40 ti rotor

Vesicle isolation protocol was modified from Hendrix’s report [15 (link)]. In brief, cells (2 × 10⁸) were sonicated and supernatants were obtained by centrifugation (3,000 g for 10 min at 4°C) and vesicles were enriched from supernatants by high speed centrifugation (30,000 g for 60 min at 4°C) using a 40-Ti rotor (Beckman Coulter, CA, USA). The vesicles-containing solution (800 μg) was incubated with anti-Flag antibody to isolate Rab37-specific vesicles and the cargos in vesicles were analyzed by immunoblotting using indicated antibodies (Supplementary Table 2). For cellular fractionation, intact cells suspended in the lysis buffer (25 mM Hepes, pH6.5, 250 mM sucrose, 1mM EDTA and protease inhibitor cocktail) were pelleted by centrifugation (250 g for 10 min). The cytoplasmic (supernatant) and membrane (pellet) fractions were then obtained by centrifugation at 16000 g for 60 min at 4°C.

Vesicle isolation protocol was as described in our previous study 23 (link). Jurkat (2 × 10⁶), THP1 (1 × 10⁷) or RAW264.7 cells (1 × 10⁷) were sonicated and subjected for vesicle enrichment by two-speed of centrifugations (3,000 g for 10 min followed by 30,000 g for 60 min at 4 °C) using a 40-Ti rotor (Beckman). The vesicles-containing solution (500 μg) was incubated with anti-V5-tag antibody to isolate Rab37-specific vesicles and the CHI3L1 cargos in vesicles were analyzed by immunoblotting (Table S2).

Vesicle isolation protocol was modified from Kuo’s report [16 (link)]. Jurkat T cells (2 × 10⁷) expressing His-V5-tagged Rab37 were resolved in 300 μl of 200 mM sucrose and gently sonicated. The supernatants were obtained by centrifugation (3000×g for 10 min at 4 °C). Vesicles were enriched from supernatants by high-speed centrifugation (30,000×g for 60 min at 4 °C) using a 40-Ti rotor (Beckman, Coulter, CA, United States). The vesicles-containing solution was incubated with anti-V5 antibody to isolate Rab37-specific vesicles and the cargos in vesicles were analyzed by Western blot using the indicated antibodies.