Appendix 8 - Fixatives and Solutions
- Kahle's Fixative (modified)
- Ethanol (100%) - 15 ml
- Formaldehyde (40%) - 6 ml
- Glacial acetic acid - 1 ml
- Seawater - 31 ml
- Natural filtered seawater
Natural flowing seawater can be filtered by one or two simple filtering devices arranged in tandem. Attach the inlet seawater spout to a glass tube mounted in a one hole stopper fitted on the end of an 18 inch plastic pipe (3 in diameter) which is loosely packed with glass wool (angel hair). The wool will need periodic changing. Place a similar tube and stopper at the outlet. The latter flow can be further filtered by leading it to an inverted aspirator jar 1/3 filled with clean sea sand and plugged at the bottom by a wad of glass wool and a lower tube outlet.
- Membrane filtered seawater
Prefiltered seawater (above) can be membrane filtered-sterilized by passage through a 0.2 µm polycarbonate filter and refrigerated in glass stoppered bottles. It can be used for dilution of isotopes, preparation of maturation factor, etc.
- Artificial Sea Water (Moore's Calcium Free; Cavanaugh, 1956)
- NaCl - 25.48 g/l
- KCl - 0.72 g/l
- MgCl2.6H2O - 6.94 g/l
- MgSO4.7H2O - 4.ll g/l
- Methyl-benzoate/celloidin solution
Add 1 g of celloidin (dry) to 100ml of methyl benzoate. Shake and repeat process until dissolved (up to 24 hours).
- Jenner-Giemsa stain (Humason, 1962)
A. Stock Jenner's dye: 1 g dye in 400 ml methyl alcohol
- Working solution: 25ml stock dye + 25 ml distilled water
B. Stock Giemsa dye: 1 g dye in 66 ml methyl alcohol + 66 ml glycerin
- Working solution: 2.5ml stock dye + 50 ml distilled water
- Must be prepared fresh!
Deparaffinize sections and pass to 50% methyl alcohol. Stain in Jenner's working solution 6 min. Pass to Giemsa working solution 45 min. Rinse briefly in distilled water and dehydrate quickly in 95% ethyl alcohol, into 100% ethyl alcohol, clear and mount. This procedure is compatible with staining through autoradiographic dipping films.