Optically-pumped saturable absorber for fast switching between continuous-wave and passively mode-locked regimes of a Nd:YVO4 laser

We report on the fast (~50 μs) remote-controlled switching between continuous-wave (cw), cw mode-locked (ML) and Q-switched ML modes of operation of a Nd:YVO4 laser using an optically-pumped saturable absorber (SA). Pulses as short as 40 ps with an average output power of 0.5 W are obtained in cw ML regime

1.6 W continuous-wave Raman laser using low-loss synthetic diamond

Low-birefringence (Δn<2x10−6), low-loss (absorption coefficient <0.006cm−1 at 1064nm), single-crystal, synthetic diamond has been exploited in a CW Raman laser. The diamond Raman laser was intracavity pumped within a Nd:YVO4 laser. At the Raman laser wavelength of 1240nm, CW output powers of 1.6W and a slope efficiency with respect to the absorbed diode-laser pump power (at 808nm) of ~18% were measured. In quasi-CW operation, maximum on-time output powers of 2.8W (slope efficiency ~24%) were observed, resulting in an absorbed diode-laser pump power to the Raman laser output power conversion efficiency of 13%

Large radius of curvature micro-lenses on single crystal diamond for application in monolithic diamond Raman lasers

The design and fabrication of large radii of curvature micro-lenses in single crystal chemical vapour deposition diamond is described. An optimised photoresist reflow process and low selectivity inductively coupled plasma etching are used to actualize a uniform array of micro-lenses with radii of curvature of 13 mm or more and a high quality surface of a root-mean-square roughness of 0.18 nm. The processes developed have the potential to achieve diamond micro-lenses with an even larger radius of curvature. These new diamond micro-lenses enable the pulse energy scalable monolithic diamond Raman laser where a large radius of curvature of the micro-lenses is critical

Database Search Strategies for Proteomic Data Sets Generated by Electron Capture Dissociation Mass Spectrometry

Large data sets of electron capture dissociation (ECD) mass spectra from proteomic experiments are rich in information; however, extracting that information in an optimal manner is not straightforward. Protein database search engines currently available are designed for low resolution CID data, from which Fourier transform ion cyclotron resonance (FT-ICR) ECD data differs significantly. ECD mass spectra contain both z-prime and z-dot fragment ions (and c-prime and c-dot); ECD mass spectra contain abundant peaks derived from neutral losses from charge-reduced precursor ions; FT-ICR ECD spectra are acquired with a larger precursor m/z isolation window than their low-resolution CID counterparts. Here, we consider three distinct stages of postacquisition analysis: (1) processing of ECD mass spectra prior to the database search; (2) the database search step itself and (3) postsearch processing of results. We demonstrate that each of these steps has an effect on the number of peptides identified, with the postsearch processing of results having the largest effect. We compare two commonly used search engines: Mascot and OMSSA. Using an ECD data set of modest size (3341 mass spectra) from a complex sample (mouse whole cell lysate), we demonstrate that search results can be improved from 630 identifications (19% identification success rate) to 1643 identifications (49% identification success rate). We focus in particular on improving identification rates for doubly charged precursors, which are typically low for ECD fragmentation. We compare our presearch processing algorithm with a similar algorithm recently developed for electron transfer dissociation (ETD) data

Liquid Chromatography Electron Capture Dissociation Tandem Mass Spectrometry (LC-ECD-MS/MS) versus Liquid Chromatography Collision-induced Dissociation Tandem Mass Spectrometry (LC-CID-MS/MS) for the Identification of Proteins

Electron capture dissociation (ECD) offers many advantages over the more traditional fragmentation techniques for the analysis of peptides and proteins, although the question remains: How suitable is ECD for incorporation within proteomic strategies for the identification of proteins? Here, we compare LC-ECD-MS/MS and LC-CID-MS/MS as techniques for the identification of proteins.Experiments were performed on a hybrid linear ion trap–Fourier transform ion cyclotron resonance mass spectrometer. Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin,lysozyme, cytochrome c, alcohol dehydrogenase, and β-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides. The results show that although protein coverage was lower for LC-ECDMS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags,providing greater confidence in protein assignment

Targeted online liquid chromatography electron capture dissociation mass spectrometry for the localization of sites of in vivo phosphorylation in human Sprouty2

We demonstrate a strategy employing collision-induced dissociation for phosphopeptide discovery, followed by targeted electron capture dissociation (ECD) for site localization. The high mass accuracy and low background noise of the ECD mass spectra allow facile sequencing of coeluting isobaric phosphopeptides, with up to two isobaric phosphopeptides sequenced from a single mass spectrum. In contrast to the previously described neutral loss of dependent ECD method, targeted ECD allows analysis of both phosphotyrosine peptides and lower abundance phosphopeptides. The approach was applied to phosphorylation analysis of human Sprouty2, a regulator of receptor tyrosine kinase signaling. Fifteen sites of phosphorylation were identified, 11 of which are novel

Energy scaling of yellow emission from monolithic diamond Raman lasers

Recent advances in the growth of low loss single crystal diamond [1] coupled with its high Raman gain and high thermal conductivity have led to the material becoming an established Raman laser material. With applications often having to adapt to available laser wavelengths, Raman lasers can be used as a simple means to shift ubiquitous commercial laser sources to the hard to reach but application rich spectral regions. The authors recently demonstrated a compact, robust monolithic diamond Raman laser shifting 20μJ nanosecond pulses from a Q-switched 532nm laser to Raman wavelengths of 573nm, 620nm and 676nm with a conversion efficiency of 84%. This work investigates the energy scalability of such a system

Activated Ion Electron Capture Dissociation (AI ECD) of proteins: synchronization of infrared and electron irradiation with ion magnetron motion.

Here, we show that to perform activated ion electron capture dissociation (AI-ECD) in a Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometer equipped with a CO(2) laser, it is necessary to synchronize both infrared irradiation and electron capture dissociation with ion magnetron motion. This requirement is essential for instruments in which the infrared laser is angled off-axis, such as the Thermo Finnigan LTQ FT. Generally, the electron irradiation time required for proteins is much shorter (ms) than that required for peptides (tens of ms), and the modulation of ECD, AI ECD, and infrared multiphoton dissociation (IRMPD) with ion magnetron motion is more pronounced. We have optimized AI ECD for ubiquitin, cytochrome c, and myoglobin; however the results can be extended to other proteins. We demonstrate that pre-ECD and post-ECD activation are physically different and display different kinetics. We also demonstrate how, by use of appropriate AI ECD time sequences and normalization, the kinetics of protein gas-phase refolding can be deconvoluted from the diffusion of the ion cloud and measured on the time scale longer than the period of ion magnetron motion

Pervasive protein thermal stability variation during the cell cycle

Quantitative mass spectrometry has established proteome-wide regulation of protein abundance and post-translational modifications in various biological processes. Here, we used quantitative mass spectrometry to systematically analyze the thermal stability and solubility of proteins on a proteome-wide scale during the eukaryotic cell cycle. We demonstrate pervasive variation of these biophysical parameters with most changes occurring in mitosis and G1. Various cellular pathways and components vary in thermal stability, such as cell-cycle factors, polymerases, and chromatin remodelers. We demonstrate that protein thermal stability serves as a proxy for enzyme activity, DNA binding, and complex formation in situ. Strikingly, a large cohort of intrinsically disordered and mitotically phosphorylated proteins is stabilized and solubilized in mitosis, suggesting a fundamental remodeling of the biophysical environment of the mitotic cell. Our data represent a rich resource for cell, structural, and systems biologists interested in proteome regulation during biological transitions

Energy scaling, second Stokes oscillation and Raman gain-guiding in monolithic diamond Raman lasers

Energy scaling of the 1st Stokes oscillation is compared in micro-lensed and plane-plane monolithic diamond Raman lasers under 532 nm pumping. A maximum Raman pulse energy of 92 µJ at 573 nm was achieved with the micro-lensed device, while in a plane-plane configuration the maximum Raman pulse energy was 3.1 mJ. 2nd Stokes generation at 620 nm in 2 and 1 mm long micro-lensed monolithic diamond Raman lasers is also reported. The best conversion efficiency from the pump at 532 nm, namely 63 %, was observed in a 2 mm long crystal at the pump pulse intensity of 4.5 GW/cm2. By measuring the output Raman laser beam caustic it was found that the 2nd Stokes intracavity beam radius at the output coupler of the micro-lensed device is at least two times smaller than that expected from the ABCD matrix calculations of the resonator mode. A Raman gain-guiding mechanism is suggested to explain this difference