ASSESSMENT OF THE ANTIELASTASE ACTIVITIES IN BRONCHOALVEOLAR LAVAGE FLUID - EFFECT OF ASSAY BUFFER IONIC-STRENGTH

Objective: To evaluate the effect of assay buffer ionic strength on as sessment of the antielastase activities in bronchoalveolar lavage flui d. Method: An improved assay protocol in which elastase (in Tris-HCI b uffer) is added to increasing volumes of test samples (made up to equa l volume with phosphate-buffered saline) was used. Results: The percen t NE activity inhibited by BALF decreased with increasing NaCl concent ration of the buffer. Inhibition of pancreatic elastase (PE) was not a ffected. One hundred percent inhibition of NE by pure AAT and SLPI sta ndards occurred at molar ratios of 0.91 +/- 0.03 for AAT-to-NE and 0.8 3 +/- 0.02 for SLPI-to-NE when assayed in buffer with less than or equ al to 0.15 mol NaCl/L, compared to ratios of 0.99 +/- 0.02 and 1.06 +/ - 0.02, respectively, for assays in buffers with 0.5-1.00 mol NaCl/L ( p < 0.05 for AAT-to-NE; p < 0.02 for SLPI-to-NE). The AAT-to-PE molar ratio at 100% inhibition of PE was not affected. Assays in buffer with less than or equal to 0.15 mol NaCl/L indicated that 86.9 +/- 4.1% of AAT and 100.9 +/- 4.9% of SLPI in BALF were active against NE, while assays in buffer with 0.50-1.00 mol NaCl/L showed that 84.4 +/- 3.5% o f AAT and 81.6 +/- 5.9% of SLPI present were active. AAT inhibited NE and PE equally only in buffer with 0.5-1.00 mol NaCl/L. Conclusions: T he results of assays of BALF antielastase activities depend on the ass ay buffer Nacl concentration, which may account for the conflicting re ports in the literature. The buffer with 0.50-1.00 mol NaCl/L appear t o be optimal for valid quantitation of anti-NE activities in BALF.