Histology FAQ
Staining, Histochemistry and Histotechnology
(Frequently Asked Questions)
Dr. John A. Kiernan
Department of Anatomy
and Cell Biology
The University of Western Ontario
London, Canada
Stain for
Chlamydia (Castaneda's method)
Question.
How do you carry out the Castaneda stain for Chlamydia?
Answer.
Castaneda's stain for elementary bodies and Rikettsiae (1930)
Castaneda's staining solution
Solution A
Potassium dihydrogen phosphate, anhydrous
1 g
Disodium hydrogen phosphate
25 g
Distilled water
1000 ml
Formalin
1 ml
Dissolve the potassium dihydrogen phosphate
in 100 ml distilled water and the disodium hydrogen phosphate
in 900 ml distilled water. Mix the two solutions to give a
buffer pH 7.5, and add formaldehyde as a preservative.
Solution B
Methylene blue 1 g
Methanol
100 ml
Staining solution
Solution A 20
ml
Solution B 0.15 ml
Formalin
1 ml
Safranine-acetic acid
Safranine (0.2% aqueous solution)
1 part
Acetic acid (0.1% aqueous solution)
3 parts
Procedure.
* Prepare films from infected tissue and dry in air
* Apply the stain for 3 min.
* Drain, do not wash
* Counterstain for a 1-2 seconds in safranine-acetic acid
* Wash in running water, blot dry.
Rickettsiae, elementary bodies of psittacosis: blue. Cell nuclei and cytoplasm: red.
Reference: "Biological stains and staining methods." BDH leaflet, 1966.
Several modifications of Castaneda's original technique are given in: Langeron, M.:"Precis de Microscopie", 1934 and 1948.
Yvan Lindekens
(yvan.lindekens[AT]rug.ac.be)