis. Preliminary mass spectra show resolution (m/Deltam) of 60-75 when the trap is operated at 1.9 MHz and 500 Vp-p.     

Handheld rectilinear ion trap mass spectrometer

A shoebox-sized, 10-kg, handheld mass spectrometer, Mini 10, based on a rectilinear ion trap mass analyzer has been designed, built, and characterized. This instrument has evolved from a decade-long experimental and simulation program in mass spectrometer miniaturization. The rectilinear ion trap has a simplified geometry and high trapping capacity, and when used with a miniature and ruggedized pumping system, it allows chemical analysis while the instrument is being carried. Compact electronics, including an air core RF drive coil, were developed to control the instrument and to record mass spectra. The instrument runs on battery power, consuming less than 70 W, similar to a laptop computer. Wired and wireless networking capabilities are implemented. The instrument gives unit resolution and a mass range of over m/z 500. Tandem mass spectrometry capabilities are implemented using collision-induced dissociation, and they are used to provide confirmation of chemical structure during in situ analysis. Continuous monitoring of air and solution samples is demonstrated, and a limit of detection of 50 ppb was obtained for toluene vapor in air and for an aqueous naphthalene solution using membrane sample introduction.     

Miniature cylindrical ion trap mass spectrometer

A miniature cylindrical ion trap mass spectrometer is described, and preliminary data are presented. Functionality and performance of laboratory-scale instruments have been maintained to the extent possible in this battery-operated mass spectrometer. Capabilities include tandem mass spectrometry experiments. Custom-designed electronic components include the RF scanning and amplification system, data acquisition components, and lens power supplies, as well as a custom-software application. Direct leak and membrane introduction inlet systems are used for sample introduction. A mass/charge range of approximately 250 Th with unit mass resolution has been demonstrated.     

Sampling wand for an ion trap mass spectrometer

A new sampling wand concept for ion trap mass spectrometers equipped with discontinuous atmospheric pressure interfaces (DAPI) has been implemented. The ion trap/DAPI combination facilitates the operation of miniature mass spectrometers equipped with ambient ionization sources. However, in the new implementation, instead of transferring ions pneumatically from a distant source, the mass analyzer and DAPI are separated from the main body of the mass spectrometer and installed at the end of a 1.2 m long wand. During ion introduction, ions are captured in the ion trap while the gas in which they are contained passes through the probe and is pumped away. The larger vacuum volume due to the extended wand improves the mass analysis sensitivity. The wand was tested using a modified hand-held ion trap mass spectrometer without additional power or pumping being required. Improved sensitivity was obtained as demonstrated with nano-electrospray ionization (ESI), atmospheric pressure chemical ionization (APCI), and low temperature plasma (LTP) probe analysis of liquid, gaseous, and solid samples, respectively.     

Atmospheric pressure MALDI/ion trap mass spectrometry

A new sample ionization technique, atmospheric pressure matrix-assisted laser desorption/ionization (AP MALDI), was coupled with a commercial ion trap mass spectrometer. This configuration enables the application-specific selection of external atmospheric ionization sources: the electrospray/APCI (commercially available) and AP MALDI (built in-house), which can be readily interchanged within minutes. The detection limit of the novel AP MALDI/ion trap is 10-50 fmol of analyte deposited on the target surface for a four-component mixture of peptides with 800-1700 molecular weight. The possibility of peptide structural analysis by MS/MS and MS3 experiments for AP MALDI-generated ions was demonstrated for the first time.     

Performance evaluation of a hybrid linear ion trap/orbitrap mass spectrometer

Design and performance of a novel hybrid mass spectrometer is described. It couples a linear ion trap mass spectrometer to an orbitrap mass analyzer via an rf-only trapping quadrupole with a curved axis. The latter injects pulsed ion beams into a rapidly changing electric field in the orbitrap wherein they are trapped at high kinetic energies around an inner electrode. Image current detection is subsequently performed after a stable electrostatic field is achieved. Fourier transformation of the acquired transient allows wide mass range detection with high resolving power, mass accuracy, and dynamic range. The entire instrument operates in LC/MS mode (1 spectrum/s) with nominal mass resolving power of 60,000 and uses automatic gain control to provide high-accuracy mass measurements, within 2 ppm using internal standards and within 5 ppm with external calibration. The maximum resolving power exceeds 100,000 (fwhm). Rapid, automated data-dependent capabilities enable real-time acquisition of up to three high-mass accuracy MS/MS spectra per second.     

Multiplexed rectilinear ion trap mass spectrometer for high-throughput analysis

A multichannel mass spectrometer based on the rectilinear ion trap (RIT) analyzer was designed and constructed for simultaneous high-throughput analysis of multiple samples. The instrument features four parallel ion source/mass analyzer/detector channels assembled in a single vacuum chamber and operated using a common set of control electronics, including a single rf amplifier and transformer coil. This multiplexed RIT mass spectrometer employs an array of four millimeter-sized ion traps (x(o) = 5.0 mm and y(o) = 4.0 mm, where x(o) and y(o) are the half-distances in the x and y dimensions, respectively). Mass spectra are acquired from four different samples simultaneously. The available mass/charge range is m/z 15-510 with excellent linearity of the mass calibration (R2 = 0.999 999). The peak width is less than 0.3 mass/charge units at m/z 146, corresponding to a resolution of approximately 500. Simultaneous MS/MS of ions due to four compounds (3-fluoroanisole, 4-fluoroanisole, 2-fluorobenzyl alcohol, 2,6-dimethylcyclohexanone) with the same nominal molecular radical cation but distinctive fragmentation patterns was demonstrated. Isolation and fragmentation efficiencies were approximately 25 and approximately 75%, respectively, measured in the typical case of the molecular radical cation of acetophenone. Preacquisition differential data were obtained by real-time subtraction of the ion signals from two channels of the multiplexed mass spectrometer. The differential experiment presented offers proof of principle of comparative mass spectra in high-throughput screening applications while reducing data storage requirements.     

An interface with a linear quadrupole ion guide for an electrospray-ion trap mass spectrometer system

A new ion sampling interface for an electrospray ionization 3D ion trap mass spectrometer system is described. The interface uses linear rf quadrupoles as ion guides and ion traps to enhance the performance of the 3D trap. Trapping ions in the linear quadrupoles is demonstrated to improve the duty cycle of the system. Dipolar excitation of ions trapped in a linear quadrupole is used to eject unwanted ions. A resolution of ejection of up to 254 is demonstrated for protonated reserpine ions (m/z 609.3). A composite waveform with a notch in frequency space is used to eject a wide range of matrix ions and to isolate trace analyte ions in a linear quadrupole before ions are injected into the 3D trap. This is useful to overcome space charge problems in the 3D trap caused by excess matrix ions. For trace reserpine in a 500-fold molar excess of poly(propylene glycol) (PPG), it is demonstrated that the resolution and sensitivity of the 3D trap can be increased dramatically with ejection of the excess PPG matrix ions. In comparison to ejection of matrix ions in the 3D trap with a similar broad-band waveform, a 5-fold increase in sensitivity with a 7 times shorter acquisition time was achieved.     

High-throughput miniature cylindrical ion trap array mass spectrometer

A fully multiplexed cylindrical ion trap (CIT) array mass spectrometer with four parallel ion source/mass analyzer/detector channels has been built to allow simultaneous high-throughput analysis of multiple samples. A multielement external chemical ionization/electron ionization source was coupled to a parallel array of CITs each of equal size (internal radius 2.5 mm), and the signal was recorded using an array of four miniature (2-mm inner diameter) electron multipliers. Using external electron ionization, the spectra of four separate samples were recorded simultaneously in real time using a four-channel preamplifier system and a data acquisition program written using LabVIEW software. These experiments mark the first demonstration of externally generated ions being successfully trapped in a miniature CIT mass analyzer. The instrument currently provides mass/charge range of approximately m/z 50-500. Average peak width is m/z 0.3, corresponding to a resolution of 1000 at m/z 300. The four-channel mass spectrometer is housed in a single vacuum manifold and operated with a single set of control electronics. The modular design of this instrument allows scale-up to many more channels of analysis for future applications in the areas of industrial process monitoring and combinatorial analysis and in the fields of proteomics and metabolomics.     

MALDI analysis of Bacilli in spore mixtures by applying a quadrupole ion trap time-of-flight tandem mass spectrometer

A novel ion trap time-of-flight hybrid mass spectrometer (qIT-TOF MS) has been applied for peptide sequencing in proteolytic digests generated from spore mixtures of Bacilli. The method of on-probe solubilization and in situ proteolytic digestion of small, acid-soluble spore proteins has been recently developed in our laboratory, and microorganism identification in less than 20 min was accomplished. In this study, tryptic peptides were generated in situ from complex spore mixtures of B. subtilis 168, B. globigii, B. thuringiensis subs. Kurstaki, and B. cereus T, respectively. MALDI analysis of bacterial peptides generated was performed with an average mass resolving power of 6200 and a mass accuracy of up to 10 ppm using a trap-TOF tandem configuration. Precursor ions of interest were usually selected and stored in the quadrupole ion trap with their complete isotope distribution by choosing a window of +/- 2 Da. Sequence-specific information on isolated protonated peptides was gained via tandem MS experiments with an average mass resolving power of 4450 for product ion analysis, and protein and bacterial sources were identified by database searching.     

Planar multipole ion trap/time-of-flight mass spectrometer

We present a novel, hybrid ion trap/time-of-flight mass spectrometer that is based on a planar multipole design. Compared with Paul trap/time-of-flight instruments, this design possesses the principal advantages of higher injection efficiency and more homogeneous extraction fields. We demonstrate the viability of the concept and describe the characterization of a first prototype. Ions can be injected into the trap with little mass discrimination and stored for several minutes. A resolution of over 1300 is achieved in reflectron mode, and the influence of the RF amplitude and pressure on the resolution is analyzed. We suggest several applications in which this new instrument could offer advantages over existing technology.     

Field-emission cold-cathode el source for a microscale ion trap mass spectrometer

A cold-cathode electron impact ionization source based on field emission from an array of diamond-coated silicon whiskers is described. The source is coupled to a microscale ion trap mass spectrometer (r0 = 0.50 mm, z0 = 0.50 mm). An electron beam of 250 nA could be obtained through the 0.45-mm diameter opening in the end cap electrode.     

Ion soft landing using a rectilinear ion trap mass spectrometer

A new ion soft landing instrument has been built for the controlled deposition of mass selected polyatomic ions. The instrument has been operated with an electrospray ionization source; its major components are an electrodynamic ion funnel to reduce ion loss, a 90-degree bent square quadrupole that prevents deposition of fast neutral molecules onto the landing surface, and a novel rectilinear ion trap (RIT) mass analyzer. The ion trap is elongated (inner dimensions: 8 mm x 10 mm x 10 cm). Three methods of mass analysis have been implemented. (i) A conventional mass-selective instability scan with radial resonance ejection can provide a complete mass spectrum. (ii) The RIT can also be operated as a continuous rf/dc mass filter for isolation and subsequent soft landing of ions of the desired m/ z value. (iii) The 90-degree bent square quadrupole can also be used as a continuous rf/dc mass filter. The mass resolution (50% definition) of the RIT in the trapping mode (radial ion ejection) is approximately 550. Ions from various test mixtures have been mass selected and collected on fluorinated self-assembled monolayers on gold substrates, as verified by analysis of the surface rinses. Desorption electrospray ionization (DESI) has been used to confirm intact deposition of [Val (5)]-Angiotensin I on a surface. Nonmass selective currents up to 1.1 nA and mass-selected currents of up to 500 pA have been collected at the landing surface using continuous rf/dc filtering with the RIT. A quantitative analysis of rinsed surfaces showed that the overall solution-to-solution soft landing yields are between 0.2 and 0.4%. Similar experiments were performed with rf/dc isolation of both arginine and lysine from a mixture using the bent square quadrupole in the rf/dc mode. The unconventional continuous mass selection methods maximize soft landing yields, while still allowing the simple acquisition of full mass spectra.     

Preparative linear ion trap mass spectrometer for separation and collection of purified proteins and peptides in arrays using ion soft landing

A preparative mass spectrometer for microarray fabrication is reported. The instrument includes an atmospheric pressure ionization source, a linear ion trap mass analyzer, an ion collection surface positioning system, and a surface loading chamber with independent vacuum pumping. It was designed for the production of protein arrays using the ion soft-landing technique to collect ions on a surface after separation by mass/charge ratio. Small microarrays have been prepared by isolating and soft landing individual protein or peptide ions after electrospray ionization of mixtures. The composition and purity of the separated materials has been confirmed using independent external mass spectrometric analysis of rinse solutions of the collected spots, either by the new method of electrosonic spray ionization MS or by nanospray ionization MS. The ability to retain bioactivity in the mass-selected and collected biomolecules has been demonstrated in particular cases. The reported instrument has also been characterized as an analytical mass spectrometer.     

Electron capture dissociation in a digital ion trap mass spectrometer

Electron capture dissociation was implemented in a digital ion trap without using any magnetic field to focus the electrons. Since rectangular waveforms are employed in the DIT for both trapping and dipole excitation, electrons can be injected into the trap when the electric field is constant. Following deceleration, electrons reach the precursor ion cloud. The fragment ions produced by interactions with the electron beam are subsequently analyzed by resonant ejection. [Glu(1)]-Fibrinopeptide B and substance P were used to evaluate the performance of the current design. Fragmentation efficiency of 5.5% was observed for substance P peptide ions. Additionally, analysis of the monophosphorylated peptide FQ[pS]EEQQQTEDELQDK shows that in the resulting c- and z-type ions, the phosphate group is retained on the phophoserine residue, providing information on which amino acid residue the modification is located.