5 Insecta (Lepidoptera)
69
Subprotocol 13
Neonate Larvae
The first ins tar larvae just after hatching are used like embryos. Neonate
larvae, as a whole, are used as the starting material for primary culture .
• • Materials
The materials used are the same as those given for subprotocol12 .
• 11 Procedure
1. Follow steps 1 and 2 for sterilization as given in subprotocol 12.
2. Transfer the surface-sterilized eggs into a sterile tube with a small
amount of culture medium (Fig. 22).
3. Incubate the tube at 25°C until the eggs hatch.
1. Catch hatched larvae and dunk them in culture medium placed on a
Maximov slide or in a small petri dish (3-5 cm diameter).
2. Cut the larvae into several pieces with a pair of scissors or a knife.
3. Transfer the fragments, with the culture medium, into culture vessels.
4. Culture at 25°C.
Fig. 22. An apparatus for the
preparation of sterile neonate larvae (Sato, 1989). Reproduced with permission
from Invertebrate Cell System Applications. Copyright CRC Press, Boca
Raton, Florida, USA
Fig. 23. Cell migration from an explant (EX) of neonate larvae of the
Tortricid Homona magnanima.
(Sato, 1989) x120. Reproduced
with permission from Invertebrate Cell System Applications.
Copyright CRC Press, Boca Raton,
Florida, USA
Production
of sterile
neonate
larvae
Culture
set-up
69
Subprotocol 13
Neonate Larvae
The first ins tar larvae just after hatching are used like embryos. Neonate
larvae, as a whole, are used as the starting material for primary culture .
• • Materials
The materials used are the same as those given for subprotocol12 .
• 11 Procedure
1. Follow steps 1 and 2 for sterilization as given in subprotocol 12.
2. Transfer the surface-sterilized eggs into a sterile tube with a small
amount of culture medium (Fig. 22).
3. Incubate the tube at 25°C until the eggs hatch.
1. Catch hatched larvae and dunk them in culture medium placed on a
Maximov slide or in a small petri dish (3-5 cm diameter).
2. Cut the larvae into several pieces with a pair of scissors or a knife.
3. Transfer the fragments, with the culture medium, into culture vessels.
4. Culture at 25°C.
Fig. 22. An apparatus for the
preparation of sterile neonate larvae (Sato, 1989). Reproduced with permission
from Invertebrate Cell System Applications. Copyright CRC Press, Boca
Raton, Florida, USA
Fig. 23. Cell migration from an explant (EX) of neonate larvae of the
Tortricid Homona magnanima.
(Sato, 1989) x120. Reproduced
with permission from Invertebrate Cell System Applications.
Copyright CRC Press, Boca Raton,
Florida, USA
Production
of sterile
neonate
larvae
Culture
set-up