Binding of control 12-mer peptides (L33 and P33) extended with a biotinylated
linker peptide were incubated with DRB3*01:01-positive cell lines (STEINLIN
and D4BL4) or control cell lines (DUCAF, EMJ) at 5 μM peptide in the
presence of 2.5 μM AdEtOH. The efficiency of binding to APCs was assessed by
flow cytometry with streptavidin-conjugated R-PE. (A)
Representative raw data of one of 3 independent peptide binding assays (1 of
2 replicates shown). (B) Comparison of percent specific
binding. Data points from independent experiments are presented as dots,
with bars representing mean ± SEM of at least 3 experiments. Raw data values
were median R-PE fluorescence intensity on B-LCLs (gated by light scatter
cytogram). Background intensity (cells only, no peptide) was subtracted
before calculating the specific binding within each experiment (L33 peptide
on STEINLIN defined as 100%). AdEtOH, adamantane ethanol; APC, antigen
presenting cell; R-PE, R-phycoerythrin; B-LCL, B-lymphoblastoid cell
lines.