MODIFIED FRAZER-LENDRUM METHOD FOR FIBRIN

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Image Examples:

FIXATION:                10% buffered neutral formalin

TECHNIQUE:                Paraffin sections cut at 5 μm.

SOLUTIONS:

Zenker’s Solution, Modified

Zinc acetate --------------------------------------------------------------------                3.0 gm

Potassium dichromate ----------------------------------------------------------                7.5 gm

Distilled water -----------------------------------------------------------------                300.0 gm

Acetic acid, glacial -------------------------------------------------------------                15.0 ml

       Dissolve the zinc acetate and potassium dichromate in the distilled

       water and then add the acetic acid.

Modified Weigert’s Iron Hematoxylin

(See Gomori’s One Step Trichrome Method)

0.5% Acid Alcohol Solution

(See Truant’s Method for Acid Fast Bacilli)

Orange G-Picric Acid Solution

Picric acid, saturated aqueous -------------------------------------------------                100.0 ml

Orange G, C.I. 16230 -----------------------------------------------------------        1.0 gm

1% Acid Fuchsin Solution

Acid fuchsin, C.I. 42685 -------------------------------------------------------                1.0 gm

Distilled water ------------------------------------------------------------------                100.0 ml

Glacial acetic acid ---------------------------------------------------------------                1.0 ml

Tartarazine Differentiating

Tartarazine, C.I. 19140 --------------------------------------------------------                1.0 gm

Picric acid, saturated aqueous -------------------------------------------------                100.0 ml

0.2% Light Green Solution

Light green, SF yellowish, C.I. 42095 ------------------------------------------                0.2 gm

Distilled water ------------------------------------------------------------------                100.0 ml

Glacial acetic acid ---------------------------------------------------------------                1.0 ml

• Use control slide.

1. Deparaffinize and hydrate to distilled water.
2. Place in Zenker’s prepared with zinc acetate overnight at room temperature.  An alternate method is to place in 40 ml. of Zenker’s solution in a glass Coplin jar and microwave at power level 1 (60W) for 1 minute.  Transfer to a 37o C oven for 3 hours.
3. Wash in running water for 3 minutes and rinse in two changes of distilled water.
4. Modified Weigert’s iron hematoxylin for 5 minutes.
5. Wash in running water for 1 minute and rinse in distilled water.
6. Place in acid alcohol for 5 seconds.
7. Wash in running water for 1 minute and rinse in distilled water.
8. Orange G-picric acid solution for 5 minutes.
9. Rinse in 3 changes of distilled water.
10. Acid fuchsin solution for 5 minutes.
11. Rinse in 3 changes of distilled water.
12. Tartarazine differentiating solution for 2 minutes.
13. Rinse with 3 changes of distilled water.
14. Light green solution for 15 seconds.
15. Rinse in 3 changes in distilled water.
16. Dehydrate in graded alcohols.
17. Clear in xylene, three or four changes.
18. Mount with synthetic resin.

Fibrin, keratin, some cytoplasmic granules ------------------------------------------                red

Erythrocytes -----------------------------------------------------------------------                orange

Muscle ----------------------------------------------------------------------------                may stain red

Collagen ---------------------------------------------------------------------------                green

We have found this staining method to be useful in demonstrating fibrin.  More consistent staining of fibrin, and better contrast with background is obtained than with the Mallory PTAH method.

The use of Zenker’s prepared with zinc acetate does not produce the precipitate that results when mercuric chloride is used to prepare Zenker’s solution.  This eliminates the necessity of treating the tissue sections with Lugol’s iodine and sodium thiosulfate.

This modified method greatly reduces the amount of picric acid needed to prepare the solutions that require picric acid.  The staining results are as good as, if not better, than the original Frazer-Lendrum technique.

Lendrum, A.C., et al.:  Studies on the character and staining of fibrin.  J. Clin. Path. 15:401-413, 1962.

Luna, L.:  Manual of Histologic Staining Methods of the Armed Forces Institute of Pathology, 3rd Ed., New York, McGraw-Hill, 1968, pp. 81-82.