The Molecular Pathology Core is a fully established facility, allowing for the study of gene expression within diseased animal and human tissues. The PixCell IIe Laser Capture Microdissection System by Arcturus, plays a pivotal role in the facility, and has been fully integrated into the specialty core in Research at St. Michael’s Hospital.
The PixCell II Laser Capture Microdissection System and Leica CM 3050S Cryostat.
Laser capture microdissection (LCM) provides users with a reliable method to procure pure populations of cells from unique regions of tissue sections during direct microscopic visualisation (resolution 7.5 microns). Cells of interest are captured by transfer to a heat-sensitive ethylene vinyl acetate plastic polymer using an infra-red laser. Samples are then extracted in microtubes that are suitable for nucleic acid analyses (PCR and RT-PCR for DNA or RNA, respectively) or protein study (Western immunoblotting or 2D-analysis).
A second laser dissection system, the Leica LMD 6500, is also available for use in the core. This system combines high laser power and high repetition rates to collect contact-free specimens by gravity. Laser microdissection using the LMD 6500 requires the use of uv absorbing, membrane coated slides on which tissue sections are mounted. Along with the LCM, the lab consists of a dedicated Cryostat (Leica CM 3050S), an ND-1000 spectrophotometer (NanoDrop Technologies Inc.), as well as a real-time PCR machine (7900 HT SDS, Applied Biosystems/Life Technologies). We are therefore equipped to take fresh, snap-frozen tissue samples, section (5-10 microns thickness) and stain them, dissect out the cells of interest, and proceed with further downstream analysis of the extracted DNA or RNA. This major advance in linking pathology and molecular biology is currently being exploited by users in Research at St. Michael’s Hospital under the guidance of Brent Steer who co-ordinates this specialty service.
Use of the molecular pathology core lab is restricted to those previously experienced or trained on the equipment. One can receive training in-house with pre-arranged scheduling.
| Fee for service | User fee | |
|---|---|---|
| LCM | $100 per hour plus caps* | $50 per hour plus caps* |
| Real-Time PCR | $50 per 384 well plate** | |
| Cryosectioning | $50 per hour | $25 per hour |
| Training (Real-Time PCR,LCM, cryosectioning): | $50 per hour |
* caps @ $4.76 ea.
** one 384 well run approx. 2 hrs.
plates @ $4.45 ea.
| PicoPure RNA Isolation Kit 40 isolations | ARCTURUS Cat.# KIT0204 | List Price $283.50 CDN |
| 10 pk, Taqman Universal PCR MasterMix, 10,000 reactions | Applied Biosytems Cat. # 4324020 | $2,854.80 CDN |
| 10 pk, Power SYBR Green PCR MasterMix, 10,000 reactions | Applied Biosytems Cat. # 4368708 | $2,100.00 CDN |
| PicoPure RNA Isolation Kit 40 isolations | ARCTURUS Cat.# KIT0204 | List Price $283.50 CDN |
| 10 pk, Taqman Universal PCR MasterMix, 10,000 reactions | Applied Biosytems Cat. # 4324020 | $2,854.80 CDN |
N.B. 10,000 reactions based on an assay volume of 10 microlitres
University of Toronto/St. Michael’s Hospital
Li Ka Shing Knowledge Institute Room 522
Brent Steer, coordinator
Phone: 416-847-1735
brent.steer@utoronto.ca
