INFLUENCE OF ACRIDINE TRACER DYES ON NEUTROPHIL FUNCTION

A study was performed to elucidate the effect of two commonly used flu orescent dyes in in vivo microscopic studies, acridine orange (AO) and acridine red (AR), on the ability of phorbol myristate acetate (PMA)o r formyl peptide (fMLP)-stimulated human neutrophils to adhere to a bo vine serum albumin matrix and to generate superoxide anions (SOX). Unl abeled stimulated human neutrophils showed 36 +/- 9% (PMA, 10(-7) M). 11 +/- 7% (fMLP, 10(-7) M) adherence to the matrix. This adhesion was CD18 dependent as evidenced by 98% and 92% reduction, respectively, wh en the anti-CD18 antibody IB4 was included. A dose-dependent inhibitio n of stimulated human neutrophil adhesion was evident after 30 min of in vitro dye labeling and the EC(50) was approximately 70 mu g/ml (AR) and 145 mu g/ml (AO). SOX generation by PMA-stimulated neutrophils wa s unaffected up to 100 mu g/ml AR and AO but was reduced by 40-60% at higher doses. Rabbit neutrophils labeled in vivo or in vitro with 100 mu g/ml AR exhibited 41% and 61% lower SOX generation, respectively. T he study indicates that neutrophil function, in terms of ability to ad here to a BSA matrix using CD11/CD18 integrins and to generate SOX upo n stimulation, is reduced depending on the dose and choice of fluoresc ent dye. Caution should be exercised when using these compounds at hig h concentrations in studies of PMN function.