Derek F. Hopkins

An LC-IMS-MS Platform Providing Increased Dynamic Range for High-Throughput Proteomic Studies

A high-throughput approach and platform using 15 min reversed-phase capillary liquid chromatography (RPLC) separations in conjunction with ion mobility spectrometry-mass spectrometry (IMS-MS) measurements was evaluated for the rapid analysis of complex proteomics samples. To test the separation quality of the short LC gradient, a sample was prepared by spiking 20 reference peptides at varying concentrations from 1 ng/mL to 10 microg/mL into a tryptic digest of mouse blood plasma and analyzed with both a LC-Linear Ion Trap Fourier Transform (FT) MS and LC-IMS-TOF MS. The LC-FT MS detected 13 out of the 20 spiked peptides that had concentrations >or=100 ng/mL. In contrast, the drift time selected mass spectra from the LC-IMS-TOF MS analyses yielded identifications for 19 of the 20 peptides with all spiking levels present. The greater dynamic range of the LC-IMS-TOF MS system could be attributed to two factors. First, the LC-IMS-TOF MS system enabled drift time separation of the low concentration spiked peptides from the high concentration mouse peptide matrix components, reducing signal interference and background, and allowing species to be resolved that would otherwise be obscured by other components. Second, the automatic gain control (AGC) in the linear ion trap of the hybrid FT MS instrument limits the number of ions that are accumulated to reduce space charge effects and achieve high measurement accuracy, but in turn limits the achievable dynamic range compared to the IMS-TOF instrument.

Application of coherent antistokes Raman scattering (CARS) to imaging mammalian cells: a means for gaining molecular selectivity in multiphoton imaging

Virtually all laser based microscopy imaging methods involve a single laser, with ultrafast lasers emerging as the enabling tool for a variety of methods. Two-photon fluorescence is a high sensitivity method with selectivity depending on a chromophore that is either added or produced by genetic engineering. While there are fundamental advantages over white light or other fluorescence microscopies, there are unavoidable limitations such as bleaching, photoinduced damage to the cell, and the inability to label some major constituents of the cell, particularly the abundant species. Raman imaging affords chemical selectivity but application is limited due particularly to its low sensitivity and unavoidable fluorescence background. Adding a second laser beam, shifted from the first laser by a molecular vibrational frequency, increases the detected Raman signal by many orders of magnitude and in addition shifts the detected signal to the high energy (blue) side of both lasers, removing fluorescence artifacts. Signal levels sufficient to acquire high signal-to-noise ratio images of 200 by 200 pixels in one minute requires sub-nanojoule pulse energy. A convenient, tunable source of the Stokes-shifted beam is provided by an Optical Parametric Amplifier (OPA), which requires an amplified laser. 250-kHz sources have ample energy and in addition keep the average sample power on the order of 0.1 mW, a level that even sensitive biological systems tolerate at the focal spot diameter of 0.3 micrometers . Long-term viability of mammalian cells has been demonstrated during dozens of scans in a single plane. Two-photon fluorescence provides a useful complimentary data channel that is acquired simultaneously with the Raman image. Several dyes and green fluorescence protein have been used for this purpose. Interpretation of images, acquiring three dimensional images, and identification of cellular features are ongoing activities.

Hanford Tank Farms Waste Feed Flow Loop Phase VI: PulseEcho System Performance Evaluation

This document presents the visual and ultrasonic PulseEcho critical velocity test results obtained from the System Performance test campaign that was completed in September 2012 with the Remote Sampler Demonstration (RSD)/Waste Feed Flow Loop cold-test platform located at the Monarch test facility in Pasco, Washington. This report is intended to complement and accompany the report that will be developed by WRPS on the design of the System Performance simulant matrix, the analysis of the slurry test sample concentration and particle size distribution (PSD) data, and the design and construction of the RSD/Waste Feed Flow Loop cold-test platform.

EVALUATING CONCENTRATION PROFILES DURING UNSTEADY MIXING

Pulse jet mixing tests to suspend noncohesive solids in Newtonian liquid were conducted at three geometric scales. To understand the solids suspension process an ultrasonic concentration probe was used to measure the concentration of solids in the cloud during a pulse at various elevations and radial positions. The data are being analyzed to provide a model for predicting concentration as a function of elevation. This paper presents a simple single frequency ultrasonic measurement application that demonstrates the ability of ultrasonic sensors to measure slurry concentration based on signal attenuation. Sensor calibration data show that ultrasonic signal attenuation is proportional to the applied frequency and to the slurry volume fraction. Real-time measurements of ultrasonic signal attenuation were used to track the process of slurry mixing using single sensors and sensor arrays. Comparison of means of the ultrasonic measurements with means obtained from discrete extractive measurements show that the distributions overlap and cannot be statistically distinguished. The real-time ultrasonic sensor can be used as a primary measurement method or to reduce reliance upon extractive methods to measure slurry density.

A Network Layer for Teleoperations in High Speed Environments

Teleoperation systems allow an operator to control a device at a remote location via a network. System stability is highly dependent on data loss and delay since command messages and the associated device feedback must remain synchronized. Several network protocols are available for teleoperation; however, none is exactly suitable for transmitting the variety of information (e.g. commands, video, and force feedback) needed by advance teleoperation systems. Furthermore, maintaining synchronization is more problematic in a high speed environment since end systems are the primary source of loss and delay. This paper introduces a new network layer designed to control remote devices in a high speed environment. Unlike previous teleoperation approaches, the proposed layer separates control and feedback into separate channels. As a result, multiple feedback channels are possible (e.g. video and force) where each channel can use the transport protocol best suited for its data (e.g. RTP for video). These channels are managed by the new layer ensuring flow control is provided between the operator and the remote device; therefore, different remote control approaches (event and predicted based) are supported. Furthermore, the proposed layer is lightweight and requires no changes to the networking infrastructure since it is only implemented at the end systems (operator and remote device). Experimental results showing the control of a microscope over an 10 Gbps network will demonstrate the teleoperation layer can easily manage communications in a high speed environment.