19 Coelenterata
223
After 10 min, the living larvae that have collected near the light are
siphoned off through a piece of rubber tubing into a fine net (125
mesh), where they are washed briefly with tap water. The net is then
inverted into a small bowl (112.5 mm diameter), and the cleaned brine
shrimps are fed to the hydra without delay, because Artemia larvae
gradually lose their motility when placed in fresh water.
- Feeding hydra
Approximately 30 min after feeding the hydra with the washed brine
shrimps, the cultures of hydra should be inverted over a sink and refilled with clean culture solution. This operation serves to remove the
uningested brine shrimps from the hydra, which meanwhile remain
firmly attached to the bottom and sides of the culture dish. If this is
not done, the brine shrimps die in the fresh water of the hydra cultures, and soon begin to decompose. In general, hydra demand strict
cleanliness, daily feeding, and frequent changes of water if they are to
increase logarithmically.
Unattached hydra that pour off in the culture water on inversion may
be collected by either of two methods. With small cultures, the best
method is to swirl the decanted water in round bowls; it will be found
that the hydra is collected in the center as the water comes to rest.
With large amounts of water, a net may be used to retain the hydra. A
38 mesh net, for example, retains almost all the hydra while not retaining the smaller brine shrimps. A 125 mesh net retains all hydra
and is useful when removal of shrimps is not involved. It is advisable
to clean the hydra 30 min after feeding because the fully fed hydra
differ maximally in size at this time compared with the smaller brine
shrimps.
Approximately once a week, the culture vessels themselves should be
cleaned to minimize the gradual accumulation of slime within a culture. For this, the attached hydra are loosened with the fingertips and
then poured into a series of large bowls (187.5 mm diameter). After
the culture vessel has been cleaned, the spun-down hydra are returned
to the now clean culture vessel. Mechanical damage to the hydra during this operation is not important because hydra have an almost total
ability to regenerate following mechanical injury.
- Trypsinization: the following is the trypsinization method ofYoungner
(1954): Mince animals or organs using scissors into pieces approximately 4-5 mm in diameter
Wash the tissue fragments several times with saline, and suspend the
fragments in trypsin solution (0.25% Bacto-Trypsin 1 : 250 (Difco
Laboratories) in a Ca 2 + - and Mg2+ -free salt solution).
Discard the trypsin solution after 10 min at 37°C.
Add fresh trypsin solution, and transfer the suspension to a Waring
blender.
Operate the blender at a speed that produces agitation of the tissue
without cutting or foaming.
223
After 10 min, the living larvae that have collected near the light are
siphoned off through a piece of rubber tubing into a fine net (125
mesh), where they are washed briefly with tap water. The net is then
inverted into a small bowl (112.5 mm diameter), and the cleaned brine
shrimps are fed to the hydra without delay, because Artemia larvae
gradually lose their motility when placed in fresh water.
- Feeding hydra
Approximately 30 min after feeding the hydra with the washed brine
shrimps, the cultures of hydra should be inverted over a sink and refilled with clean culture solution. This operation serves to remove the
uningested brine shrimps from the hydra, which meanwhile remain
firmly attached to the bottom and sides of the culture dish. If this is
not done, the brine shrimps die in the fresh water of the hydra cultures, and soon begin to decompose. In general, hydra demand strict
cleanliness, daily feeding, and frequent changes of water if they are to
increase logarithmically.
Unattached hydra that pour off in the culture water on inversion may
be collected by either of two methods. With small cultures, the best
method is to swirl the decanted water in round bowls; it will be found
that the hydra is collected in the center as the water comes to rest.
With large amounts of water, a net may be used to retain the hydra. A
38 mesh net, for example, retains almost all the hydra while not retaining the smaller brine shrimps. A 125 mesh net retains all hydra
and is useful when removal of shrimps is not involved. It is advisable
to clean the hydra 30 min after feeding because the fully fed hydra
differ maximally in size at this time compared with the smaller brine
shrimps.
Approximately once a week, the culture vessels themselves should be
cleaned to minimize the gradual accumulation of slime within a culture. For this, the attached hydra are loosened with the fingertips and
then poured into a series of large bowls (187.5 mm diameter). After
the culture vessel has been cleaned, the spun-down hydra are returned
to the now clean culture vessel. Mechanical damage to the hydra during this operation is not important because hydra have an almost total
ability to regenerate following mechanical injury.
- Trypsinization: the following is the trypsinization method ofYoungner
(1954): Mince animals or organs using scissors into pieces approximately 4-5 mm in diameter
Wash the tissue fragments several times with saline, and suspend the
fragments in trypsin solution (0.25% Bacto-Trypsin 1 : 250 (Difco
Laboratories) in a Ca 2 + - and Mg2+ -free salt solution).
Discard the trypsin solution after 10 min at 37°C.
Add fresh trypsin solution, and transfer the suspension to a Waring
blender.
Operate the blender at a speed that produces agitation of the tissue
without cutting or foaming.