Recipes for Voucher Preservation
PVLG is used to permanently mount whole or broken spores on glass slides. For best results, mounted specimens should not be studied for 2-3 days after they were mounted to give time for spore contents to clear. Whole spores will change color, generally darkening to varying degrees, and shrink or collapse with plasmolysis of spore contents. Discrete layers of the spore wall or flexible inner walls of broken spores will swell to varying degrees and appear fused after long storage in some instances.
|Distilled water||100 ml|
|Lactic acid||100 ml|
|Polyvinyl alcohol (PVA)||16.6 g|
It is most important to mix all ingredients in a dark bottle BEFORE adding the polyvinyl alcohol. The PVA should have the following properties: 99-100% hydrolyzed, and a viscosity of 24 – 32 centipoise in a 4% aqueous solution at 20°C. The PVA is added as a powder to the other liquid ingredients. The PVA dissolves slowly, and then only when placed in a hot water bath (70 – 80°C). The solution will be clear in 4-6 hours; we usually prepare the mix in the evening and letting it incubate in the water bath overnight. PVLG stores well in dark bottles for approximately one year.
Rick Koske writes that the PVA powder can be added to the water, followed by autoclaving for 15 minutes. The lactic acid and glycerin are added, and the solution then is stored at room temperature for at least 24 hr before using.
Melzer’s reagent has become an important diagnostic tool in morphology-based diagnoses. Iodine staining reactions will vary from pale pink (weak reaction) to dark red-brown (moderate reaction) to dark reddish-purple (intense dextrinoid reaction). The iodine binds to hydrophobic regions of macromolecules (in spore or germinal walls) and intensity of the reaction is related in part to the length of carbohydrate chains. In most instances to date, the intensity of the reaction is directly correlated to plasticity of the structure in acidic mountants.
|Chloral hydrate||100 g|
|Distilled water||100 ml|
|Potassium iodide||5.0 g|
When Melzer’s reagent is used according to the recipe above, staining reactions will be most intense. However, mounts are temporary even when a coverslip is sealed, and often dries out within 1-2 years of storage. More permanent mounts are made by mixing Melzer’s reagent with PVLG in a volume ratio of 1:1 (and storing the mixture in a dark bottle). The staining reaction is diminished slightly, but not enough to cause any confusion as to the intensity of the reaction. In structures staining weakly, the color reaction fades within a year or two of storage.