2 h 2:1 Lowicryl/methanol
2 h Lowicryl (need 5 ml/capsule holder)
Overnight Lowicryl (need 5 ml/capsule holder)
Collect all for hazardous waste disposal.
Day 5
Required materials:
100% Ethanol
Oven-dried gelatin capsules, spider covers, stem holders, bottom spacer
plates, cans, chamber for gelatin capsules
Cryomanipulator with M4 thread
Red temperature tube
Preparations:
10 ml Lowicryl (see recipe above)
Set a stem holder in each of 3 curing containers (label cans) and place a
bottom spacer plate on top of its base.
Procedures (T2, S2, T3):
Chill Lowicryl, pipets and chamber for gelatin capsules.
Gently jiggle/tap capsule holders to encourage tissue to settle at the bottom
of capsules.
Insert gelatin capsules in chamber.
Fill gelatin capsules with Lowicryl.
Screw cryomanipulator into socket of spider cover.
Align arrow of spider cover with position #1 of capsule holder (between 2 dots).
Push spider cover down into capsules, lift slightly and carefully check that
all capsules are attached (you may have to try more than once; don’t lose
your alignment). When possible, move the spider cover with capsules to
the gelatin capsule chamber and insert tissue capsules into gelatin capsules,
pushing down to attach the gelatin capsules to the spider cover. Again, lift
slightly and check for attachment. Unscrew the cryomanipulator, but leave
the spider cover in place. Remove the empty capsule holder, mop up any
spilled Lowicryl, and insert a labeled curing container (prepared as above)
to which 10 ml 100% ethanol has been added. Chill. When chilled, use the
insulated tweezers to transfer the loaded spider cover (again, lift slightly to
check attachment and push down to re-engage if necessary) to the stem
holder. The bottom spacer plate is used to help ensure that even if a gelatin/
tissue capsule detaches in the curing container it will remain in place and
not fall over into the ethanol. Insert and fill the next set of gelatin capsules
(transfer the Lowicryl within the gelatin chamber as needed). Then repeat
as above until all spider cover/capsule sets are seated on their stem holders.
172
K. D. Micheva and K. D. Phend
2 h Lowicryl (need 5 ml/capsule holder)
Overnight Lowicryl (need 5 ml/capsule holder)
Collect all for hazardous waste disposal.
Day 5
Required materials:
100% Ethanol
Oven-dried gelatin capsules, spider covers, stem holders, bottom spacer
plates, cans, chamber for gelatin capsules
Cryomanipulator with M4 thread
Red temperature tube
Preparations:
10 ml Lowicryl (see recipe above)
Set a stem holder in each of 3 curing containers (label cans) and place a
bottom spacer plate on top of its base.
Procedures (T2, S2, T3):
Chill Lowicryl, pipets and chamber for gelatin capsules.
Gently jiggle/tap capsule holders to encourage tissue to settle at the bottom
of capsules.
Insert gelatin capsules in chamber.
Fill gelatin capsules with Lowicryl.
Screw cryomanipulator into socket of spider cover.
Align arrow of spider cover with position #1 of capsule holder (between 2 dots).
Push spider cover down into capsules, lift slightly and carefully check that
all capsules are attached (you may have to try more than once; don’t lose
your alignment). When possible, move the spider cover with capsules to
the gelatin capsule chamber and insert tissue capsules into gelatin capsules,
pushing down to attach the gelatin capsules to the spider cover. Again, lift
slightly and check for attachment. Unscrew the cryomanipulator, but leave
the spider cover in place. Remove the empty capsule holder, mop up any
spilled Lowicryl, and insert a labeled curing container (prepared as above)
to which 10 ml 100% ethanol has been added. Chill. When chilled, use the
insulated tweezers to transfer the loaded spider cover (again, lift slightly to
check attachment and push down to re-engage if necessary) to the stem
holder. The bottom spacer plate is used to help ensure that even if a gelatin/
tissue capsule detaches in the curing container it will remain in place and
not fall over into the ethanol. Insert and fill the next set of gelatin capsules
(transfer the Lowicryl within the gelatin chamber as needed). Then repeat
as above until all spider cover/capsule sets are seated on their stem holders.
172
K. D. Micheva and K. D. Phend