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Product for the preparation of cyto-histological samples for optical microscopy.
Recommended method to show argyrophilic reticular fibres in connective tissue.
This method produces a selective evident impregnation in a very short time thanks to two factors: the preliminary impregnation
with an iron salt and the use as silver source of an unstable diaminic complex (ammoniacal solution), which is more reactive than silver nitrate.
1) Pre-treatment with trivalent iron.
After a preparatory oxidation with potassium permanganate, the section is treated with trivalent iron (ferric ammonium
sulphate). Iron ions, more reactive than silver ions, quickly bind affine functional groups in argyrophilic structures.
2) Treatment with ammoniacal solution.
Silver is present in ammoniacal solution in the form of complex hydrosoluble oxide – [Ag(NH3)2]2 O. This complex silver cation
replaces iron previously bound to tissues. In the next step, formic aldehyde acts as reducing agent: it removes oxygen from the complex and releases metallic silver that deposits on argyrophilic structures.
[Ag(NH3)2]2 O + HCHO = 2 Ag + 4 NH3 + HCOOH Any excess silver in the unprecipitated state is removed by treating with gold chloride Unreduced silverdiamine cation is then removed by sodium thiosulfate (Na2S2O3). Both form a complex, which is highly soluble but cannot be oxidized any more. A nuclear red staining completes the method.
For good results, follow these rules:
– Always use excellent and chlorine-free distilled or deionized water.
– Use only perfectly clean glassware.
– Avoid deposit of dust on sections. Never touch solutions with metallic objects (tweezers etc.)
1) Bring the section to distilled water.
2) Put on the section 5 drops of reagent A and 5 drops of reagent B: leave to act 5 minutes.
3) Wash the slide in distilled water.
4) Put on the section 10 drops of reagent C: leave to act 1 minute.
5) Double washing in distilled water.
6) Put on the section 10 drops of reagent D: leave to act 3 minutes.
7) Double washing in distilled water.
8) Put on the section 10 drops of reagent E: leave to act 3 minutes.
9) Wash in distilled water.
10)Put on the section 10 drops of reagent F: leave to act 5 minutes.
11)Double washing in distilled water.
12)Put on the section 10 drops of reagent G: leave to act 2 minutes
13)Wash in distilled water.
14)Put on the section 10 drops of reagent H: leave to act 2 minutes
15)Wash in distilled water
16)Put on the section 10 drops of reagent I: leave to act 5 minutes
17)Dehydrate through ascending alcohols: clear in xylene and mount.