104
4.
EMBEDDING
prepolymerized syrup can be stored for days or weeks in a refrigerator
and used as needed. The latter procedure is regarded as a useful convenience in many laboratories for all embedding, quite apart from questions of specimen damage. But refrigerated containers must be allowed
to warm almost to room temperature before opening them or water condensation may result. Large bubbles are apt to be formed and trapped in
capsules containing prepolymerized methacrylate unless heat is applied
to their bottoms at least as fast as to their tops. Therefore preheated racks
should be used.
Prepolymerized methacrylate has the additional advantage that it can
be adapted to an open-face, flat embedding (Borysko, 1960). In this way
smears, spreads, hanging drop cultures, and the like can be embedded
while attached to glass surfaces. After impregnating the material on a
slide with monomer, aluminum foil is bent and folded around the slide
to create a shallow tray. Prepolymerized plastic is then poured into this
to a depth of 3 or 4 mm. The temperature is slowly raised to approximately 100°C which is maintained for 10-24 hours, after which it is
slowly cooled. (It would not be possible to use monomer in this way in
uncovered containers for it would evaporate quickly.) Finally, the layer
of plastic is stripped away from the glass carrying the specimen material
with it. A variant method proposed by Howatson and Almeida (1958) inverts the lower half of a capsule containing prepolymerized methacrylate
over the desired cells. The capsule in contact with the glass is then put
into the oven for hardening. If a thin glass surface such as a cover slip
has been used, it can be detached from the methacrylate easily by placing it on a piece of solid carbon dioxide for a minute or two. They warn
that it is then important to prevent water condensing on the cool block
as this may cause swelling and distortion of the cells. They avoid condensation by transferring the blocks immediately to a desiccator and leaving them there until they have regained room temperature.
Efforts have been made to eliminate polymerization damage in various
other ways. Shipkey and Dalton (1959) have advocated the use of azodiisobutyronitrile as a nonoxidant catalyst for methacrylate embedding.
They use it in concentrations varying from 0.5 to 1.5% with heat polymerization at 47°C (higher polymerization temperatures are apt to produce bubbles). They particularly recommend it for tissue culture material, blood cells, and cells on free surfaces where polymerization damage
is a problem. Since this catalyst can be used to make prepolymerized
syrup, one can perhaps obtain a double measure of protection. The au-
4.
EMBEDDING
prepolymerized syrup can be stored for days or weeks in a refrigerator
and used as needed. The latter procedure is regarded as a useful convenience in many laboratories for all embedding, quite apart from questions of specimen damage. But refrigerated containers must be allowed
to warm almost to room temperature before opening them or water condensation may result. Large bubbles are apt to be formed and trapped in
capsules containing prepolymerized methacrylate unless heat is applied
to their bottoms at least as fast as to their tops. Therefore preheated racks
should be used.
Prepolymerized methacrylate has the additional advantage that it can
be adapted to an open-face, flat embedding (Borysko, 1960). In this way
smears, spreads, hanging drop cultures, and the like can be embedded
while attached to glass surfaces. After impregnating the material on a
slide with monomer, aluminum foil is bent and folded around the slide
to create a shallow tray. Prepolymerized plastic is then poured into this
to a depth of 3 or 4 mm. The temperature is slowly raised to approximately 100°C which is maintained for 10-24 hours, after which it is
slowly cooled. (It would not be possible to use monomer in this way in
uncovered containers for it would evaporate quickly.) Finally, the layer
of plastic is stripped away from the glass carrying the specimen material
with it. A variant method proposed by Howatson and Almeida (1958) inverts the lower half of a capsule containing prepolymerized methacrylate
over the desired cells. The capsule in contact with the glass is then put
into the oven for hardening. If a thin glass surface such as a cover slip
has been used, it can be detached from the methacrylate easily by placing it on a piece of solid carbon dioxide for a minute or two. They warn
that it is then important to prevent water condensing on the cool block
as this may cause swelling and distortion of the cells. They avoid condensation by transferring the blocks immediately to a desiccator and leaving them there until they have regained room temperature.
Efforts have been made to eliminate polymerization damage in various
other ways. Shipkey and Dalton (1959) have advocated the use of azodiisobutyronitrile as a nonoxidant catalyst for methacrylate embedding.
They use it in concentrations varying from 0.5 to 1.5% with heat polymerization at 47°C (higher polymerization temperatures are apt to produce bubbles). They particularly recommend it for tissue culture material, blood cells, and cells on free surfaces where polymerization damage
is a problem. Since this catalyst can be used to make prepolymerized
syrup, one can perhaps obtain a double measure of protection. The au-