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FACS trafficking MIN6B1-SNAP-GLP-1R or CHO-SNAP-GLP-1R cells were pre-incubated in HEPES-bicarbonate buffer plus 3 mM glucose and 1% BSA before labeling at 37 C with 1 M cleavable BG-SS-488 SNAP-Surface probe (a gift from New England Biolabs) and stimulation with 100 nM agonist in HEPES-bicarbonate buffer plus 11 mM glucose and 1% BSA, and trafficking assays performed as follows 27 : Briefly, for receptor internalization, cells were placed at 4 C to arrest endocytosis following incubation at 37 C for the indicated times, or directly put at 4 C for 0 min time-point, and treated with ice-cold alkaline TNE buffer 100 mM MesNa to strip surface-exposed label (plus MesNa) or to measure total labeled receptor (minus MesNa)

In between-drug comparisons, all tirzepatide doses were comparable with semaglutide 2.0 mg and superior to semaglutide 1.0 mg and 0.5 mg