Wilder Modification of the Bielschowsky Stain for Reticulum
EMS Catalog #: 26392-Series
Cut paraffin at 6µm sections.
- Deparaffinize and hydrate slides to distilled water.
- Oxidize in Phosphomolybdic Acid, 10% (#26392-07) for 1 minute. Rinse well in distilled water or sunning water to prevent the cells from holding yellow colors.
- Sensitize in Uranyl Nitrate, 1% (#26392-08) for 1 minute or less. Rinse 10 – 20 seconds in distilled water.
- Stain in fresh Ammonical Silver for one minute (changing solution frequently). The solution may be prepared as follows:
- Add concentrate Ammonium Hydroxide (#26392-02) to 5.0 ml of Silver Nitrate, 10.2% (#26392-01) drop by drop, until the forming precipitate is almost dissolved.
- Add 5.0 ml of Sodium Hydroxide, 3.1% (#26392-03) and just dissolve the resultant precipitate by slowly adding conc. Ammonium Hydroxide dropwise. Bring volume up to 50 ml with distilled water.
- Use at once or filter and store in refrigerator in amber bottle.
- Dip quickly in 95% alcohol and transfer to freshly prepared reducing solution. Prepare the reducing solution as follows:
- Mix 50.0 ml distilled water, 1.5ml Uranyl Nitrate, 1% (#26392-08) and 0.5 ml of Formalin, 40% neutralized (#26392-04).
- Prepare just before use and change frequently during the process.
- Rinse in distilled water.
- Tone in Gold Chloride, 0.2% (#26392-09) approx. 1 minute, until the yellow brown color leaves the sections and they turn lavender. Check the slides at this point individually with a microscope to prevent over toning; this could lead to red sections.
- Rinse in distilled water and transfer to Sodium Thiosulfate, 5% (#26392-10) – approx. 1 minute – to fix the color. Wash in tap water.
- Counter-stain in Nuclear Fast Red (#26392-06) if desired, rinsing well in distilled water. Alternatively Harris Hematoxylin (#26392-05) with either Van Gieson (#26392-5A) or phloxine can be used as a counterstain.
- Dehydrate in 95% and 100% alcohol, 2 changes and clear in xylene, 2 changes.
- Mount. If celloidin is used, clear in oil of origanium followed by 25% alcohol, usually mounted in balsam with Permount.
|Reticulum fiber (If beading is observed and/or dark background, replace the ammonium Hydroxide Silver Nitrate solution and the reducing solution||Black|
|Other tissue elements||Red|
Wilder, H.C., Amer. J. Clin. Path., 11:817, 1935.
Mallory, F.B. Pathological Technique, Phil., W.B. Saunders & Co, c. 1938
AFIP Manual of Histological Staining Techniques, NY, McGraw-Hill Publ., c. 1968/3rd ed. P.92
Wilder Modification of the Bielschowsky Stain for Reticulum is available online from the EMS Catalog. For ordering or product information, click here.