270
Organ Culture
• • Results
According to Daig and Spindler (1979), the integument can be kept alive
for several days. During this period, the cultured integument synthesized protein, RNA and DNA. Ecdysone added to the culture medium
was converted to 20-hydroxyecdysone by the integument.
Subprotocol 2
Pre-Exuvial Integument of Crabs (Crustacea)
• • Materials
Equipment - Incubator
- Aquarium recirculating salt water
- Culture tubes with caps (15 ml)
- Pasteur pipettes
- Petri dishes
- Tissue culture dishes (no. 3001; Falcon; Becton-Dickinson)
Solutions - Crab integument medium I and II (see Appendix 1)
- Salt water
- 20% Betadyne (= povidone iodine; Sigma) solution
Animals - Blue crab Callinectes sapidus
Preparation
of crabs
Sterilization
• • Procedure
1. Maintain animals in a recirculating salt water aquarium at room temperature.
2. Select crabs in the DJ stage according to the methods of Drach and
Tchernigovtzeff (1967) and Stevenson (1968).
1. Immerse the animals in ice-cold 20% Betadyne solution for 5 min to
surface sterilize and anesthetize them.
2. Blot dry the crabs, swab the carapace with 70% ethanol, and air-dry
the crabs in a laminar flow cabinet.
Fig. 1. Crab outer morphology. 1, frontal
area; 2, hepatic area, 3, branchial area; 4, cardiac area; 5, intestinal area; 6, coxopodite; 7,
ischiopodite; 8, meropodite; 9, carpopodite;
10, propodite; 11, dactylopodite
Organ Culture
• • Results
According to Daig and Spindler (1979), the integument can be kept alive
for several days. During this period, the cultured integument synthesized protein, RNA and DNA. Ecdysone added to the culture medium
was converted to 20-hydroxyecdysone by the integument.
Subprotocol 2
Pre-Exuvial Integument of Crabs (Crustacea)
• • Materials
Equipment - Incubator
- Aquarium recirculating salt water
- Culture tubes with caps (15 ml)
- Pasteur pipettes
- Petri dishes
- Tissue culture dishes (no. 3001; Falcon; Becton-Dickinson)
Solutions - Crab integument medium I and II (see Appendix 1)
- Salt water
- 20% Betadyne (= povidone iodine; Sigma) solution
Animals - Blue crab Callinectes sapidus
Preparation
of crabs
Sterilization
• • Procedure
1. Maintain animals in a recirculating salt water aquarium at room temperature.
2. Select crabs in the DJ stage according to the methods of Drach and
Tchernigovtzeff (1967) and Stevenson (1968).
1. Immerse the animals in ice-cold 20% Betadyne solution for 5 min to
surface sterilize and anesthetize them.
2. Blot dry the crabs, swab the carapace with 70% ethanol, and air-dry
the crabs in a laminar flow cabinet.
Fig. 1. Crab outer morphology. 1, frontal
area; 2, hepatic area, 3, branchial area; 4, cardiac area; 5, intestinal area; 6, coxopodite; 7,
ischiopodite; 8, meropodite; 9, carpopodite;
10, propodite; 11, dactylopodite