Iron-Alum Aceto-Carmine Staining for Chromosomes and Other Anatomical Features of Rhodophyceae

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Abstract
The chromosomes, certain intracellular structures and gross anatomical details of many red algae, which, as a class, have proved technically difficult material, can be demonstrated by staining with aceto-carmine after a mordant bath of iron alum. Acetic-alcohol mixtures are used as nuclear fixatives and formalin-acetic-alcohol and other similar fluids for preservation of anatomical features. The tougher more cartilaginous thalli of some species can be softened, if squashes are desired, by prolonging fixation (24-48 hr.) in acetic alcohol and subsequent washing. The fixatives are washed out of the material before the latter is transferred to 0.5-5.0% ferric ammonium sulphate, the concentration of which may be altered according to the material. Excess mordant is removed by washing and the material stained in Belling's aceto-carmine containing a trace of ferric acetate as a “ripener”. The degree of heating before covering is critical as it controls the quality of the staining. Squashing must be very thorough to spread the chromosomes which are usually very small but only slight controlled pressure is necessary when diffuse structures such as carposporophytes, nemathecia or medullary filaments are being demonstrated. Paraffin sections mounted on slides can also be stained by this method.