Staining Procedure

1. Deparaffinize sections if necessary and hydrate to distilled water.

Pneumocystis Stain Kit 2. Place slide in freshly mixed Sulfation Reagent for 10 minutes.
Note: Agitate Staining Jar every few minutes to keep acids mixed.
Catalog Number: KT030 3. Rinse in several changes of distilled water.
4. Incubate slide in Cresyl Echt Violet Solution (0.1%) for 10-15 minutes.
Note: Agitate slide several times during incubation step.
5. Rinse quickly in distilled water.
Document #: DS-3023-A 6. Dip slide in Naphthol Yellow S Solution for 4 seconds.
Effective Date: 02/015/15 Note: Excessive Naphthol Yellow S Solution decolorizes
Pneumocystis.
Intended Use 7. Rinse very quickly in distilled water.
For In Vitro Diagnostic Use 8. Dehydrate very quickly in 2 changes of fresh Absolute Alcohol.
Alternative Method: Dip slide twice in Absolute Alcohol and air-dry
Summary and Explanation slide.
The Pneumocystis Stain Kit is intended for use in the histological visualization of 9. Clear, and mount in synthetic resin.
Pneumocystis carinii in cytology smears, and paraffin or frozen tissue sections.
Limitations of the Procedure
Pneumocystis carinii: Violet / Purple 1. Histological staining is a multiple step diagnostic process that
Connective Tissue: Blue / Green requires specialized training in the selection of the appropriate
Erythrocytes: Yellow reagents, tissue selections, fixation, processing, preparation of the
Mucin: Rose / Purple slide, and interpretation of the staining results.
Cartilage: Rose / Purple 2. Tissue staining is dependent on the handling and processing of the
tissue prior to staining.
Control Tissue 3. Improper fixation, freezing, thawing, washing, drying, heating,
Tissues fixed in 10% formalin are suitable for use prior to paraffin embedding. sectioning, or contamination with other tissues or fluids may
Consult references (Kiernan, 1981: Sheehan & Hrapchak, 1980) for further produce artifacts or false negative results.
details on specimen preparation. 4. The clinical interpretation of any positive staining, or its absence,
1. Cut sections, usually 3 to 5 µm and pick the sections up on glass must be evaluated within the context of clinical history, morphology
slides. and other histopathological criteria. It is the responsibility of a
2. Bake the slides for at least 30 minutes at approximately 70°C. qualified pathologist to be familiar with the special stain and
3. Allow to cool. methods used to produce the slide.
5. Staining must be performed in a certified licensed laboratory under
Recommended Positive Control the supervision of a pathologist who is responsible for reviewing the
1. Pnuemocystis carinii stained slides and assuring the adequacy of positive and negative
controls.
Reagents Provided
Kit Contents Volume Storage Precautions
Cresyl Echt Violet Solution (0.1%) 125 mL 2-8°C 1. Consult local and/or state authorities with regard to recommended
Naphthol Yellow S Solution 125 mL 15-30°C method of disposal.
Staining Jar 60 mL N/A 2. Materials of human or animal origin should be handled as
biohazardous materials and disposed of with proper precautions.
Storage and Handling 3. Avoid microbial contamination of reagents. Contamination could
Do not use product after the expiration date printed on vial. If reagents are produce erroneous results.
stored under conditions other than those specified here, they must be verified 4. This reagent may cause irritation. Avoid contact with eyes and
by the user. Diluted reagents should be used promptly. mucous membranes.
5. If reagent contacts these areas, rinse with copious amounts of water.
Prepare Of Sulfation Reagent Immediately Before Use (Not Included in Kit): 6. Do not ingest or inhale any reagents.
1. Pour 15 ml of Glacial Acetic Acid into the Staining Jar provided with
this kit. Troubleshooting
2. Slowly add 5 ml of Sulfuric Acid to the Staining Jar. If unexpected staining is observed which cannot be explained by variations in
3. Screw cap tightly on staining jar and invert several times to laboratory procedures and a problem is suspected, contact Diagnostic
thoroughly mix acids. BioSystems Technical Support at (925) 484-3350, extension 2
4. Wait 5-10 minutes before proceeding with stain procedure to allow or [email protected].
mixed acids to cool.
5. Use Staining Jar only for Sulfation procedure. References
6. Wear protective clothing, gloves, and eyewear when mixing and I. Sheenan, D.C., Hrapchak, B.B. Theory and Practice of
handling this reagent. Make fresh for each use. Histotechnology, 2nd Edition. Battelle Press, Columbus, OH.

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