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The purified non-acylated tirzepatideGIPRmini-G s Nb35 complex at a concentration of 14-16 mg/mL was mixed with 100 M non-acylated tirzepatide at 4 C and applied to glow-discharged holey carbon grids (Quantifoil R1.2/1.3, Au 300 mesh) that were subsequently vitrified by plunging into liquid ethane using a Vitrobot Mark IV (ThermoFisher Scientific)