title: Isolation of human alveolar macrophages and lymphocytes from bronchoalveolar lavage fluid by centrifugal elutriation.
abstract: Cell populations obtained by bronchoalveolar lavage (BAL) from humans showed varying proportions of lymphocytes and macrophages, about 5% and 94%, respectively, in healthy controls (n = 12), and about 24% and 75%, respectively, in patients with sarcoidosis (n = 13). The present study was carried out to examine centrifugal elutriation as a means to obtain from human BAL fluids highly purified populations of viable cells, required for further biochemical investigations. After centrifugation of BAL fluids at 400 g for 5 min, cells were resuspended in Hank-Tris solution, loaded into a Beckman elutriator rotor (rotor speed 2800 rpm) and elutriated with Hank-Tris in approximately 10 min. Cells were collected at flow rates of 20 ml/min (fraction I), 32 ml/min (fraction II) and, after stopping the rotor, 80 ml/min (fraction III). Fraction I contained mainly lymphocytes, about 54% and 74% in controls and sarcoidosis, respectively. Fraction III consisted almost solely of macrophages, while fraction II contained mixed populations. Total cell recovery was about 74% and 55% in controls and sarcoidosis, respectively. Half of the missing cells were lost during centrifugation, the rest during elutriation. Cell viability was 85-90% after the first centrifugation. Following elutriation, the viability was found unchanged in fraction III, but decreased to about 50% in fraction I of controls. The decrease may be due to the accumulation of dead cells with decreased density in this fraction. The results showed that centrifugal elutriation is a highly suitable technique for the isolation of macrophages from human BAL fluids. Enriched lymphocyte populations may be obtained from sarcoid BAL fluids.
mesh_terms: D001980:Bronchi / Q000378:metabolism*; D002452:Cell Count; D002469:Cell Separation; D002498:Centrifugation; D005122:Exudates and Transudates / Q000166:cytology*; D006801:Humans; D007958:Leukocyte Count; D008171:Lung Diseases / Q000473:pathology; D008214:Lymphocytes; D008264:Macrophages; D011650:Pulmonary Alveoli / Q000378:metabolism*; D012507:Sarcoidosis / Q000473:pathology; D007507:Therapeutic Irrigation