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Methods for Setting Up Primary Cultures Specific to Animal Groups
5. Place the capillary tubes containing the hemolymph in 30-ml culture
flasks containing RML-375 culture medium.
6. Culture at 28°C.
Results
According to Moulton (1978), hemocyte cultures grow best when cells
are left in the capillary tubes. When hemocytes are first explanted, they
attach, become spindle shaped, and develop small unipolar or bipolar
cytoplasmic processes. The cells remain like this for approximately two
weeks (Fig.4). The cells showed phagocytotic activity. Some cultures survived for 9 months.
Subprotocol 5
Tick (Arachnida) Preimaginal Bodies
• • Materials
Equipment - Dissecting microscope
- Centrifuge and centrifuge tube (15 ml)
- Petri dishes (60 mm diameter)
- Iris scissors
- Forceps
- Pasteur pipettes
Solution - Culture medium (RML-375; see Appendix 1)
An ima Is - Nymphal ticks Rhipicephalus sanguineus, Dermacentor andersoni, and
Amblyomma macula tum
Others - Roccal (a quaternary ammonium compound; Winthrop Laboratories)
- Quick-drying glue
Procedure
The following procedure is based on the methods of Moulton (1978).
Sterilization 1. Surface sterilize the ticks by submersion in 10% Roccal for 30 min.
2. Wash the ticks in two changes of 70% ethanol.
3. Rinse the ticks in several changes of sterile distilled water.
Dissection 1. Dry and glue the tick, ventral side down, in the middle of a 60-mm
petri dish with quick -drying glue (the nozzle of the tube of glue should
be disinfected with UV irradiation before squeezing out the glue).
Methods for Setting Up Primary Cultures Specific to Animal Groups
5. Place the capillary tubes containing the hemolymph in 30-ml culture
flasks containing RML-375 culture medium.
6. Culture at 28°C.
Results
According to Moulton (1978), hemocyte cultures grow best when cells
are left in the capillary tubes. When hemocytes are first explanted, they
attach, become spindle shaped, and develop small unipolar or bipolar
cytoplasmic processes. The cells remain like this for approximately two
weeks (Fig.4). The cells showed phagocytotic activity. Some cultures survived for 9 months.
Subprotocol 5
Tick (Arachnida) Preimaginal Bodies
• • Materials
Equipment - Dissecting microscope
- Centrifuge and centrifuge tube (15 ml)
- Petri dishes (60 mm diameter)
- Iris scissors
- Forceps
- Pasteur pipettes
Solution - Culture medium (RML-375; see Appendix 1)
An ima Is - Nymphal ticks Rhipicephalus sanguineus, Dermacentor andersoni, and
Amblyomma macula tum
Others - Roccal (a quaternary ammonium compound; Winthrop Laboratories)
- Quick-drying glue
Procedure
The following procedure is based on the methods of Moulton (1978).
Sterilization 1. Surface sterilize the ticks by submersion in 10% Roccal for 30 min.
2. Wash the ticks in two changes of 70% ethanol.
3. Rinse the ticks in several changes of sterile distilled water.
Dissection 1. Dry and glue the tick, ventral side down, in the middle of a 60-mm
petri dish with quick -drying glue (the nozzle of the tube of glue should
be disinfected with UV irradiation before squeezing out the glue).