Vesicle budding reactions were performed according to the protocol developed by , using the non-hydrolysable GTP analogue, GTP-γ-S

<p><b>Copyright information:</b></p><p>Taken from "Two Human ARFGAPs Associated with COP-I-Coated Vesicles"</p><p></p><p>Traffic (Copenhagen, Denmark) 2007;8(11):1644-1655.</p><p>Published online 29 Aug 2007</p><p>PMCID:PMC2171037.</p><p>© 2007 The Authors Journal compilation © 2007 Blackwell Publishing Ltd</p> Vesicles produced in the reaction were separated from the Golgi donor membranes on sucrose density gradients and peaked at 40–43% sucrose. Fractions were analysed by silver stain (upper panel) and immunoblotting with antibodies as indicated (lower panel). The positions of coatomer bands are indicated on the left and also by asterisks. Note the abundant presence of ARFGAP2 and ARFGAP3 in the vesicle fractions defined by the presence of coatomer subunits (fractions 8 + 9) but the absence of ARFGAP1