Human small cell lung cancer A549 cell line and
nonsmall cell lung cancer cell line NCI-H249, obtained from American
Type Culture Collection (Manassas, VA), were grown in RPMI-1640 supplemented
with 10% fetal bovine serum. For FACS analysis, adherently grown A549
cells were detached by applying nonenzymatical citric saline buffer.21 (link) NCI-H249 cells were grown in suspension. To
minimize nonspecific uptake, the procedures were performed on ice.
Aliquots of cells were blocked with 10% normal goat serum, incubated
with anti-Axl primary antibody h173 (kindly provided by Vasgene Therapeutics
Inc., Los Angeles, CA) produced using composite human antibody technology20 (link) and goat antihuman Alexa Fluor 488 (Invitrogen,
Paisley, Scotland) in sequence. Subsequently, cells were measured
by flow cytometry (CyAn analyzer, Beckman Coulter). Cells without
the incubation of primary antibody were used as negative controls.
nonsmall cell lung cancer cell line NCI-H249, obtained from American
Type Culture Collection (Manassas, VA), were grown in RPMI-1640 supplemented
with 10% fetal bovine serum. For FACS analysis, adherently grown A549
cells were detached by applying nonenzymatical citric saline buffer.21 (link) NCI-H249 cells were grown in suspension. To
minimize nonspecific uptake, the procedures were performed on ice.
Aliquots of cells were blocked with 10% normal goat serum, incubated
with anti-Axl primary antibody h173 (kindly provided by Vasgene Therapeutics
Inc., Los Angeles, CA) produced using composite human antibody technology20 (link) and goat antihuman Alexa Fluor 488 (Invitrogen,
Paisley, Scotland) in sequence. Subsequently, cells were measured
by flow cytometry (CyAn analyzer, Beckman Coulter). Cells without
the incubation of primary antibody were used as negative controls.