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We report on the quantitative proteomic analysis of single mammalian cells. Fluorescence-activated cell sorting was employed to deposit cells into a newly developed nano-droplet sample processing chip, after which samples were analyzed by ultrasensitive nanoLC-MS. An average of circa 670 protein groups were confidently identified from single HeLa cells, which is a far greater level of proteome coverage for single cells than has been previously reported. We demonstrate that the single-cell proteomics platform can be used to differentiate cell types from enzyme-dissociated human lung primary cells and identify specific protein markers for epithelial and mesenchymal cells.

RATIONALE Electrospray ionization (ESI) results in the formation of multiply charged cluster ions which then fragment via two competing processes: fission (charge separation) via Coulomb repulsion, and evaporation of one or more neutrals from the fragmenting ion, as described by Rayleigh's liquid drop model. Unimolecular and collision-induced dissociation (CID) of doubly charged lithium acetate clusters, (CH3 COOLi)n Li2 (2+) , demonstrated that Coulomb fission via charge separation is the dominant dissociation process with no contribution from the neutral evaporation processes for all such ions from the critical limit to larger cluster ions, although the latter process was normally observed in most earlier studies of multiply charged clusters. METHODS Doubly and higher charged lithium acetate cluster ions were generated by ESI. Unimolecular and CID of mass selected doubly charged cluster ions above the critical limit were recorded at high resolution using an LTQ Velos Orbitrap mass spectrometer. RESULTS Unimolecular and collision-induced dissociation of doubly charged lithium acetate clusters proceed via symmetric fission processes and the intensity of the fragment ions decreases as the cluster size increases. Dissociations via neutral evaporation of a monomer or larger clusters are not observed. CONCLUSIONS Doubly charged clusters of lithium acetate above the critical limit dissociate via symmetrical fission processes and not neutral evaporation. These results are clearly in disagreement with the Rayleigh liquid drop model that has been used previously to predict the critical size and explain the fragmentation behavior of multiply charged clusters. Copyright © 2016 John Wiley & Sons, Ltd.

Two unexpected singly charged ions at m/z 1103 and 944 have been observed in mass spectra obtained from electrospray ionization mass spectrometric analysis of liquid chromatography effluents with mobile phases containing trifluoroacetic acid (TFA) that severely interfered with sample analysis. Accurate mass measurement and tandem mass spectrometry studies revealed that these two ions are composed of three components; clusters of trifluoroacetic acid, clusters of mass 159 and iron. Formation of these ions is inhibited by removing TFA from the mobile phases and using formic acid in its place, replacing the stainless steel union with a titanium union or by adding a small blank fused-silica capillary column between the chromatography column and the electrospray tip via a stainless steel union without any adverse effects to chromatographic separation, peak broadening or peptide identifications.

Aim: The aim of the present study was to evaluate the effect of occlusal splint therapy by determining the cross-sectional dimension of masseter muscle using ultrasound in patients with temporomandibular joint disc displacement with reduction. Methods: Twenty-seven patients aged between 20 and 40 years were included in the study. A detailed history was obtained, and a comprehensive clinical examination was carried out. Subjective assessment with structured pro forma was performed pre-occlusal and post-occlusal splint therapy. The objective measurement of the bilateral masseter muscle thickness was recorded using real-time ultrasonography before and after splint therapy. Results: The mean ultrasound thickness of the masseter muscle in the pre-clenching state before splint therapy was 9.45 mm (SD 1.39), and the post-clenching state was 13.15 mm (SD 2.23). After splint therapy, the mean thickness in the pre-clenching state was 9.14 mm (SD 1.31), and the post-clenching state was 12.78 mm (SD 2.23; P

Unimolecular (metastable) and collision-induced dissociation of 1,3, 5-trinitrobenzene molecular ion was studied using linked scans and mass-analyzed ion kinetic energy spectrometry on a hybrid instrument of EBEqQ geometry. An unusual ortho effect leading to the loss of OH radical from the parent molecular ion is observed as a unimolecular dissociation process only in the first-field free region between the ion source and the electric sector, although corresponding dissociation in the ion source is of negligible abundance (

This review presents a brief historical introduction to the development of tandem mass spectrometry and its principal applications. It is placed in the context of the general principles underlying mass spectrometry, particularly the relationships between internal energy and fragmentation kinetics. The center-of-mass framework is presented as a convenient means of applying conservation of momentum to the energy transfer problem in tandem mass spectrometry as a means of deducing energy transfer in the collisional activation step and kinetic energy release as activated ions dissociate into fragment ions and neutrals. The principles of molecular beam methods are summarized and illustrative examples are given for which definitive information on reaction dynamics is available. The importance of scattering-very little appreciated in early discussions of tandem mass spectrometry-is shown to be the natural consequence of impulsive collisions, which appears to be a general mechanism for energy exchange in collisional activation. It is shown that the average energy transferred in single collisions is much less than the theoretical maximum given by the center-of-mass collision energy and the Massey criterion is presented as a simplistic rationale for understanding the essentially exponential decline in the energy transfer function above and below the relative velocity at which the probability for energy transfer is maximized. The issues of energy transfer in collisions of large molecular ions with low-mass neutrals are reviewed and a general description of energy transfer in multiple collisions is presented. It is shown that the center-of-mass and Massey criterion limitations are pragmatically overcome by multiple collision activation in ion traps. Surface-induced dissociation is presented as a viable alternative to multiple collision activation which is especially attractive for activation of large molecular ions. Finally, a few of the emerging dynamics principles governing energy transfer and dissociation of peptides are summarized. Copyright 2000 John Wiley & Sons, Ltd.

Collision-induced dissociation of nitromethane molecular ions to NO+ and NO has been studied at 19 eV center-of-mass collision energy using a crossed-beam tandem mass spectrometer. Center-of-mass velocity contour maps for the two dissociation processes are distinctly different; NO has its intensity maximum at 7 degrees with energy transfer endothermicities of 1.8 eV while NO+ has intensity maxima at zero and 8 degrees with energy transfer endothermicities of 0.3 and 5.5 eV, respectively. The observation of two distinct peaks in the NO+ contour map clearly demonstrates that nitromethane ions excited to 5.5 eV internal energy do not dissociate from the ground state potential surface. Dissociation is faster than intramolecular relaxation, in contradiction to the fundamental postulate of statistical theories (RRKM, QET) for unimolecular dissociation.