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Influence of acridine tracer dyes on neutrophil function
Author(s) -
Hansell Peter,
Berger Elaine,
Chambers J. David,
Arfors KarlE.
Publication year - 1994
Publication title -
journal of leukocyte biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.819
H-Index - 191
eISSN - 1938-3673
pISSN - 0741-5400
DOI - 10.1002/jlb.56.4.464
Subject(s) - acridine orange , cd18 , in vivo , in vitro , microbiology and biotechnology , superoxide , stimulation , bovine serum albumin , neutrophile , biochemistry , n formylmethionine leucyl phenylalanine , adhesion , biology , chemistry , flow cytometry , integrin alpha m , endocrinology , apoptosis , enzyme , organic chemistry
A study was performed to elucidate the effect of two commonly used fluorescent dyes in in vivo microscopic studies, acridine orange (AO) and acridine red (AR), on the ability of phorbol myristate acetate (PMA)‐or formyl peptide (fMLP)‐stimulated human neutrophils to adhere to a bovine serum albumin matrix and to generate superoxide anions (SOX). Unlabeled stimulated human neutrophils showed 36 ± 9% (PMA, 10 ‐7 M) and 11 ± 7% (fMLP, 10 ‐7 M) adherence to the matrix. This adhesion was CD18 dependent as evidenced by 98% and 92% reduction, respectively, when the anti‐CD18 antibody IB4 was included. A dose‐dependent inhibition of stimulated human neutrophil adhesion was evident after 30 min of in vitro dye labeling and the EC 50 was approximately 70 μ g/ml (AR) and 145 μ g/ml (AO). SOX generation by PMA‐stimulated neutrophils was unaffected up to 100 μ g/ml AR and AO but was reduced by 40–60% at higher doses. Rabbit neutrophils labeled in vivo or in vitro with 100 μ g/ml AR exhibited 41% and 61% lower SOX generation, respectively. The study indicates that neutrophil function, in terms of ability to adhere to a BSA matrix using CD11/CD18 integrins and to generate SOX upon stimulation, is reduced depending on the dose and choice of fluorescent dye. Caution should be exercised when using these compounds at high concentrations in studies of PMN function. J. Leukoc. Biol. 56: 464–468; 1994.