FIG 1.

HPLC chromatograms and HEK-TLR2 stimulation with lipoproteins and nonlipidated proteins. (A) Overlay of RP-HPLC chromatograms. Samples were run on a C₈ reverse-phase column using an acetonitrile-trifluoroacetic acid (TFA) mobile-phase system, and protein signal was detected at 280 nm using a diode array detector. The peaks between 15 and 20 min reflect the lipidated proteins. (B) HEK-TLR2 and HEK293 (expressing no TLR) cells were stimulated with decreasing doses of Pam₃CSK₄, SP_2108 LP (lipoprotein), SP_0148 LP, or nonlipidated (Δlipid) proteins. Following an 18-h stimulation, IL-8 was measured from the supernatants. Median values with interquartile ranges from at least 4 independent stimulations are shown. The experiments performed with all shown stimuli were repeated in HEK293 cells, and the results were at or below the limit of detection in the assay.