Broncho-alveolar lavage specimens (BAL) for CMV surveillance collected on day 35 post-bone marrow transplant patients or routine screening of solid organ transplant patients are examined for CMV only. http://bahankuliahkesehatan.blogspot.com/
II. Collection and Transport
BAL fluid should be collected into a clean, sterile container. If a delay in transport or processing is anticipated, keep the specimen at 4 oC.
A. Processing of Specimen:
a) Approximately 2-3 mL of centrifuged sediment should be received from the specimen receiving planting area.
b) Transfer 2 mL of sediment to a sterile freezer vial containing 4 drops (0.2 mL) gentamicin (1 mg/mL) and 2 drops (0.1 mL) of fungizone (250 mg/mL) to a final concentration of 100 mg/mL and 10mg/mL respectively.
c) Allow to stand at room temperature for 10 minutes
d) Refer to Appendix II for Shell Vial inoculation.
B. Direct Examination:
C. Isolation and Identification:
Incubation at 36oC
aMRC-5 = Human fibroblast cells
bCMV-IE= Monoclonal antibody IFA stain for CMV immediate early antigen
Refer to Appendix II for Shell Vial staining and interpretation.
D. Interpretation and Processing of Cultures:
a) At 48 hours observe shell vial coverslips using the inverted microscope:
i) If <50% or no CPE - Perform DFA for CMV immediate early antigen
(See Appendix V).
ii) If >50% CPE - Prepare a cytospin slide of scraped cells and then perform DFA for CMV immediate early antigen.
iii) If positive, freeze shell vial supernate.
IV. Reporting Results
Shell Vial: Negative report: “Negative for Cytomegalovirus.”
Positive report*: “POSITIVE for Cytomegalovirus.”
1. Gleaves, Curt A. et al. Cumitech 15A “Lab Diagnosis of Viral Infections”.
American Society for Microbiology, August 1994.