226
Methods for Setting Up Primary Cultures Specific to Animal Groups
4. Draw off the Instant Ocean, and replace with Necco's physiological
solution.
5. Remove the Necco's solution after 15 s, and replace with fresh solution.
6. Wash the excised coenosarc six times in the same manner.
7. Cut the tissue lengthwise with cataract knives so that it lies flat.
Culture 1. Transfer the cut tissue to a cover glass, and arrange so that the
set-up
gastrodermis faces upward.
2. Remove the fluid with a Pasteur pipette leaving the fluid sufficient to
cover the tissues.
3. Place one drop of Necco's solution in a depression slide glass to prevent eventual evaporation of the hanging drop. Care must be taken so
that the drop in the depression does not touch the hanging drop.
4. Invert the cover glass.
5. Place the inverted cover glass over the depression, and seal the rim of
the cover glass with a melted mixture of paraffin and vaseline in equal
proportions.
6. Keep the culture in the dark at 18°C.
• • Results
According to Burnett et al. (1968), attachment of the migrated cells starts
within 4-6 h after explanting. During the first 24 h in culture, the spreading monolayer consists of only digestive cells. Divisions of digestive cells
within the monolayer into epidermal epitheliomuscular cells and interstitial cells were observed, but not frequently. After 24 h i-cells began to
invade the monolayer. At this stage, the epidermal cells also started to
extend over the monolayer. From days 3 to 6, cell division and differentiation continued. Before Burnett et al. (1968), Martin and Tardent (1963)
cultured the coenosarc of T. larynx in filtered seawater to which serine,
taurine, glycine, TC-yeastolate, and glucose had been added. In this culture system, cnidoblasats multiplied at a fast rate after the death of all
the other cell types for at least 50 days. Partial differentiation of nematocysts was also observed. Cnidoblasts showed division even when the
nematocyst capsule was already well developed.
Subprotocol 3
Septa of Sea Anemones (Anthozoa)
• • Materials
Equipment - Scissors
- Forceps
- Needles
- Pasteur pipettes
- Tissue culture dishes
Methods for Setting Up Primary Cultures Specific to Animal Groups
4. Draw off the Instant Ocean, and replace with Necco's physiological
solution.
5. Remove the Necco's solution after 15 s, and replace with fresh solution.
6. Wash the excised coenosarc six times in the same manner.
7. Cut the tissue lengthwise with cataract knives so that it lies flat.
Culture 1. Transfer the cut tissue to a cover glass, and arrange so that the
set-up
gastrodermis faces upward.
2. Remove the fluid with a Pasteur pipette leaving the fluid sufficient to
cover the tissues.
3. Place one drop of Necco's solution in a depression slide glass to prevent eventual evaporation of the hanging drop. Care must be taken so
that the drop in the depression does not touch the hanging drop.
4. Invert the cover glass.
5. Place the inverted cover glass over the depression, and seal the rim of
the cover glass with a melted mixture of paraffin and vaseline in equal
proportions.
6. Keep the culture in the dark at 18°C.
• • Results
According to Burnett et al. (1968), attachment of the migrated cells starts
within 4-6 h after explanting. During the first 24 h in culture, the spreading monolayer consists of only digestive cells. Divisions of digestive cells
within the monolayer into epidermal epitheliomuscular cells and interstitial cells were observed, but not frequently. After 24 h i-cells began to
invade the monolayer. At this stage, the epidermal cells also started to
extend over the monolayer. From days 3 to 6, cell division and differentiation continued. Before Burnett et al. (1968), Martin and Tardent (1963)
cultured the coenosarc of T. larynx in filtered seawater to which serine,
taurine, glycine, TC-yeastolate, and glucose had been added. In this culture system, cnidoblasats multiplied at a fast rate after the death of all
the other cell types for at least 50 days. Partial differentiation of nematocysts was also observed. Cnidoblasts showed division even when the
nematocyst capsule was already well developed.
Subprotocol 3
Septa of Sea Anemones (Anthozoa)
• • Materials
Equipment - Scissors
- Forceps
- Needles
- Pasteur pipettes
- Tissue culture dishes