15 Mollusca
163
- Hyamine 1622 (Rohm and Haas)
Chemicals
- Penicillin
- Streptomycin
- Fungizone (Amphotericin B)
- Chlortetracycline
- 0.2% Iodine solution (Robinson Laboratory)
Solutions
- Buffered physiological solution with antibiotics (BPSA; see Appendix 1)
- Trypsin-EDTA solution (sodium gluconate 1.5 g,NaCI2.8 g,KCI 0.15 g,
Na2HP04 0.07 g, NaHC03 0.05 g, glucose 0.5 g, trehalose 0.5 g, galactose 0.5 g, phenol red 5.0 mg; trypsin (2x crystallized; Sigma) 2.4 g,
EDTA tetrasodium salt (MW = 380) 38.0 mg in 1,000 ml H20)
- Culture media (Basch and Diconza's medium C; see Appendix 1)
- Hansen's M-182, M260 or S-301 media (see Appendix 1)
- Antibiotic solution (penicillin 100 IV/ml, streptomycin 100 flg/ml,
Fungizone 0.25 flg/ml, chlortetracycline 5 flg/ml)
- Lettuce
Miscellaneous
- Rat diet granules (Geisler Pet Products)
- Eggs of snails Biomphalaria glabrata
Animals
Procedure
The following procedure is based on the techniques of Hansen (1979).
1. Maintain the snails on lettuce and rat diet granules.
Oviposition
2. Allow oviposition on plastic rafts overnight.
3. Remove egg sacs carefully (eggs are deposited in groups in gelatinous
sacs).
1. Rinse the egg sacs briefly in 0.2% iodine.
Sterilization
2. Rinse the egg sacs briefly in 0.001 % hyamine 1622.
3. Incubate the egg sacs at 23°C for 4-5 days in a sterile solution of antibiotics in a petri dish.
4. Change the antibiotic solution daily.
5. Select embryos at the trochophore stage.
6. Place approximately 15 egg sacs (containing more than 40 embryos)
in a BPI dish containing 0.4 ml BPSA.
7. Tease the egg sacs apart and release the embryos from the egg capsules.
8. Rinse the embryos by transferring them to another BPI dish containing BPSA.
9. Repeat this rinsing procedure three more times.
163
- Hyamine 1622 (Rohm and Haas)
Chemicals
- Penicillin
- Streptomycin
- Fungizone (Amphotericin B)
- Chlortetracycline
- 0.2% Iodine solution (Robinson Laboratory)
Solutions
- Buffered physiological solution with antibiotics (BPSA; see Appendix 1)
- Trypsin-EDTA solution (sodium gluconate 1.5 g,NaCI2.8 g,KCI 0.15 g,
Na2HP04 0.07 g, NaHC03 0.05 g, glucose 0.5 g, trehalose 0.5 g, galactose 0.5 g, phenol red 5.0 mg; trypsin (2x crystallized; Sigma) 2.4 g,
EDTA tetrasodium salt (MW = 380) 38.0 mg in 1,000 ml H20)
- Culture media (Basch and Diconza's medium C; see Appendix 1)
- Hansen's M-182, M260 or S-301 media (see Appendix 1)
- Antibiotic solution (penicillin 100 IV/ml, streptomycin 100 flg/ml,
Fungizone 0.25 flg/ml, chlortetracycline 5 flg/ml)
- Lettuce
Miscellaneous
- Rat diet granules (Geisler Pet Products)
- Eggs of snails Biomphalaria glabrata
Animals
Procedure
The following procedure is based on the techniques of Hansen (1979).
1. Maintain the snails on lettuce and rat diet granules.
Oviposition
2. Allow oviposition on plastic rafts overnight.
3. Remove egg sacs carefully (eggs are deposited in groups in gelatinous
sacs).
1. Rinse the egg sacs briefly in 0.2% iodine.
Sterilization
2. Rinse the egg sacs briefly in 0.001 % hyamine 1622.
3. Incubate the egg sacs at 23°C for 4-5 days in a sterile solution of antibiotics in a petri dish.
4. Change the antibiotic solution daily.
5. Select embryos at the trochophore stage.
6. Place approximately 15 egg sacs (containing more than 40 embryos)
in a BPI dish containing 0.4 ml BPSA.
7. Tease the egg sacs apart and release the embryos from the egg capsules.
8. Rinse the embryos by transferring them to another BPI dish containing BPSA.
9. Repeat this rinsing procedure three more times.