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Freezing of sputum as a way to improve the applicability of sputum studies

Todor A. Popov, Aleksandar Petlichkovski, Tihomir B. Mustakov, Tanya Z. Kralimarkova


Rationale: Sputum examination is a valuable research tool to study airway diseases, but the requirement to process the samples within 2 hours of their collection poses limitations to its wider applicability. A way to bypass this hurdle would be to freeze the sample at the time of collection and to examine it at a later stage.
Methods and Subjects: We developed a protocol for freezing of sputum upon collection by adding dimethylsulfoxide to it. We tested the reproducibility of cell counts in frozen samples and in fresh portions from the same sputum specimens. We took sputum from 41 asthmatics (18 males) with different levels of control of their disease: 19 of the samples were spontaneously produced and 22 were induced with hypertonic saline.
Results: Significant correlations (p<0.05) were established between the total cell counts, the relative and absolute number of neutrophils, eosinophils and macrophages in the paired fresh and frozen sputum samples. Cell viability in frozen sputum was slightly but consistently lower. Only one frozen sample had viability < 50%. Outcomes in paired samples from induced sputum had better reproducibility than the spontaneous ones.
Conclusion: Examination of frozen sputum samples does not change total cell counts and differential cell counts, despite consistently affecting cell viability compared with fresh sputum cellularity. Still cell viability in frozen sputum was above 50% in all but one examined specimens.


sputum, cellular content of sputum, frozen sputum, induced sputum, asthma, COPD

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About The Authors

Todor A. Popov
University Hospital St. Ivan Rilski, Sofia, Bulgaria

Aleksandar Petlichkovski
Stt. Kiril and Metodij University, Skopje, FYROM
North Macedonia

Institute for Immunobiology and Human Genetics, Faculty of Medicine

Tihomir B. Mustakov
Medical University, Sofia, Bulgaria

Clinical Center Allergology

Tanya Z. Kralimarkova
Medical University, Sofia, Bulgaria

Clinical Center Allergology

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