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To exclude dead cells, LIVE/DEAD Fixable Aqua Dead Cell Stain (Molecular Probes™, ThermoFisher Scientific) was added simultaneously to the surface staining step.
Surface staining was performed for DC-SIGN, in some of the experiments surface staining of CD3 or CD28 was also included.
Cell surface staining for FACS analysis was performed with MHC class I antibody from eBioscience.
Intracellular staining was performed after cell surface staining as previously described [65].
Cells were air dried, fixed in 4%PFA before surface staining using previously described antibodies.
The cells were counted and 1×106 cells were used for surface staining.
Lymphocyte phenotypes were measured by five-colour surface staining.
Directly conjugated antibodies were utilized for cell surface staining.
PBMC were thawed and rested overnight before surface staining.
Similar(2)
All the antibodies were used for cell-surface staining.
Aside from these details, the GLUT4myc surface staining is homologous to the method of CD36 cell-surface staining.
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