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NC: negative control group; PE: phycoerythrin; EGFP: enhanced green fluorescent protein.
(The y-axis logarithmic numbers refer to antibodies bound per cell.) Abbreviations: FITC, fluorescein isothiocyanate; PE, phycoerythrin.
Abbreviations CD: Cluster of differentiation ELISA: Enzyme-linked immunosorbent assay IgG: Immunoglobulin G LPS: Lipopolysaccharide MFI: Mean fluorescence intensity PBMC: Peripheral blood mononuclear cell PCR: Polymerase chain reaction PE: Phycoerythrin RFLP: Restriction fragment length polymorphism SNP: Single nucleotide polymorphism TAE: Tris-acetate-EDTA TNF[alpha]: Tumor necrosis factor alpha TNFR: Tumor necrosis factor receptor sTNFR: Soluble tumor necrosis factor receptor.
Anti-CD4 conjugated to chlorophyll protein (PerCP), anti-CD8 conjugated to phycoerythrin (PE), and anti-CD127 conjugated to phycoerythrin (PE), anti-CD25 conjugated to fluorescein isothiocyanate (FITC) and isotype-matched control antibodies were obtained from Beckman Coulter.
 Nonstandard abbreviations: CEC, circulating endothelial cell; SLE, systemic lupus erythematosus; PBMC, peripheral blood mononuclear cell; HUVEC, human umbilical vein endothelial cell; PE, phycoerythrin; FITC, fluorescein isothiocyanate.
Briefly, the prepared cell suspensions of MSC were incubated with specific antibodies using phycoerythrin (PE)-anti-CD34, PE-anti-CD45, fluorescein isothiocyanate (FITC)-anti-CD29, and FITC-anti-CD44 (e-BIOSCIENCE, USA) before measurement of PMN characteristics.
The coupled microspheres were pelleted by microcentrifugation at 11 340g for 4 min, the supernatant was removed, and the microspheres were resuspended in 75 [micro]L of reporter solution [10 [micro]g/mL streptavidin-conjugated phycoerythrin in hybridization buffer (1X TMAC)] and incubated at room temperature for 15 min.
FITC PE PE-Cy5 APC 1 [kappa] CD34 CD62L CD38 2 [kappa] CD14 CD44 CD38 3 [kappa] CD126 HLA-DR CD38 4 [kappa] CD117 CD128 CD38 5 [kappa] CD11b CD38 CD3 6 [kappa] CD184 CD38 CD19 7 [kappa] CD45ro CD38 CD10 8 [kappa] CD31 CD38 CD54 9 [kappa] CD50 CD38 CD49d 10 [kappa] CD130 CD38 CD44 11 [kappa] CD56 CD38 CD20 12 [lambda] CD34 CD62L CD38 13 [lambda] CD14 CD44 CD38 14 [lambda] CD126 HLA-DR CD38 15 [lambda] CD117 CD128 CD38 16 [lambda] CD11b CD38 CD3 17 [lambda] CD184 CD38 CD19 18 [lambda] CD45ro CD38 CD10 19 [lambda] CD31 CD38 CD54 20 [lambda] CD50 CD38 CD49d 21 [lambda] CD130 CD38 CD44 22 [lambda] CD56 CD38 CD20 Abbreviations: APC, allophycocyanin; FITC, fluorescein isothiocyanate; PE, phycoerythrin; PE-Cy5, phycoerythrin-Cy5.
The microspheres were incubated for 1 h, washed, resuspended in 50 [micro]L of 4mg/L streptavidin phycoerythrin, and incubated for 20 min.
Antibodies were conjugated with fluorescein isothiocyanate, phycoerythrin, peridinin chlorophyll protein, or allophycocyanin.
The CD19 antibody was conjugated with phycoerythrin and the immunoglobulin light chain antibodies were conjugated with fluorescein isothiocyanate.
 Nonstandard abbreviations: COX, cyclooxygenase; MPO, myeloperoxidase; TLR, Toll-like receptor; PSGL-1, P-selectin glycoprotein ligand 1; ARIC, Atherosclerosis Risk in Communities; PC5, phycoerythrin-cyanin 5.1; FITC, fluorescein isothiocyanate; PE, phycoerythrin; L, ligand; MFI, mean fluorescence intensity; FBS, fetal bovine serum; FS, forward scatter; SSC, side scatter; PMA, platelet-monocyte aggregate; PLA, platelet-lymphocyte aggregate; PGA, platelet-granulocyte aggregate.
 Nonstandard abbreviations: scFv, single-chain fragment variable; VH, variable heavy; VL, variable light; FACS, fluorescence activated cell sorting; LOQ, limit of quantification; LOD, limit of detection; SOE, single overlap extension; WT, wild-type; bt-tacro, biotinylated tacrolimus; SA-PE, streptavidin-R- phycoerythrin; MFI, mean fluorescence intensity; [k.sub.off], dissociation rate constant; [K.sub.d], equilibrium affinity constant; CDR, complementarity determining region; mAb, monoclonal antibody; GaM-488, Alexa Fluor 488 goat antimouse.
We then analyzed annexin V binding via flow cytometry (EPICS XL-MCL; Beckman Coulter) with CD45 gating [excitation, 385 nm (argon laser); emission, 525 nm (for phycoerythrin)] and side scatter and monitored the emission signals at 488 nm (fluorescein isothiocyanate) and 675 nm (propidium iodide).
Antibody combinations were CD45 (conjugated to fluorescein isothiocyanate)/CD14 (conjugated to phycoerythrin), CD5/CD19, [Kappa]/CD19, [Lambda]/CD19, CD3/CD23, CD10/CD34, CD25/ CD20, CD22/CD11c, and CD4/CD8.
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