ELISA OF COMPLEMENT C3A IN BRONCHOALVEOLAR LAVAGE FLUID

Activated complement factors within the lung may induce several local biological effects. In order to investigate local complement activation we have developed non-competitive two-site ELISAs of C3a and total C3 in bronchoalveolar lavage fluid (BALF). For the assay of C3a, both C3 and C3(H2O) were removed from the samples by precipitation with polyethylene glycol. It was necessary to add carrier proteins to BALF to remove C3 and C3(H2O) fully. The ELISA of C3a has the lowest limit of detection reported thus far, namely 0.045 nM (= 0.405 ng/ml). In BALF from healthy persons (n = 9) the C3a concentration was 0.20 nM (0.12-0.31 nM) (median, range). C3a was higher in BALF from patients with asthma or with sarcoidosis; asthma (n = 10), 0.45 nM (0.20-5.79 nM); sarcoidosis (n = 19), 1.31 nM (0.095-5.65 nM) (Mann-Whitney U test, p < 0.005). In BALF from patients with Pneumocystis carinii pneumonitis (n = 10) the C3a concentration was 0.18 nM (0.07-0.57 nM). C3a concentrations in BALF may reflect local complement activation in the lung and/or diffusion into the lumen. This was studied by normalizing C3a concentrations in BALF into values for epithelial lining fluid (ELF), and calculating serum-to-ELF quotients of C3a, and C3a/total C3 quotients.