OTF 5000/HS/LS-001/2/3/4 Iss 4 June 2012
27
3.2 FITTING THE SPECIMEN
For setting up tissue, proceed as follows:
1 Remove the frost elimination cover from the quick freezer plate.
2 Make sure that the object holder is above 0ºC, if it is at room temperature that is fine and it
just takes a little longer. It is important to note that embedding compound does not stick to a
surface that is below 0ºC.
3 Place a small amount of embedding compound on the top surface of the object holder and
make sure that it covers all of it in order to get the maximum area for adhesion. Try to avoid
spilling embedding compound over the side.
4 Place the object holder in one of the semicircular recesses of the quick freezer plate so that
the machined slot fits with the top of the object holder a few mm above the level of the plate.
Alternatively if the quick freezer plate has frosted over too much then object holders may be
stood on the top of it.
5 Before the embedding compound starts to freeze place the tissue in the correct orientation,
holding it with a pair of forceps. Take care that the forceps do not freeze to your tissue. If
your tissue is fresh you will see the freezing isotherm gradually rise through the tissue, as a
white line.
6 Remove the object holder from the quick freezer and place it in the V block or cryomatic
block and tighten the screw firmly to hold it in position. Wait a few minutes for temperatures
to equalise before trimming.
It is often best to set up a new blade, knife or anti-roll plate using some embedding compound
first. Proceed as follows:
1 Follow steps 1 to 4 above.
2 Watch the embedding compound and suddenly it will be seen to freeze as it turns from
transparent to a dense white colour. Watch the freezing isotherm until just before it reaches
the top surface of the embedding compound and while it is still liquid, carefully apply some
more embedding compound until it forms a mound on the top. Then let it continue to freeze
until it is completely solid.
3 Remove the object holder from the quick freezer and place it in the microtome.
Note 1: If you are unlucky enough to have your object holder freeze solidly into ice on the quick
freezer plate, a little 90% ethanol solution around the base or neck of the chuck will act as an
antifreeze to release it. Be careful if you do this because the ethanol will also lower the melting
point of your specimen.
Note 2: If you are using pre-frozen tissue then it will be much colder than its optimum cutting
temperature and therefore plenty of time must be allowed for temperature to equalise in the
specimen before cutting.
Note 3: It is important that after use the frost elimination cover should be replaced on the quick
freezer plate.
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