24
edits
Changes
no edit summary
== Protocol 1 ==
#After harvesting tissue from animal, place into pathology cassette and fix in 4% formaldehyde, 4% paraformaldehyde or 4% formalin O/N
#After fixation, begin dehydrating the tissue in 70% Ethanol
#Remove wax embedded sample from the mold (should just slip out)
#Store at room temperature until prepped to section
== Paraffin embedding protocol 2 ==
#Harvest tissue from animal, place into cassette and submerge in 10% neutral buffered formalin for 24-48 hrs (depending on thickness of sample).
#Move cassettes into 70% ethanol for 24 hrs. Samples can be left in 70% for long term storage. Note that the subsequent dehydration steps involve long incubation periods and therefore should be started first thing in the morning (unless you plan on being in the lab overnight!).
#Move cassettes into 75% ethanol for 30 minutes.
#Move cassettes into 95% ethanol for 75 minutes. Repeat this step a second time, using fresh 95% ethanol.
#Move cassettes into 100% ethanol for 60 minutes. Repeat this step twice more, using fresh 100% ethanol each time.
##During these incubation steps, prepare beakers with 60 deg C paraffin.
#Move cassettes into xylenes for 30-60 minutes. Repeat this step a second time, using fresh xylenes.
#Move cassettes into 60 deg C paraffin for 60 minutes. Repeat this step twice more, using fresh paraffin each time. The second paraffin incubation step can be extended over night, if necessary.
##During these incubation steps, turn on the paraffin wax machine (Link Building, Room 311).
#Immediately prior to embedding, spray wax mold with "Mold Grease" and coat the bottom layer of the mold with wax.
#Take your sample and orient it on top of the base wax as desired. Partially cover with more paraffin.
#Place pathology cassette on top of the mold and completely fill the mold.
#Place mold with sample on "Cold Side" of embedding station .
##Wax will solidify in 10-15 minutes.
#Remove wax embedded sample from the mold (should just slip out).
#Store at room temperature until ready to section.