sentence: The FLAG-tagged receptors , Golgi , and TGN were labeled for 1 h in blocking buffer with anti – FLAG M1 antibody ( 1:1000 ; F3040 , Sigma-Aldrich ) conjugated with Alexa 647 ( A20186 ; Molecular Probes ) , anti-GPP130 ( a gift from Adam Linstedt , Carnegie Mellon University , Pittsburgh , PA ) and anti – TGN-38 rabbit polyclonal antibody ( 1:2000 ; T9826 ; Sigma-Aldrich ) , respectively .