Isolation of human alveolar macrophages and lymphocytes from bronchoalveolar lavage fluid by centrifugal elutriation

Abstract

Blaschke E, Eklund A, Skog S, Danielsson B. Isolation of human alveolar macrophages and lymphocytes from bronchoalveolar lavage fluid by centrifugal elutriation. Scand J Clin Lab Invest 1985; 45: 691–696.

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.

Key Words:

• alveolar lymphocytes
• alveolar macrophages
• bronchoalveolar lavage
• centrifugal elutriation