Fig 3. HPLC analysis of 5-LO WT and 5-LOΔ4 product formation.

(A) 3 μg purified 5-LO WT (red line) or Δ4 isoform (black line) expressed in E. coli were stimulated by 20 μM AA in the presence of 1 mM ATP and 1 mM calcium chloride. (B) 5 × 10⁶ HEK293T cells transfected with 5-LO WT (red line) or 5-LOΔ4 isoform (black line) were stimulated by 5 μM A23187 and 10 μM AA for 10 min at 37°C. The formed 5-LO products were purified by solid phase extraction and subjected to HPLC analysis using PGB₁ as internal standard. Absorbance was monitored at 280 nm (1–8 min) and 235 nm (8–22 min).