ENG Scientific Pneumocystis Carinii Stain Kit

Description

• Pneumocystis carinii is an opportunist which causes a diffuse interstitial pneumonia in patients with impaired immune systems
• The disease itself can be classified as epidemic or sporadic; the latter occurring in patients who have an underlying immunosuppression
• In this procedure, a sulfation reagent of glacial acetic acid and sulfuric acid (not included) is used for the removal of background material
• This allows the Pneumocystis carinii cysts to be visualized more easily after staining with Toluidine Blue 0
• Diagnosis of Pneumocystis carinii pneumonia can frequently be made on bronchoalveolar lavage (BAL) or on touch preparations of pulmonary tissue (open lung biopsies and transbronchial biopsies)
• Recommended Procedures: BAL Specimens: Using a 50 mL tube, centrifuge for 15 minutes at 2000 x g
• Aspirate all but the bottom 5 mL of supernatant
• With a Pasteur pipette, aspirate the sediment plus approximately 1 mL of the remaining 5 mL of fluid
• Transfer material to a 15 mL centrifuge tube, gently mix and prepare smear by spreading a drop over a 1 cmm area
• If concentrated specimen is very thick or mucoid, spread over entire slide with care being taken not to make the smear too thick
• Dry the slides on a heating block at 50 - 55°C
• Allow to cool before staining
• Touch Preparations: When dry, process in same manner as slides of BAL
• Preparation of Sulfation Reagent: In a dry coplin jar submerged in a plastic tub filled with cool water (not below 10°C), mix 45 mL of glacial acetic acid with 15 mL of concentrated sulfuric acid and stir gently with glass rod
• Carefully place smears in reagent for 10 minutes
• Mix immediately and again after 5 minutes
• Gently rinse with water for 10 minutes
• Drain excess water and stain with Sol
• I for 3 - 5 minutes
• Rinse with Sol
• II followed by Sol
• III for 10 seconds each
• Clear in Sol
• IV, two changes each for 10 seconds and mount
• Results: The cyst forms appear as lavender structures (cup-shaped) approximately 5 urn in diameter
• The cyst outline is distinct, and the internal region stains uniformly
• Note: A negative bronchoalveolar lavage does not rule out an infection with Pneumocystis carinii
• Other diagnostic procedures may be necessary.