Imaging Services - BIDMC

Imagine Core B- BIDMC Services

The HDDC Imaging Core B at the Beth Israel Deaconess Medical Center (BIDMC) consists of two interacting components:

The Histology core, provides service for paraffin embedding, sectioning, staining, and frozen sectioning and the Imaging/Microscopy core, provides shared equipment and service for confocal, widefield, photodocumentation, electron microscopy, and digital image processing.  In addition, the microscopy core has an immunostaining service. 

All work is done on a fee-for-service basis to recover costs for equipment upkeep, supplies, and technical assistance.

Facilities and Equipmentming

Facilities

The HDDC imaging core B at BIDMC is located on the 8th floor of the Charles A. Dana Research Building on the East Campus of the BIDMC (please note:  this is NOT at the Dana Farber Cancer Center!!) and consists of more than 2,000 square feet of space.  Maps and other information about access, pricing, and services can be found at http://www.bidmc.org/Research/CoreFacilities.aspx.  Please use this main site to access the individual component Histology or Imaging/Microscopy core links for information.

EQUIPMENT 

Histology 

1) Leica TP1020 Tissue Processor 

This tissue processor is used to dehydrate and infiltrate tissues/cells with paraffin. The instrument has vacuum function and a fume control system. The carousel configuration has 12 stations; 10 for reagents and 2 for melted paraffin. Two loading baskets can be run each night with an 80 cassette capacity per basket. Infiltration time is separately programmable for each station. There is a delayed start function up to 9 days; this allows processing over the weekend to be ready on Monday morning. The machine also has the capacity to interrupt an automatic process for reloading or removing cassettes for special applications before the end of a run. 

2) Leica EG1160 Embedding Center Dispenser and Hot Plate 

This equipment is used to embed tissues/cells that are infiltrated with paraffin. The machine has a separately heated paraffin dispensing system. It additionally has a compressor-cooled cold spot for cooling blocks and a melted paraffin holding reservoir for the newly processed cassettes. 

3) Leica XL Autostainer 

This equipment is used to stain paraffin sections. It has the capability of programming numerous special stains or customer-specific modifications of the H&E stain. There are18 reagent stations, 5 wash stations, and an integrated forced hot air oven that significantly reduces slide drying time. The instrument has the capability of continuous loading - it can load up to 11 racks (with 30 slides per rack) while processing. The capacity is 150 slides per hour with a typical H&E protocol. The machine also allows for simultaneous staining of several different protocols - the user can select any compatible program for each rack. The instrument has self-contained fume control with a replaceable filter. The microprocessor controller can handle up to 15 programs with up to 25 steps each. 

4) Leica IP S Automated Slide Printer, which is an ink-jet printing module that is used for high-throughput printing of labels for histology slides. 

5) Leica CM 1850 Cryostat, which is used to prepare sections from frozen tissue samples for immunostaining and analysis by confocal microscopy or for immunoperoxidase methods. 

6) Leica RT2125 Rotary Microtomes (3), which are used to cut paraffin sections. 

Electron Microscopy 

1) Ultracut E Ultramicrotomes (2), which are used to cut thick and ultrathin plastic sections. 

2) Ultracut-S Ultramicrotome, which is used to cut thick and ultrathin sections from frozen tissue. 

3) JEOL JEM-1400 Transmission Electron Microscope 

This microscope is used for the visualization of tissues by electron microscopy. This updated electron microscope is outfitted with both side (rapid acquisition with a wide viewing area) and bottom (high resolution) Gatan Orius 11 mexapixel ccd cameras, the latter being used to acquire images for electron tomography. The microscope is also outfitted with a 70 degree tilting goniometer, a Fischione dual axis specimen holder and a single tilt specimen holder for doing both single and dual-axis tomograms. 

4) Bal-Tec MED 020 Vacuum Evaporator, which is used to deposit a fine carbon film on formvar coated grids and to glow discharge the grids before ultrathin sections are placed on the grid. 

The electron microscopy core also has a glass knife breaker, numerous diamond knives for both plastic and cryosectioning, a custom machine to pull thin films for making formvar coated grids, and ovens, rotators, and other small equipment needed to prepare samples. 

Confocal Microscopy 

1) Zeiss LSM880 Upright Confocal System (expected delivery in March/April of 2015) 

This laser scanning confocal system will replace the upright 510 META, below. It will be outfitted with 6 laser lines (404, 458, 488, 514, 561, and 633), 3 PMT's, a motorized scanning stage and stepper motor, 10x (air), 20x (air), 40x (water corrected), 63x (oil), 100x (oil) objective lenses with DIC optics; Zen acquisition software with the tiling module, a high capacity computer and 30 inch high resolution monitor. 

2) Zeiss LSM510 Meta Live-Cell Confocal System 

This inverted confocal system is used to visualize physiological processes in live cells. The instrument has a completely enclosed incubator system with an O2/CO2 regulation for long-term support of live cell experiments. It is configured to acquire long time-lapse data, to do FRET and ratio imaging using calcium and intracellular pH dyes, and also serves as a secondary instrument for the acquisition of data from fixed tissue/fixed cell samples, when needed. The specific features of this instrument are as follows: Axiovert 200 (M27 stand) with joystick control; Life Imaging Services environmental incubator; heated stage; Brick and Cube O2,CO2, and humidity control; assortment of Ludin live imaging chambers; Eppendorf InjectMan N1 2 with FemtoJet microinjection system; 10x (air), 20x (air), 40x (water corrected), 63x (oil), 100x (oil) objective lenses and DIC optics; 405 nm (near-UV), 488 nm (green), 543 nm (red), and 633 nm (far-red) lasers; Zeiss Zen Physiology software for FRET, time lapse acquisition, etc.. 

Widefield fluorescence microscope with photodocumentation 

1) Zeiss Axioimager M1 microscope with color and B/W CCD cameras 

This upright widefield microscope is used for photodocumentation of stained paraffin sections and for fluorescence imaging of tissue sections. A sophisticated physiology software package accompanies this software to extend its processing capabilities. The specific features of this instrumentation are as follows: AxioCam MRm (B/W) and HRc (color) CCD cameras; X-Cite Mercury Vapor Short Arc Lamp; 5x (air), 10x (air), 20x (air), 40x (oil), 63x (oil), and 100x (oil) objective lenses; red, green, blue and far-red fluorescence filters; Zeiss AxioVision software including the Physiology package; additional off-line work station. 

Computers-Image Processing 

1) Dell T7610 computer workstation (expected delivery in November of 2014) . 

This computer workstation contains two Intel Xenon E5-2630 processors; Windows 7 Professional 64 bit; 256GB 1866MHz EEC RDIMM memory, PNY Gforce GTX982 Graphics Card, keyboard, 32 inch high resolution monitor, and laser mouse. 

2) Dell T7500 computer workstation 

This computer workstation contains a dual quad core processor, 24GB 1333 MHz SDRAM memory, 256MB ATI fireMV 2260 video card, Windows 7 professional 32 bit, keyboard, 30 inch monitor, and laser mouse. 

3) Dell server, which is used to run the Volocity dongle. 

Software-Image Processing 

1) Image J is freeware from the NIH that is used by most labs, thus it will not be discussed further. 

2) Volocity visualization (Improvision/Perkin Elmer) 

This software allows for the rapid, interactive, high resolution volume rendering of 3D and 4D data sets. This program will accept a range of file types from confocal, wide field, and electron microscopes. The user is able to do very high quality rendering (via the Ray Tracer), navigate through 3D volumes, create images to export, and make quickTime virtual reality movies. 

3) Volocity quantitation (Improvision/Perkin Elmer) 

This software allows for a range of options to analyze 3D and 4D image data. This is used to measure and track objects, determine whether or not objects are co-localized, analyze ratio and FRET images, and to create 2D and 3D interactive charts and graphs of the measurements. 

4) SerialEM (the University of Colorado, Boulder) 

This software is used to acquire tilt series for electron tomography, large image areas for 3-D reconstruction from serial sections, and images for reconstruction of macromolecules by single-particle methods. The main focus of the program is automated tilt series acquisition. For this, it uses an approach based on prediction of specimen position during the tilt series from the position at previous tilts. It does not count on the microscope or the specimen being particularly well-behaved, so unless a prediction appears reliable, it falls back to measuring and adjusting defocus and/or specimen position when necessary. With this method, it achieves high quality tilt series with both robustness and speed. 

5) IMOD (the University of Colorado, Boulder) 

This software is a set of image processing, modeling, and display programs used for tomographic reconstruction and for 3D reconstruction of EM serial sections and optical sections. The package contains tools for assembling and aligning data within multiple types and sizes of image stacks, viewing 3-D data from any orientation, and modeling and display of the image files. This program is free-ware, readily available to academic users. We typically use IMOD for assembling 3-D tomograms and use Volocity software for assembling 3-D stacks from the confocal system. 

6) Chimera (UCSF) 

This software is a highly extensible program for interactive visualization and analysis of molecular structures and related data, including density maps, supramolecular assemblies, sequence alignments, docking results, trajectories, and conformational ensembles. We use this program, in particular, to produce high-quality images and animations (movies). Chimera is free-ware for academic uses. 

Available through Core B: Harvard Center for Biological Imaging, HMS 

Lightsheet Microscopy 

1) Zeiss Lightsheet Z1Microscope 

This instrument provides state-of-the-art long-term, high-speed 3D imaging with virtually no phototoxicity or bleaching. This revolutionary microscope from ZEISS was the first of its kind to be installed in North America (it was prototype #3, installed after production). The samples are immobilized in agarose or tubing, immersed in a buffer, and can be freely rotated during imaging. 3D imaging that previously took hours on a point scanner can now be accomplished in minutes with this instrumentation. This instrument was updated with the latest capacity in October of 2014. 

Super-Resolution PALM/SIM (and 3-D SIM) Microscopy 

1) Zeiss ELYRA PS.1 Super-resolution Microscope system 

This expensive, state-of-the-art equipment has the capability of widefield microscopy, laser widefield, TIRF, spectral confocal imaging, and super-resolution using either structured illumination imaging (SIM) or PALM/dSTORM. Using structured illumination, this system can improve resolution in X, Y, and Z two-fold – with no special sample prep needed and is suited for high resolution 3-D imaging. Photomanipulation super-resolution techniques like PALM or dSTORM can be done to improve resolution by as much as an order of magnitude, which is applicable to single molecule imaging. 

Confocal microscopy 

1) Zeiss LMS 880 with Airyscan and GaAsP technology 

Although the BIDMC core has confocal microscopy capacity, if users need more advanced instrumentation it is available in the HDBI core. The Zeiss LSM 880 Confocal System, in comparison to the new instrument in the BIDMC core, will be outfitted with Airyscan (increases resolution) and a transmission detector for spectral imaging. At present, the latter two applications are not available on the Zeiss 880 at BIDMC. 

Whole-slide scanning (large sections with rapid acquisition) 

1) Zeiss AxioScan Z1 

This equipment can be used by HDDC investigators to do high speed slide scanning with 100-slide capacity, and can image samples stained with both colorimetric and fluorescent dyes. 

Multi-photon confocal imaging 

1) Zeiss LSM 780 NLO Multi-Photon system 

This multi-photon system is designed to image deep into living tissue. Using the 710 NLO with the specialized SCA/E objective, it is possible with this instrumentation to visualize deep into clarified tissue with incredible resolution. Even without clarifying the samples, this instrument excels at penetrating deep into live or fixed specimens, and is capable to create useful 3D datasets. 

Although there are more than 16 instruments available to HDDC users in the HCBI, we believe that the equipment listed is most likely to be used by HDDC members.