OTF 5000/HS/LS-001/2/3/4 Iss 4 June 2012
25
6 Use your left hand to press down on the top of the feed screw (where the rewind knob fits
on) and pull towards you. Use upwards counter pressure on the hinge segment with your
right hand at the same time.
7 The trunnion nut will come out of the clips in the hinge section, and the entire feed screw
assembly can now be lifted out from the left.
8 Ensure the trunnion nut is midway on the feed screw.
9 Position your right hand as in ‘5’ above and insert the alternative feed screw assembly in
from the left, directing the top of the screw through the hole in the top arm.
10 Locate the ball at the bottom of the assembly into the corresponding recess in the pawl
arm assembly.
11 With the lugs on the trunnion nut positioned towards the sides, locate them under the
clips beneath the hinge segment (this can only be done by feel).
12 Check that both lugs are held by the clips and that the trunnion nut ball is still properly
located.
13 Before refitting the cover, rewind the feed screw to the top, (i.e. furthest from the large
toothed wheel), by using the rewind knob. Remember to remove rewind knob to enable
the microtome cover to be refitted.
14 Replace the microtome cover and then place the microtome back in to the cryostat
chamber, remembering to refit the rewind knob.
3 CUTTING & COLLECTING SECTIONS
3.1 BASIC FREEZING TECHNIQUES
As a general rule, the quicker tissues are frozen the better the results will be. The aim is to
minimise the damage caused by ice crystal artefact, and the more rapid the transition from
liquid phase to solid phase, the smaller the ice crystals will be. The susceptibility to damage
varies between tissue types, with certain tissues (e.g. muscle) being very easily damaged.
Furthermore, the nature of the work may dictate the level of tissue disruption which is
acceptable. In all cases it is important to keep the specimen size minimal, i.e. at least one
dimension should be no more than a few millimetres.
3.1.1 LIQUID NITROGEN
With a boiling point of -196ºC, LN
2
is an ideal freezing agent. Tissues may be immersed directly,
or more frequently a solvent, such as hexane, is pre-cooled in LN
2
then specimens are
immersed in the solvent.
3.1.2 CARBON DIOXIDE - GAS
The traditional method of freezing. A blast of gas from a CO
2
cylinder will achieve temperatures
around -60ºC. Care must be taken with pressurised gas, and the risk of creating an aerosol of
potentially infectious tissue particles must be borne in mind.
3.1.3 CARBON DIOXIDE - SOLID
This can be used in a bath of solvent, such as hexane, to achieve temperatures around -60ºC.
Safer than CO
2
gas, solid CO
2
(cardice) remains one of the most popular methods of freezing.
3.1.4 FREEZER PLATES
Some cryostats incorporate special cold plates for rapid freezing, covering the temperature
range -45ºC to -60ºC. The OTF cryostat has a standard quick freezer plate that operates at
approximately 10ºC below chamber temperature. The /QF optional feature, available on the
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