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See also: Tissue Stains: 49,
51, 52,
55, 56,
72, 81.
PRINCIPLE OF THE TEST.
To stain paraffinized tissue sections for amyloid, required in
the diagnosis of some diseases encountered in anatomic pathology,
such as amyloidosis.
SPECIMEN REQUIRED.
All human tissue excised at surgery, outpatient clinics, and postmortems,
fresh or in fixative, along with a filled-out Tissue Examination Form
(U. S. Standard Form 515, USSF515).
REAGENTS, INSTRUMENTATION.
10% buffered neutral formalin.
Paraffin sections at 6 microns.
1% Congo red solution.
Congo red powder.
1% Sodium hydroxide solution.
Sodium hydroxide.
Alkaline alcohol solution.
50% Ethanol.
Mayer's hematoxylin solution.
Alkaline alcohol solution.
95% alcohol.
Absolute alcohol.
Clearing.
Permount.
STEP-BY-STEP DESCRIPTION.
FIXATION: 10% buffered neutral formalin.
TECHNIQUE: Cut paraffin sections at 6 micron (if brighter color is desired.)
Cut at 12 microns - useful for minute amyloid deposits.
SOLUTIONS: 1% CONGO RED SOLUTION
Congo red.......................................... 1 gm
Distilled water.................................... 100 ml
1% SODIUM HYDROXIDE SOLUTION
Sodium hydroxide solution........................... 1 gm
Distilled water..................................... 100 ml
ALKALINE ALCOHOL SOLUTION
Sodium hydroxide, 1%................................ 1 ml
Alcohol, 50%........................................ 100 ml
MAYER'S HEMATOXYLIN SOLUTION
STAINING PROCEDURE.
1. Deparaffinize and hydrate to distilled water.
2. Congo red solution for 1 hour.
3. Rinse off excess stain in water, two or three changes.
4. Differentiate in alkaline alcohol solution for 3 - 5 seconds. Agitate
constantly until the background appears clear.
5. Wash in running water for 5 minutes.
6. Counterstain in Mayer's hematoxylin solution for 5 minutes.
7. Wash in running water for 15 minutes.
8. Dehydrate in 95% alcohol, absolute alcohol and clear (two changes each).
9. Mount.
CONTROL: Amyloidosis of skin
RESULTS:
Amyloid: pink to red.
Nuclei: blue.
REFERENCES.
1. Vowles GH, Francis RJ.
Amyloid. Chapter 15, pp. 303-324.
In: Bancroft JD, Gamble M.
Theory and Practice of Histological Techniques. Fifth Edition.
Edinburgh: Churchill Livingstone. 2002;:303-324.
ISBN 0-443-06435-0, 796 pages.
2. Bennhold H.
Eine spezifische Amyloid-Färbung mit Kongorot.
[German: A specific amyloid stain with Congo red].
Münchener Med Wochenschr. 1922;33:1537-1541.
3. Chiari H.
Ein Beitrag zur sekundären fluoreszenz des lokalen Amyloids.
[German: A contribution to secondary flourescence in local amyloid].
Mikroskopie. 1947;2:79.
4. Divery P, Florkin M.
Sur les propriétés optiques de l'amyloide.
[French: Regarding the optical properties of amyloid].
Compte Rendus des Séances de la Societé de Biologie
et des Filiales. 1927;97:1808-1810
5. Friedrich N, Kékulé.
Zur Amyloidfrage. [German: On the question of amyloid.]
Virchows Arch f patholog Anat u Physiol u f klin Medizin. 1859;16:50.
6. Highman B.
Improved methods for demonstrating amyloid in paraffin sections.
Arch Pathol. 1946;41:559.
7. Reimann HA, Koucky RF, et al.
Primary amyloidosis limited to tissue of mesodermal origin.
Am J Pathol. 1935;11:977.
8. Schwartz P.
Amyloidosis.
Illinois: C.C. Thomas. 1970;:.
9. Virchow R.
Weitere Mittheilungen über das Vorkommen der pflanzlichen Cellulose
beim Menschen. [German: Further communications regarding the appearance
of plant cellulose in humans].
Virchows Arch f patholog Anat u Physiol u f klin Medizin. 1853:6:246.
10. Prophet EB, Mills B, Arrington JB, Sobin LH, eds.
Laboratory Methods in Histotechnology.
Washington, DC: Armed Forces Institute of Pathology. 1992;:.
ISBN 1-881041-00-X, 278 pages.
11. Mikel UV, ed.
Advanced Laboratory Methods in Histology and Pathology.
Washington, DC: Armed Forces Institute of Pathology. 1994;:.
ISBN 1-881041-13-1, 254 pages.