Discussion of Laser Microdissection Projects
Since ideal tissue preparation parameters for mesenchymal tissues and epithelial tissues differ and because considerations of fixation protocols may vary depending upon the ultimate purpose of the microdissected samples, discussion of each project in advance is recommended.
Identifying Areas to Microdissect
Investigators should be aware that identification of structure in uncoverslipped slides is not as easy as on coverslipped preparations. Double-scoping potential samples on the morphology reference (Map) slide to identify areas for microdissection with a pathologist is recommended. Scheduled time with a pathologist may be arranged.
Optimal Tissue Collection for Laser Microdissection
Frozen tissues are recommended for optimal recovery of DNA, RNA and protein.
Harvest tissue samples within 5 minutes of animal death in a sterile environment (RNase free blades and instruments, wear gloves and clean work area).
Place a metal disk or equivalent on dry ice to provide a flat work surface.
Place appropriate size Tissue Tek® Cryomold® on top of the metal disk.
Squeeze a thin layer of OCT into the bottom of the mold and quickly orient the tissue (Figure 1). Label the Cryomold®.
Fill the Cryomold® to capacity and allow OCT to freeze completely (Figure 2). Label Cryomold® with a permanent marker.
Transfer samples to a labeled box or plastic bag and store at -80°C.
Snap freezing in isopentane can destroy tissue morphology and is not recommended for LM procedures.