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Differentiation Between Attached And Ingested Immune Complexes By A Fluorescence Quenching Cytofluorometric Assay.

S. Sahlin, J. Hed, I. Rundquist
Published 1983 · Chemistry, Medicine

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Immune complexes attached to and ingested by human polymorphonuclear (PMN) cells were quantified by cytofluorometry using a fluorescence quenching assay which permits differentiation between attachment and ingestion. The fluorescence intensity decreased after ingestion as a result of the low pH in the phagolysosomes. When extracellular pH was lowered a slight decrease in phagolysosomal pH was detected in macrophages but not in PMN. When measuring total fluorescence, interaction at pH 5.8 for PMN and at pH 4.4 for macrophages is recommended, since the intensity of extra- and intracellular fluorescence are equal under these conditions. Thirty different dyes were tested for dye exclusion and fluorescence quenching of FITC-conjugated yeast particles, and FITC-conjugated IgG. Because of the lysosomotropic effect of basic dyes, acid and direct dyes are preferable as quenching agents. We could not find physical or chemical properties of the dyes that correlated with their quenching effect. Heat aggregated IgG was used as an immune complex analogue in the development of the assay. Trypan blue (0.2 mg/ml) at pH 4.4 was found to be the best quenching agent of extracellular fluorescence when using ingested aggregated IgG. The technique offers a simple method of quantifying ingested protein aggregates and of studying heterogeneity in phagocyte populations.
This paper references
10.1084/JEM.136.3.514
IN VITRO ADHERENCE OF SOLUBLE IMMUNE COMPLEXES TO MACROPHAGES
W. Arend (1972)
10.1038/248156A0
Detection and radioassay of soluble circulating immune complexes using guinea pig peritoneal exudate cells
I. Onyewotu (1974)
10.1016/0003-9861(69)90454-8
Fluorescent protein-dye conjugates. II. Gamma globulin conjugated with various dyes.
R. F. Chen (1969)
10.1111/j.1365-3083.1977.tb02153.x
Differences in the Mode of Phagocytosis with Fc and C3 Receptors in Macrophages
G. Kaplan (1977)
Quantitative phagocytosis by neutrophils. I. A new method with immune complexes.
P. A. Ward (1973)
Quatification of phagocytosis by human neutrophils. The use of radiolabelled staphylococcal protein A-IgG complexes.
R. Hälgren (1976)
10.1038/290406A0
The respiratory burst of phagocytic cells is associated with a rise in vacuolar pH
A. Segal (1981)
10.1016/0006-2952(74)90174-9
Commentary. Lysosomotropic agents.
C. de Duve (1974)
10.1111/J.1574-6968.1977.TB00651.X
The extinction of fluorescence by crystal violet and its use to differentiate between attached and ingested microorganisms in phagocytosis
J. Hed (1977)
10.1016/0014-4827(69)90290-0
Studies on phagocytosis. I. Uptake of radio-iodinated (131-I) human serum albumin as a measure of the degree of phagocytosis in vitro.
Y. Chang (1969)
10.1177/12.4.271
THE EFFECT OF HYDROGEN ION CONCENTRATION ON FLUORESCENT LABELLED ANTIBODIES
R. Hiramoto (1964)
10.1084/JEM.145.5.1368
Association and dissociation of aggregated IgG from rat peritoneal macrophages
D. Knutson (1977)



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R. Tanikawa (2014)
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T. Umeda (2008)
Different ffunctional a and m morphological c characteristics in a a n nonadherent s subpopulation o of h human m macrophages rrecovered by b bronchoalveolar llavage
C. Barck (1995)
10.1016/J.POWTEC.2014.01.088
Self-assembled macromolecular nanocoatings to stabilize and control drug release from nanoparticles
Schalk J. Strydom (2014)
10.1111/j.1462-5822.2010.01487.x
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V. R. Gupta (2010)
10.1117/12.431551
Fluorescence imaging of endothelial cellular responses in the intact lung microvasculature
Wolfgang M. Kuebler (2001)
10.1002/BIO.727
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M. Suzuki (2003)
10.1186/1743-8977-10-2
Deciphering the mechanisms of cellular uptake of engineered nanoparticles by accurate evaluation of internalization using imaging flow cytometry
S. Vranic (2012)
10.22028/D291-22664
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A. Neumeyer (2010)
10.1021/acsnano.7b05528
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10.1016/j.nano.2010.12.007
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A. Neumeyer (2011)
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M. M. Amaral (2007)
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10.1134/S1995078018010147
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A. S. Zhirnik (2018)
10.1016/j.jim.2016.10.006
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Andrew C. Melton (2017)
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T. Binder (2010)
10.1093/humrep/des208
Trophoblast cells induce a tolerogenic profile in dendritic cells.
G. Salamone (2012)
Characterization of staphylococcal small colony variants and their pathogenic role in biomaterial-related infections with special reference to Staphylococcus epidermidis
S. Matar (2004)
10.1016/j.imbio.2010.12.005
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K. Staples (2011)
10.1128/IAI.73.9.5547-5553.2005
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S. Al-Robaiy (2005)
A study of the coupling between morphodynamics and energy metabolism in macrophages
G. V. Helden (2014)
10.1007/BF00214490
Modulation of phagocytic and oxidative burst activities of bovine neutrophils by human recombinant TNF-α, IL-1-α, IFN-γ, G-CSF, GM-CSF
M. B. Kabbur (2004)
10.3390/nano10122417
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Ruhung Wang (2020)
10.1002/PSC.678
Investigation of penetratin peptides. Part 2. In vitro uptake of penetratin and two of its derivatives
T. Letoha (2005)
10.3109/01902148909069608
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Karl A. Nyberg (1989)
10.1016/0022-1759(87)90224-9
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J. Hed (1987)
10.1016/j.jddst.2017.04.013
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Courtney A. Follit (2017)
10.1002/(SICI)1097-0320(19970201)27:2<145::AID-CYTO6>3.0.CO;2-F
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K. Hess (1997)
10.1021/bm301294n
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D. Benoit (2012)
10.1016/J.JCONREL.2005.08.007
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