Yale University West Campus Analytical Core Laboratory
The West Campus Analytical Core (WCAC) was launched in 2012 to provide the research community at West Campus with a comprehensive analytical instrumentation facility that provides access and training to a broad collection of shared instrumentation. The mission of the WCAC is to support and accelerate ongoing research projects and to provide suitable space for incremental additions of high-value shared instrumentation.
The main instrumentation lab and staff is located at the West Campus Molecular Innovations Center (building 27) room 109 with instruments also located in MIC room 310, The Advanced Bioscience Center (building 31) and other locations on West Campus
Instruments: Mass Spectrometers
Agilent 6550 Q TOF
The Agilent 6550 Q-TOF LC/MS System,(MIC room 109) uses iFunnel technology to deliver the lowest detection levels of any high resolution LC/MS instrument currently available. Low femtogram-level sensitivity with high resolution across a relatively broad mass range (as compared to Orbitrap) as well as accurate mass make it a versatile platform, and a good choice for small molecule/metabolite ID, metabolomics and proteomics applications. We currently have from Agilent a 1290 LC system coupled to the dual spray API source.
iFunnel increases ion transfer to achieve the lowest detection levels of any high resolution LC/MS instrument, achieving low femtogram-level sensitivity
Ion Beam Compression and Shaping (IBCS) technology provides the greatest sensitivity while maintaining 40k mass resolution, sub ppm mass accuracy, and a faster scan rate up to 50 spectra per second.
Agilent Jet Stream technology combined with improved ion optics provide 5x greater on-column sensitivity and increase up time to maintain the highest sensitivity for high throughput applications.
In-spectrum dynamic range of up to 5 orders of magnitude reveals trace-level targets, even in the presence of much more abundant compounds
Agilent 6490 QQQ
The Agilent 6490 triple quadrupole instrument (located in ABC room210) can be used with an ESI source for UHPLC applications or the Chip Cube system. Like the 6550 Q ToF, it employs Agilent’s iFunnel technology which incorporates a thermal gradient “Jet Stream,” a hexabore capillary at the vacuum interface for increased ion sampling and dual stage ion funnel. The 6490 is a high throughput instrument capable of determining 400 transitions per method. It is the most sensitive triple quad available with attogram/ zeptomole level sensitivity and linearity of six orders of magnitude. These performance characteristics make it especially suited for MRM analysis.
Thermo Orbitrap Velos
The Thermo LTQ Orbitrap Velos (located in MIC rm 109) Equipped with a Waters nanoACQUITY UPLC system, allowing nano LCMS and LCMS/MS on-line LC-MS/MS, for data-dependent protein IDs using a high resolution Orbitrap mass analyzer for MS survey scans and an LTQ Velos linear ion trap for CID-based fragment , shotgun-based quantitative proteomics. The Orbitrap mass analyzer is utilized for both MS survey and MS/MS scans under HCD mode and data dependent parallel multiple-stage fragmentation (MSn) analysis. HCD is often utilized to experimentally detect and analyze low mass ions such as the reporter ions used in ITRAQ experiments or to discover other peptide modifications. High mass accuracy (often <1 ppm) and high resolution (up to 100,000) of the Oribitrap mass analyzer allows for increased speed and confidence in identification of complex samples by minimizing false positives. The Velos system in the core is dedicated primarily to proteomics analysis.
• Resolving power of >100,000
• Mass accuracy better than 1 ppm
• Multiple fragmentation techniques:
• Parallel MS and MSn analysis
Agilent 6120 LC-MS
The Agilent 6120 Quadrupole LC/MS system (located in MIC room 109) is a robust, single quad system. It is suitable for routine low resolution LC-MS applications in either Positive or Negative ESI mode, for both large and small molecule analysis. The 6120 sensitivity allows low picogram detection levels with RSDs of less than 5 percent linear response over 3 orders of magnitude. The 6120 in the core is coupled to an Agilent 1260 infinity LC system.
Gas chromatography-mass spectroscopy (GC-MS) combines two technologies to form a single method of analyzing chemical compounds or mixtures. The system is located in MIC room 109.
The Agilent 7890A GC offers state of the art performance features including advanced separation capabilities, and real-time self-monitoring instrument intelligence. Fast oven cool-down and robust back-flushing allow quick cycle times. 5th-generation Electronic Pneumatics Control (EPC) and digital electronics set a new benchmark for pressure set-point and retention time locking precision (0.001 psi) and help make the 7890A a robust instrument platform.
The Agilent 5975C inert MSD (mass spec) is equipped with a new Triple-Axis Detector delivering good MS resolution and low mass deviation, with good sensitivity and spectral integrity.
400 MHz Magnet (1 of 2)
The WCAC operates two 400 MR NMR spectrometers (MIC room 109) each has an upgraded Oxford magnet operated through an Agilent Varian console, featuring DD2 technology, including amplifier linearization, flat baseline-corrected digital filtering and compatibility with Agilent NMR automation solutions.
The 400 MRs are both operated using VNMRJ 3.2. Every data file acquired with VNMRJ has associated files with many of the relevant parameters from the acquisition. This data file can be accessed by opening the associated files in a text editor, or through VNMRJ. These files typically do not contain information about the temperature or sample rotation speed at which the data was acquired.
400 MHz (9.4 Tesla) Oxford Magnet
Two radio frequency channels
TMC antivibration system
Waveform generator for shaped pulses
Z-axis pulsed field gradient module
Variable Temperature unit.
These instruments are used for routine 1D H/C/F/P experiments
Agilent 600 MHz NMR
Agilent 600 MHz -(installed September/October 2012 in MIC room 109) -triple resonance, single-gradient system with HCN and OneNMR probes (both inverse and direct detect).
Magnet: 14.1 Tesla 54 mm bore Agilent Premium Compact Shield Superconducting Magnet
Probes: NMR Probe (1H-19F/15N-31P 5 mm PFG OneNMR Probe)Unique design provides high performance for both high frequency (1H-19F) and low frequency (15N-31P) observe and decouple experiments.
Cold probe-This is currently the primary system probe. It is a 3mm, triple resonance tuned to H,C and N.
Agilent Cary 660 FTIR
The Agilent Cary 660 FTIR (MIC room 109) is a versatile, high performance spectrometer that meets a broad range of routine spectrometry and research analysis. The instrument is controlled using high performance operating software and can also be fitted with a wide range of accessories to provide extra capabilities. The core labs 660 is equipped with a Pike Technologies GladiATR™ Single Reflection ATR accessory. The common platform of Agilent’s Cary 660/670/680-FTIR spectrometers enables the performance and capabilities of the imaging system utilized to be scaled.
Agilent Cary 300 UV-Vis spectrophotometer
The Agilent Cary 300 UV-visible spectrophotometer (MIC room 109) is equipped with a versatile set of accessories and posseses a large photometric range. It is controlled by the Cary WinUV software, modular Windows-based software, featuring powerful analytics run by an easy-to-use interface. The instrument is equipped with liquid sample holders and can be fitted with a wide range of accessories to provide extra capabilities. The instrument is well-suited for materials testing of both turbid liquid and solid samples. The Cary 300 allows routine UV-Vis measurements but has a pre-monochromator allowing an extended range past 5.0 absorbance units (Abs). The instrument is also capable of doing kinetics measurements for determining reaction rates and enzyme activity as well as for thermal denaturation/renaturation measurements.
Thermo Nanodrop 2000C
(located in MIC room 310)
Perkin Elmer EnVision
The 2104 Perkin Elmer EnVision® Multilabel Reader in the core, (MIC room 310) is a state-of-the-art dual-detector model and among the fastest HTS readers available. The instrument includes two detectors enabling simultaneous dual wavelength reading, below emission reading, barcode readers, a high speed light source and adjustment of measurement height function. The Core lab instrument is equipped with stackers, temperature control, dispenser, enhanced luminescence and/or AlphaScreen capabilities.
GE Heathcare Typhoon FLA 9500
The GE Healthcare Typhoon FLA 9500 (MIC room 310) is a variable mode laser scanner with modular access to the optical components, providing both versatile and flexible imaging for precise quantitation of proteins, nucleic acids, and other biomolecules.
- Versatility: imaging of multifluorescent-, chemifluorescent-, radioisotope-labeled, and colorimetric samples
- High resolution and quantitation: a pixel resolution of up to 10 µm and a linear signal response over five orders of magnitude provides precise quantitation in gels, blots, tissue sections, and arrays
- High sample throughput: scanning area of 40 × 46 cm enables simultaneous imaging of up to 20 gels or blots, measuring 10 × 8 cm in size. This facilitates comparisons among blots, and reduces workload and waiting time
- Flexibility: optimized performance for new applications by adapting the system with stages, detectors, filters and lasers
- 2-D DIGE imaging: simultaneously image two 2-D DIGE gels for differential expression studies
- Visible and infrared fluorescence imaging: optional near infrared excitation for imaging IRDyeTM and other infrared dyes
The system provides several imaging modes at any time, including fluorescence and chemifluorescence, filmless autoradiography, and digitization of colorimetrically-stained samples (e.g., CoomassieTM blue and silver stain). It is also adequate for quantitative chemiluminescent imaging that does not require maximum sensitivity.
The system is optimized for 2-D DIGE imaging for differential protein expression studies, ECL Plus chemifluorescent imaging for quantitative protein detection by Western blotting, and multifluorescent ECL Plex imaging for precise quantitation of two or more proteins in the same blot.
Typhoon FLA 9500 comes equipped with both bi-alkali and multialkali photomultiplier tubes (PMT). This combination provides excellent detection over a very broad spectrum. Each PMT is selected for optimal response to the detected emission wavelength.
The system accommodates four automated filters. Filters are easily accessed and exchanged without tools for attaining optimal imaging conditions.
Stages give the optimal imaging conditions for a range of samples. Samples include agarose and polyacrylamide gels, membranes, DIGE gels, radioisotope-labeled samples, as well as microplates and glass slides with the TP plug-in. The system can simultaneously scan two DIGE gels, each measuring up to 20 x 25 cm. The stages are easily removed from the system for cleaning.
A fast 1000 μm prescan function gives a rapid overview of the sample. The system provides a linear signal response over five orders of magnitude, providing a suitable basis for precise quantitation.
Lasers can be exchanged in the field to accommodate new applications and fluorophores.
Note: there is also a Typhoon 9000 located in Building 31 2nd floor cold room annex.
GE Healthcare ImageQuant LAS 4000
The GE Healthcare ImageQuant LAS 4000 (MIC room 310) is a digital imaging system for sensitive, quantitative imaging of gels, blots, and colonies.
- Exceptional sensitivity for quantitation of biomolecules labeled with chemiluminescent probes, and expandable to multifluorescent UV, visible, and IR imaging. Allows gel documentation of Coomassie and silver-stained samples
- Accurate quantitation of closely spaced bands in complex protein samples for gels and films up to 21 x 14 cm (25 x 25 cm using the optional lens). Low noise for longer exposure times and lower background, important for precise quantitation of very weak signals
- Wide linear dynamic range for precise quantitation of faint and dark bands over four orders of magnitude in the same image, avoiding time consuming multiple film exposures
- Distortion, dark frame, and flat frame corrections are applied to each imaging mode for optimal precision and uniform quantitation
- Sensitive detection of less than 40 pg of protein on a Western blot. Sensitivity may be increased by "binning" of up to 8 x 8 pixels and smoothing
- Chemiluminescent and colorimetric signals can be captured in the same image without changing the lens. Optional multifluorescence detection extends functionality to the capture of fluorophores such as Cy2, Cy3, and Cy5 and Deep Purple Total protein stain
ImageQuant LAS 4000 is designed to quantitate the highly sensitive detection of proteins using Amersham ECL Western blotting reagents. A free combination of light sources and filters can be incorporated into the imager according to your needs.
The system consists of an intelligent dark box, camera, lens, and automated four-position filter wheel with a standard filter for EtBr and open positions for the filters delivered with the optional fluorescent light sources. Focusing, filters, illuminators, and exposure time are remotely controlled by a computer. Simple operation allows adjustment without having to open the cabinet. The intelligent light box provides automation and user safety. UV light sources are housed inside the cabinet, and can only be switched on when the door is closed.
A bright, wide aperture FUJINON F0.85 lens, specially developed for chemiluminescence imaging, projects sharp images onto a patterned CCD chip. By diagonally aligning octogonal pixels, the gap between pixels in the horizontal and vertical planes is reduced. This pattern allows an effective image resolution of 6.3 Mpixels.
The user-friendly control software has several capture modes for achieving optimal sensitivity and dynamic range including increment, repetition, and program modes. For image analysis, advanced and flexible ImageQuant TL software is available separately and comprises a dedicated suite of analysis applications to provide high levels of automation and accuracy in analysis of gels, blots, and colony counting.
2D Gel Electrophoresis
2D gel electrophoresis is an analytical technique which combines the independent parameters of charge (through isoelectric focusing-IEF) and size (through sodium dodecyl sulfate-polyacrylamide electrophoresis-SDS-PAGE) to separate proteins. Complex mixtures consisting of over 2000 proteins may be separated on a single gel using the system available at the WCAC. Training is available to enable the use of 2D-DIGE (differential gel electrophoresis), using fluorescent cy-dyes to separately label and run 2 or 3 samples and run them simultaneously on the same gel. The gel may then be imaged using the core's 9410 Typhoon Imager, and the spots compared for relative abundance. The sensitivity of the dyes is comparable to silver staining, with a significantly higher dynamic range, greater than 10^3.
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