ACETOCARMINE STAINING
Acetocarmine preparation (1% solution)
Carmine is a basic dye that is prepared from the insect Coccus cacti. Dissolve 10 g carmine (Fisher C579-25) in 1 L of 45% glacial acetic acid, add boileezers, and reflux for 24 h. Filter into dark bottles and store at 4°C. This solution can be stored for a long time. Staining can be intensified by adding ferric chloride (FeCl2·6H2O); add 5 mL of a 10 % ferric chloride solution per 100 mL of % acetocarmine.
Acetocarmine staining
To stain plant chromosomes, a 1% solution of carmine in 45% acetic acid is used. Freshly fixed material is transferred into 1% acetocarmine for at least 30 min and then analyzed by the squash method. If the material was fixed for a longer time, it requires a longer staining time (up to several days) to reach good contrast. If the material is to be analyzed immediately, fix and stain the tissue in one step using the 1% acetocarmine solution.
Chromosome squash technique
Drain off the fixative and place the roots in 1% acetocarmine for 1 to 3 h. Heat until the acetocarmine begins to boil. Cut off the root cap with a razor blade and squeeze the meristematic tissue out with a lancet needle. Add a drop of acetocarmine or 45% acetic acid. Place a razor blade (double-edged) to one side and add a cover slip. Tap the cover slip gently with the needle end of a probe. Slide the razor blade out and heat to a point just below boiling (steam will form beneath the slide). Then, quickly squash with thumb or forefinger between two layers of filter paper. Be careful to not move the cover slip at this point.
