32
2. TISSUE EXPOSURE
compact mass and then to treat small aggregates as though they were
solid tissue blocks. Thus, the "buffy coat" of centrifuged blood can be
handled effectively in this way (Low and Freeman, 1958), as also can be
particulate fractions of cells (Palade and Siekovitz, 1956).
For some suspended specimens repeated centrifugation at each preparative stage might prove harmful, or at least tedious, and result in the
loss of too much material. Then it often will prove practical at one stage
or another to mix the warmed pellet (45 °C) with a drop or two of
similarly warmed 2% agar. This then can be removed to a cooled microscope slide and cut into small cubes after the agar has set. In subsequent
operations these are treated as tissue blocks.
In dealing with blood in particular, it is desirable to avoid coagulation.
Low and Freeman (1958) advocate the use of silicone-coated glassware.
This is prepared by its immersion in a 2% solution of Dow-Corning #200
antiwetting agent dissolved in carbon tetrachloride, followed by baking
for 0.5-1 hour at 450-550°C. Hypodermic needles and/or other steel
surfaces are coated with Armour Company Monocote by immersion,
followed by draining and air drying.
2.10. The Use of Cold-Blooded and Young Animals
People using tissues from cold-blooded animals have a great advantage
over those who insist upon studying mammals. These tissues apparently
can stand much more abuse. This is particularly true if the animals have
been prechilled so that the tissues are cold. Dissection then can be a
relatively leisurely affair, and the animals can even be dead for an extended period of time and still yield well-preserved material.
Reference already has been made to the success of Palay and his coworkers (1962) in perfusing fish with an osmium tetroxide fixative. It
would seem likely that the method could be adapted without much difficulty to amphibians, and even to reptiles.
To a certain extent newborn mammals can be treated similarly. Usually
tissue from such babies can be removed after decapitation and still be
well preserved in spite of the inevitable delay. This is certainly simpler
than trying to keep a very young animal alive under anesthesia while a
delicate operation is performed. If the removal of tissue is to be at all
complicated, and take unduly long, the animal can be precooled before
decapitation.
2. TISSUE EXPOSURE
compact mass and then to treat small aggregates as though they were
solid tissue blocks. Thus, the "buffy coat" of centrifuged blood can be
handled effectively in this way (Low and Freeman, 1958), as also can be
particulate fractions of cells (Palade and Siekovitz, 1956).
For some suspended specimens repeated centrifugation at each preparative stage might prove harmful, or at least tedious, and result in the
loss of too much material. Then it often will prove practical at one stage
or another to mix the warmed pellet (45 °C) with a drop or two of
similarly warmed 2% agar. This then can be removed to a cooled microscope slide and cut into small cubes after the agar has set. In subsequent
operations these are treated as tissue blocks.
In dealing with blood in particular, it is desirable to avoid coagulation.
Low and Freeman (1958) advocate the use of silicone-coated glassware.
This is prepared by its immersion in a 2% solution of Dow-Corning #200
antiwetting agent dissolved in carbon tetrachloride, followed by baking
for 0.5-1 hour at 450-550°C. Hypodermic needles and/or other steel
surfaces are coated with Armour Company Monocote by immersion,
followed by draining and air drying.
2.10. The Use of Cold-Blooded and Young Animals
People using tissues from cold-blooded animals have a great advantage
over those who insist upon studying mammals. These tissues apparently
can stand much more abuse. This is particularly true if the animals have
been prechilled so that the tissues are cold. Dissection then can be a
relatively leisurely affair, and the animals can even be dead for an extended period of time and still yield well-preserved material.
Reference already has been made to the success of Palay and his coworkers (1962) in perfusing fish with an osmium tetroxide fixative. It
would seem likely that the method could be adapted without much difficulty to amphibians, and even to reptiles.
To a certain extent newborn mammals can be treated similarly. Usually
tissue from such babies can be removed after decapitation and still be
well preserved in spite of the inevitable delay. This is certainly simpler
than trying to keep a very young animal alive under anesthesia while a
delicate operation is performed. If the removal of tissue is to be at all
complicated, and take unduly long, the animal can be precooled before
decapitation.