Nanocrystal quantum dot-conjugated anti-myeloperoxidase antibody as the detector of activated neutrophils.
Fluorescent nanocrystal quantum dots (QDs) have been applied to a wide range of biological studies by taking advantage of their fluorescence properties. Here we show that QDs conjugated with antibody against neutrophil peroxidase, myeloperoxidase (MPO). We designed a novel method to conjugate QDs to antibody without losing any antibody function including their antigen recognizing and Fc-receptor binding activities. When we applied anti-MPO antibody (Ab) with conventional organic probes in the case of immunostaining of living cells, the antibodies lost their fluorescence because of MPO enzymic activity to produce reactive oxygen species. Our QD-conjugated anti-MPO (alpha-MPO-QDs) can detect MPO on the surface of activated neutrophils. In addition, anti-MPO-QDs did not react to the inactivated neutrophils. In conclusion, we demonstrated that antibody visualized the expression of MPO on the neutrophil surface after stimulation with proinflammatory cytokines. Taken together, these techniques have the possibility that QDs can reveal the activation of neutrophils by immunostaining and flow cytometric analysis as a powerful tool for diagnosis of the neutrophil activation in vitro.
Hoshino
Akiyoshi
A
Research Institute, International Medical Center of Japan, Shinjuku-ku, Tokyo 162-8655, Japan.
Nagao
Tomokazu
T
Nakasuga
Akira
A
Ishida-Okawara
Akiko
A
Suzuki
Kazuo
K
Yasuhara
Masato
M
Yamamoto
Kenji
K
D019268:Antibodies, Antineutrophil Cytoplasmic / Q000008:administration & dosage / Q000737:chemistry; D000936:Antigen-Antibody Complex; D000954:Antigens, Surface / Q000032:analysis; D002469:Cell Separation; D005434:Flow Cytometry; D019085:Fluorescent Antibody Technique, Direct / Q000379:methods; D006801:Humans; D053758:Nanoparticles / Q000737:chemistry; D018375:Neutrophil Activation; D009504:Neutrophils / Q000276:immunology*; D009195:Peroxidase / Q000032:analysis* / Q000276:immunology; D045663:Quantum Dots; D013194:Staining and Labeling
D019268:Antibodies, Antineutrophil Cytoplasmic; D000936:Antigen-Antibody Complex; D000954:Antigens, Surface; D009195:Peroxidase