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Organ Culture
Results
According to Fyhn and Costlow (1975), ovarian cells do not change very
much after 7 days culture. Small immature oocytes adjacent to the oogonia had a normal appearance. The oogonia seemed normal, but did not
undergo mitosis. Parenchymatous cells located between the ovarioles did
not exhibit mitosis, but kept a normal appearance, even after 25 days
culture.
Cement glands in explants had a normal appearance after 7 days culture.
Subprotocol 7
Ovaries of Crabs (Crustacea)
Materials
Equipment - Dissecting microscope and dissecting tray
- Culture dish
- Pasteur pipettes
- Petri dishes
- Watch glasses
Solutions - Stock culture medium (dissolve lyophilized TC-199 medium without
serum in 100 ml artificial seawater (ASW) or saline)
Animals - Portunid crab Thalamita crenata
Miscellaneous Lens paper prepared according to the method of Chen (1954):
- Cut the lens paper into an octagon with a diameter of approximately
25mm
- Wash the lens paper in two or three changes of ether for 2 h
- Dip the lens paper in two or three changes of absolute alcohol for
2h
- Keep the lens paper in glass-distilled water overnight with five to
10 changes of water
- Dry the paper in an oven at 70°-80°C
- Sterilize the lens paper by dry heat at 120°-150°C for 1 h
Procedure
Sterilization 1. Swab a crab with 95% ethanol.
Dissection 1. Remove the carapace from the crab.
2. Dissect away the hypodermis and other supporting tissues under a
dissecting microscope.
3. Cut the anterolateral portions of the ovaries into 1-2-mm 3 pieces, and
wash the pieces in stock culture medium.