The TPCL procures and maintains a biorepository of snap frozen and paraffin embedded tissues, provides fresh tissues for research, and facilitates access to the Department's archived tissue samples. Specific investigator requirements (i.e., placement in particular media or sterile procurement) can typically be met, but requires prior approval. In many cases, malignant and matched tissue samples are available. Tissues are distributed only to UCLA researchers, and IRB approval may be required. Certain requests require approval of the TPCL Tissue Advisory Board. Once your request is approved, turnaround time for distribution of snap frozen/paraffin samples from the TPCL biorepository is 1-2 days. We require 1-2 weeks to retrieve materials from the Department of Pathology archives. Pathologic diagnosis is confirmed on all frozen and paraffin-embedded samples prior to release by four board-certified pathologists. Depending on IRB approval, it may be possible to provide investigators with additional pathology information (e.g., node or margin status) from the original tumor. The TPCL regularly monitors the RNA quality of its samples. Results indicate that RNA from TPCL tissues is more than sufficient quantity and quality for microarray analysis.
Histology-related services include fixation, processing, embedding, and sectioning of human and animal tissues, cryostat sections (frozen tissues), sectioning for RT-PCR or other molecular studies, H&E staining and decalcification of blocks. Special stains, as performed in our diagnostic histology laboratory, are available as well. TPCL staff can advise researchers on tissue preparation, preservation and handling to ensure the highest quality results. Years of experience handling samples from small animals ensure that tissues are properly embedded and precisely sectioned as requested.
These services are directed by the TPCL Co-Director Dr. Jonathan Said, an internationally recognized expert in IHC/ISH. Services include immunostaining of established antibodies as well as optimization of new antibodies and double staining for brightfield; TUNEL assay for apoptosis is also performed. We have a large library of established antibodies available for use on human and animal tissues [IHC Core Antibodies]. Several chromogens (DAB, Fast Red, AP Blue, AP Green) and counterstains (hematoxylin, methyl-green, nuclear fast red) are available. The detection system most frequently used is the HP system using DAB with hematoxylin counterstain. Need to insert instructions for investigators submitting this request (see current page).
Laser capture microdissection (LCM) allows researchers to analyze specific cells within a larger sample. In 2010, TPCL and the California NanoSystems Institutes Advanced Light Microscopy/Spectroscopy (ALMS) core lab jointly purchased a Leica Laser-Microdissection (LMD) 7000. This state-of-the-art LMD technology is housed in the ALMS core, and can be accessed via keycard 24/7/365 once users complete a training class. New features of this instrument include: (1) ability to capture into varied devices, including single PCR tube caps, lab-on-a-chip devices and Petri dishes; (2) a more powerful laser that allows cutting of thick (> 100 micron) sections in a single cut; (3) more precise control of the laser settings to enable collection of individual nuclei; (4) patented beam-steering technology that enables more precise and faster dissection at every magnification; and (5) gravity collection of samples that reduces contamination. Users can sign up for training at the ALMS on-line registration site. Special slides are required for this, and TPCL will prepare all slides for you. Please contact the TPCL for more information.
The TPCL also houses an Arcturus PixCell IIe with single-cell and fluorescence capabilities. The TPCL teaches researchers how to perform LCM, and offers free advice on further molecular analysis (i.e., DNA/RNA extraction) of LCM samples and LCM-related research design. The goal is to make researchers self-reliant in using this technology.
Digital imaging and image analysis services include state-of-the-art virtual microscopy (VM) and digital pathology (DP) (image analysis) services to the UCLA community. In VM, whole glass slides are converted to high resolution digital images (either brightfield or fluorescence) for easy archiving and retrieval, detailed magnification up to 40X, remote viewing via a web-based interface and preparation for publications and teaching - most of which can be performed from a single computer, using free software. DP includes performance of (or instruction in performing) quantitative digital image analysis studies. This includes both quantitative immunohistochemistry and analysis of other cellular characteristics (e.g., cell size). Dr. Clara Magyar directs the digital imaging/image analysis services for the core. The TPCL provides assistance with image acquisition and analysis, tips on sample preparation, and training on our image analysis systems. The TPCL houses several different scanners and image analysis programs (discussed in detail below).Scanners include the Applied Imaging Ariol SL-50 high throughput scanning system (fluorescence, brightfield, image analysis) and the Aperio ScanScope AT high throughput scanning system (brightfield, web enabled).Our automated digital analysis software includes Definiens, Ariol and Molecular Devices MetaMorph.
Why use digital imaging/image analysis? VM/DP permits real-time discussions of histology images posted on secure websites, eliminating barriers to the exchange of pathology information with national/international collaborators/consultants, helping to reduce inter/intra-observer variability that adversely affects quantitative histopathological analysis, improving the overall efficiency of our researchers and clinicians, and eliminating delay, damage, or loss of often irreplaceable slides during shipping between UCLA faculty and extramural sites. Images can be archived, stored indefinitely, and easily retrieved - unlike glass slides, which may be lost, misfiled, damaged or fade with time.
The TPCL houses several different scanners and image analysis programs (discussed in detail below).Scanners include the Applied Imaging Ariol SL-50 high throughput scanning system (fluorescence, brightfield, image analysis) and the Aperio ScanScope AT high throughput scanning system (brightfield, web enabled).Our automated digital analysis software includes Definiens, Ariol and Molecular Devices MetaMorph.
Image Analysis Guidelines:
Assisted sessions for analysis: the user will bring prepared samples and a staff member will scan and perform analysis.
Unassisted sessions for analysis: can be booked only by users that have undergone the training. If users need help, we try to answer questions right away. For questions regarding advanced image analysis, please plan ahead and set up an appointment.
Users who have been trained and who have reserved use times may use our facility 24/7/365 with key card access. Please contact Dr. Clara Magyar or Dr. Dry to obtain keycard access and to reserve use times. .
The download of any programs/files from the PCs is strictly forbidden.
Any instrument malfunction should be reported immediately.
Training for analysis: An appointment must be made in advance to discuss the nature of the project. After slides have been scanned, a training session (duration: 1-2 hours) will be scheduled. Follow-up sessions are encouraged, especially for new users and for people who want to get instruction on topics not covered during the basic session. Please contact Dr. Clara Magyar (firstname.lastname@example.org) for an appointment.
Rules for slides to be scanned:
Aperio: Best results are obtained with glass slides with tissues of regular thickness (4-5 microns), and coverslipped. Slides with marks (such as ink dots), extra mounting medium, non-glass slides, plastic cover slips, thick labels and labels that are going around the slide present challenges for proper imaging, and may need to be removed in order to obtain an optimal scan. Images are individually reviewed once the scanning is completed. Scans with a poor quality will be rescanned manually to achieve the maximum quality possible. Faint stains, air bubbles, thick sections, or marks on the glass surface can cause lower quality scans. In some cases, we recommend sample recut or restaining.
Ariol: Best results are obtained with glass slides with tissues of regular thickness (4-5 microns). Slides must be coverslipped. Coverslips cannot be mounted anywhere on the slide label region. The slide label must be completely dry and absent of any mounting residue. Avoid sample placement near slide edges as samples may be beyond the physical scanning limits of the microscope.
Recommended software for your computer:
Recommended Imaging books: