(A) Neutrophils were isolated from two healthy donors and treated with the indicated concentrations of MLi-2 or PF-06447475 for 30 min. Cells were then lysed and 10 µg of whole cell extract subjected to quantitative immunoblot analysis with the indicated antibodies (all at 1 µg/ml antibody) and the membranes were developed using the Odyssey CLx scan Western Blot imaging system. (B) Quantitation of immunoblots was undertaken by analysing phospho-Thr73 Rab10/total Rab10 ratio (Bi) and phospho-Ser935 (full length)/total LRRK2 (full length) ratio (Bii). (C and D) Quantification of immunoblots as above, phospho-Thr73 Rab10/total Rab10 ratio (Di) and phospho-Ser935 (full length)/total LRRK2 (full length) ratio (Dii), except that neutrophils were isolated from two healthy donors and treated with MLi-2 at concentrations of 100 nM for the indicated times. Similar results were obtained in two independent experiments.