5 Insecta (Lepidoptera)
59
~. Cut the heart into pieces smaller than 1 mm.
4. Transfer the fragments, with the culture medium, into culture vessels.
5. Culture at 25°C.
Results
Some cells may migrate from the explants and survive for more than 1
month (Fig. 10). It is difficult to determine the exact origin of the migrated cells. It is also difficult to obtain a continuous cell line from a
primary culture of circulatory tissues (Miltenburger et aI., 1977).
Subprotocol 8
Hemocytes
Hemocytes are freely circulating cells in hemocoeI. They are cultured as
a free cell culture.
Materials
- Low-speed centrifuge and centrifuge tubes
- Pasteur pipettes
- Culture dish
- Culture vessels
- Carlson's solution (see Appendix 1)
- Culture media
- Larval, pupal, or imaginal lepidopteran insects
Procedure
Equipment
Solutions
Animals
1. Surface sterilize insects by submersion in 70% ethanol for 5 min. The Sterilization
precautions given in step 1 of the sterilization procedure in subprotocol
1 should be undertaken.
1. Hold the larvae dorsally bent back (Fig. 11), and make an incision Bleeding
with scissors or a puncture with a pin on a proleg. In the case of pupae, make an incision on the head, while for adults, cut a leg. Only a
small amount of blood can be obtained from adult insects.
2. Gently press the body of the insect.
3. Drop the blood that oozes out onto Carlson's solution placed in a culture dish.