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Kermit K Murray

Publications and source records attributed to Kermit K Murray.

17 recordsLinked to original sources
PubMed2006

Interfacing capillary gel microfluidic chips with infrared laser desorption mass spectrometry.

We report on the fabrication and performance of a gel microfluidic chip interfaced to laser desorption/ionization (LDI) mass spectrometry with a time-of-flight mass analyzer. The chip was fabricated from poly(methylmethacrylate) with a poly(dimethyl siloxane) cover. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed in the channel of the microfluidic chip. After electrophoresis, the cover was removed and either the PDMS chip or the PMMA cover was mounted in a modified MALDI ion source for analysis. Ions were formed by irradiating the channel with 2.95 microm radiation from a pulsed optical parametric oscillator (OPO), which is coincident with IR absorption by N-H and O-H stretch of the gel components. No matrix was added. The microfluidic chip design allowed a decrease in the volume of material required for analysis over conventional gel slabs, thus enabling improvement in the detection limit to a pmol level, a three orders of magnitude improvement over previous studies in which desorption was achieved from an excised section of a conventional gel.

Comet Assay↗
PubMed2006

Particle formation by infrared laser ablation of glycerol: implications for ion formation.

The quantity and size distribution of micrometer-sized particles ejected from thin films of glycerol were measured using light scattering particle sizing. Thin glycerol films were irradiated at atmospheric pressure with an infrared optical parametric oscillator at wavelengths between 2.95 and 3.1 microm. Particulate material resulting from the ablation was sampled directly into a particle-sizing instrument and particles with diameters greater than 500 nm were detected and sized by light scattering. The fluence threshold for particle formation was between 2000 and 3000 J/m2 for all laser wavelengths. At threshold, fewer than 100 particles/cm3 were detected and this value increased to several thousand particles/cm3 at twice the threshold fluence. The average size of the coarse particles ranged from 900 nm to 1.6 microm at threshold and decreased by 10-20% at twice the threshold fluence. The coarse particle formation observations were compared with ion formation behavior in matrix-assisted laser desorption ionization and interpreted in terms of a photomechanical mechanism for material ablation and ion formation.

Atmospheric Pressure↗
PubMed2006

IR-MALDI-LDI combined with ion mobility orthogonal time-of-flight mass spectrometry.

Most MALDI instrumentation uses UV lasers. We have designed a MALDI-IM-oTOF-MS which employs both a Nd:YAG laser pumped optical parametric oscillator (OPOTEK, lambda = 2.8-3.2 microm at 20 Hz) to perform IR-LDI or IR-MALDI and a Nd:YLF laser (Crystalaser, lambda = 249 nm at 200 Hz) for the UV. Ion mobility (IM) gives a fast separation and analysis of biomolecules from complex mixtures in which ions of similar chemical type fall along well-defined "trend lines". Our data shows that ion mobility allows multiply charged monomers and multimers to be resolved; thus, yielding pure spectra of the singly charged protein ion which are virtually devoid of chemical noise. In addition, we have demonstrated that IR-LDI produced similar results as IR-MALDI for the direct tissue analysis of phospholipids from rat brain.

Animals↗
PubMed2005

Direct coupling of polymer-based microchip electrophoresis to online MALDI-MS using a rotating ball inlet.

We report on the coupling of a polymer-based microfluidic chip to a MALDI-TOF MS using a rotating ball interface. The microfluidic chips were fabricated by micromilling a mold insert into a brass plate, which was then used for replicating polymer microparts via hot embossing. Assembly of the chip was accomplished by thermally annealing a cover slip to the embossed substrate to enclose the channels. The linear separation channel was 50 microm wide, 100 microm deep, and possessed an 8 cm effective length separation channel with a double-T injector (V(inj) = 10 nL). The exit of the separation channel was machined to allow direct contact deposition of effluent onto a specially constructed rotating ball inlet to the mass spectrometer. Matrix addition was accomplished in-line on the surface of the ball. The coupling utilized the ball as the cathode transfer electrode to transport sample into the vacuum for desorption with a 355 nm Nd:YAG laser and analyzed on a TOF mass spectrometer. The ball was cleaned online after every rotation. The ability to couple poly(methylmethacrylate) microchip electrophoresis devices for the separation of peptides and peptide fragments produced from a protein digest with subsequent online MALDI MS detection was demonstrated.

Bombesin↗
PubMed2005

Matrix-assisted laser desorption/ionization mass spectrometry of collected bioaerosol particles.

A method was developed for collection and analysis of bioaerosols by matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry using a modified Andersen N6 bioaerosol collector. The overall goal of the study was to develop methods for obtaining mass spectra with minimal reagents and treatment steps for potential use in remote collection and analysis systems. Test bioaerosol particles were generated from a nebulized E. coli bacterial suspension and collected on MALDI targets placed in an Andersen N6 single-stage aerosol impactor. The bioaerosols were mixed with matrix either by deposition on a bare target with the matrix solution added later, or by deposition on a target pre-coated with matrix. The matrix compounds alpha-cyano-4-hydroxycinnamic acid (CHCA) and sinapic acid (SA) were tested and the SA matrix was found to give the best results in number of peaks, resolution, and signal-to-noise ratio. Deposition of bioaerosol particles onto the matrix pre-coated target did not produce signal in the m/z region above 1000, but the signal could be recovered with the addition of a 1:1 (v/v) acetonitrile/water solvent. Addition of solvent by pipette to the pre-coated targets after particle deposition recovered signal comparable to the dried-droplet sample preparations, whereas solvent sprayed into the impactor recovered fewer peaks. Deposition on pre-coated targets with post-collection solvent addition was superior to deposition on bare target followed by post-collection addition of matrix solution.

Aerosols↗
PubMed2004

Affinity labeling the dopamine transporter ligand binding site.

Photoaffinity labeling is a positive function approach that has been used in an effort to identify the cocaine-binding site on the dopamine transporter (DAT). Radioactive and non-radioactive analogs of cocaine and other dopamine uptake blockers are used to irreversibly label the DAT ligand-binding site and the protein is subjected to chemical or enzymatic treatments that cleave at specific amino acid residues. Analysis of cleavage products from radioactively photolabeled DAT using epitope-specific immunoprecipitation, gel electrophoresis, and autoradiography has identified the site of origin in the primary sequence of labeled fragments as small as 4 kDa. More precise localization of the site of labeling is done by subjecting photolabeled DAT to parallel or serial digestion with multiple cleavage methods, followed by analysis of radiolabeled peptides by reverse-phase HPLC. Fragment retention times are compared to calculated retention times of predicted digest peptides and to chemically or photochemically labeled synthetic peptides. The presence of authentic DAT sequence in HPLC fractions of digests from DAT labeled with non-radioactive ligands is further supported by MALDI and nanoelectrospray mass spectrometry. Using these methods we have identified two distinct regions of DAT that interact with multiple structurally related and diverse irreversible ligands, suggesting that these regions may be involved in the formation of ligand binding sites.

Affinity Labels↗
PubMed2004

Online CE-MALDI-TOF MS using a rotating ball interface.

We report on the construction and performance of a rotating ball interface for online coupling of capillary electrophoresis (CE) to matrix-assisted laser desorption ionization (MALDI) mass spectrometry with a time-of-flight (TOF) mass analyzer. The interface is based on a rotating stainless steel ball that transports samples from atmospheric pressure to the high vacuum of the mass spectrometer for desorption and ionization. The sample is deposited directly from a 50-microm-i.d. separation capillary onto the 19-mm ball that is rotating at 0.03 to 0.3 rpm. The sample is mixed online with matrix flowing from a separate 50-microm-i.d. capillary. The sample deposit dries before it is rotated past a polymer gasket and into the laser ionization region. Cleaning of the interface is accomplished using solvent-saturated felt, which cleans the ball surface after it rotates out of the ionization chamber. On-line CE-MALDI is demonstrated, and the performance is evaluated with the analysis of a mixture of three peptides: [Lsy8] vasopressin, substance P, and neurotensin. The rotating ball interface to MALDI-TOF MS demonstrated mass detection limit in the high femtomole range. The interface has negligible memory effect and shows no significant electrophoretic peak broadening when operated under optimized conditions.

Electrochemistry↗
PubMed2004

Direct from polyacrylamide gel infrared laser desorption/ionization.

The direct combination of gel electrophoresis and infrared laser desorption/ionization time-of-flight mass spectrometry has been demonstrated. We present results for infrared laser desorption and ionization mass spectrometry of peptides and proteins directly from a polyacrylamide gel without the addition of a matrix. Analyte molecules up to 6 kDa were ionized directly from a vacuum-dried sodium dodecyl sulfate-polyacrylamide gel after electrophoretic separation. Mass spectra were obtained at the wavelength of 2.94 microm, which is consistent with IR absorption by N-H and O-H stretch vibrations of water and other constituents of the gel. A 5-nmol quantity of peptide or protein was loaded per gel slot, although it was possible to obtain mass spectra from a small fraction of the gel spot. This technique shows promise for the direct identification of both parent and fragment masses of proteins contained in polyacrylamide gels.

Animals↗
PubMed2004

On-line laser desorption/ionization mass spectrometry of matrix-coated aerosols.

Matrix-assisted laser desorption/ionization (MALDI) was used for the on-line analysis of single particles. An aerosol was generated at atmospheric pressure and particles were introduced into a time-of-flight (TOF) mass spectrometer through a single-stage differentially pumped capillary inlet. Prior to entering the mass spectrometer, a matrix was added to the particles using a heated saturator and condenser. A liquid matrix, 3-nitrobenzyl alcohol (NBA), and a solid matrix, picolinic acid (PA), were used. Particles were ablated with a 351 nm excimer laser and the resulting ions were mass-separated in a two-stage reflectron TOF mass spectrometer. Aerosol particles containing the biomolecules erythromycin and gramicidin S were analyzed with and without the matrix addition step. The addition of NBA to the particles resulted in mass spectra that contained an intact molecular ion mass peak. In contrast, PA-coated particles did not yield molecular ion peaks from matrix-coated particles.

Aerosols↗
PubMed2004

On-line single droplet deposition for MALDI mass spectrometry.

A single droplet generator was coupled with a rotating ball inlet matrix-assisted laser desorption/ionization (MALDI) time of flight (TOF) mass spectrometer. Single droplets with 100 picoliter volume were ejected by a piezoelectric-actuated droplet generator and deposited onto a matrix-coated rotating stainless steel ball at atmospheric pressure. The single droplet deposit was transported to the vacuum side of the instrument where ionization was accomplished using a UV pulsed laser. Using this on-line interface, it was possible to obtain protonated molecule signal from as little as 10 fmol analyte.

Angiotensin I↗
PubMed2004

Characterization of infrared matrix-assisted laser desorption ionization samples by Fourier transform infrared attenuated total reflection spectroscopy.

Fourier transform infrared attenuated total reflection (FT-IR ATR) spectroscopy was used to characterize thin films of succinic acid, a matrix compound commonly used with infrared matrix-assisted laser desorption ionization (IR-MALDI) mass spectrometry. IR spectra of succinic acid thin films deposited alone and in combination with the analyte biomolecules insulin and cytochrome c were obtained by FT-IR ATR spectroscopy. Spectra of analyte and matrix alone were similar to those obtained previously from KBr pellets, Nujol mull, or thin-film absorption, although the ATR spectra have significantly lower background interferences. Thin films deposited from mixtures of water and methanol have additional peaks compared to films deposited from a methanol solution. These additional peaks are attributed to carboxylate groups stabilized by residual water molecules. No evidence was found to suggest that residual water absorption contributes to absorption at wavelengths typically used for IR-MALDI. Absorption of energy by analyte vibrational modes with rapid energy transfer to the matrix is suggested as a contributor to desorption and ionization consistent with the FT-IR ATR results.

Journal Article↗
PubMed2003

Two-laser infrared and ultraviolet matrix-assisted laser desorption/ionization.

Matrix-assisted laser desorption/ionization (MALDI) was performed using two pulsed lasers with wavelengths in the IR and UV regions. A 10.6 micro m pulsed CO(2) laser was used to irradiate a MALDI target, followed after an adjustable delay by a 337 nm pulsed nitrogen laser. The sample consisted of a 2,5-dihydroxybenzoic acid matrix and bovine insulin guest molecule. The pulse energy for both of the lasers was adjusted so that the ion of interest, either the matrix or guest ion, was not produced by either of the lasers alone. The delay time for maximum ion yield occurs at 1 micro s for matrix and guest ions and the signal decayed to zero in approximately 400 micro s. A mechanism is presented for enhanced UV MALDI ion yield following the IR laser pulse based on transient heating.

Animals↗
PubMed2002

Matrix addition by condensation for matrix-assisted laser desorption/ionization of collected aerosol particles.

Condensation of an ultraviolet absorbing liquid matrix onto aerosol particles was used to enhance the ionization efficiency of large molecules. Laboratory-generated particles were coated with matrix, deposited on a sample target, and analyzed by laser desorption mass spectrometry with no other matrix addition. The aerosol was generated in a Collison nebulizer, and the particles were dried in a diffusion dryer before entering a heated region saturated with the liquid matrix 3-nitrobenzyl alcohol (NBA) and then entering a cooled condensation region. Matrix-coated particles were collected on a sample target and analyzed using a 337-nm laser and a time-of-flight mass spectrometer. Particles containing the peptides gramicidin S and gramicidin D were analyzed both with and without the matrix addition step. Condensation addition of matrix increased the biomolecule ion signal and resulted in mass spectra with less fragmentation and low-mass ion interference.

Aerosols↗
PubMed2002

Infrared laser desorption/ionization on silicon.

Laser desorption/ionization from a single-crystal silicon surface was performed using a laser operating in the 3-microm region of the mid-infrared. Analyte molecules up to 6 kDa were ionized with no added matrix. As with ultraviolet desorption/ionization from porous silicon (DIOS), IR laser desorption from silicon does not produce matrix ions that can interfere with analysis of low-mass analytes. However, in contrast to UV DIOS, silicon porosity or roughness is not required for ionization using an IR laser. Mass spectra were obtained in the wavelength range between 2.8 and 3.5 microm, which is consistent with energy absorption by a hydrogen-bonded OH group. A mechanism based on desorption of adsorbed solvent molecules is postulated.

Animals↗
PubMedDate not supplied

Laser capture microdissection MALDI for direct analysis of archival tissue.

MALDI mass spectra were obtained from cancer cells isolated by laser capture microdissection (LCM) of archived tissue. Frozen human lung tissue from adenocarcenoma and squamous cell carcenoma cases were cut into 5 to 15 microm thick sections, stained with hematoxylin and dehydrated. Cancer cells were isolated by LCM, mixed with matrix solution, and deposited on a MALDI target for mass spectrometric analysis. For comparison with LCM isolated cells, tissue sections were placed directly on the MALDI target without microdissection. Tissue sections frozen in optimal cutting temperature (OCT) solution and cut into 8 microm thick sections gave the best performance with direct MALDI analysis. Between 15 and 20 peaks were observed in the mass region between 1,000 and 4,000 Da, and roughly half of these peaks were common to either squamous cells or adenocarcenoma. Additional peaks were observed in the non-LCM mass spectra and these may result from biomolecules in the healthy tissue. When compared to fresh tissue, both LCM and non-LCM archived tissue produced fewer peaks, possibly due to degradation of the biomolecules in the archived tissue.

Adenocarcinoma↗