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After packing, the column was washed with buffer containing 20 mM HEPES, pH 7.4, 100 mM NaCl, 10% (v/v) glycerol, 5 M tirzepatide, 25 M TCEP, 5 mM MgCl 2 , 1 mM MnCl 2 , 0.1% (w/v) LMNG and 0.02% (w/v) CHS first (10 CVs), and then with decreased concentrations of detergents, 0.03% (w/v) LMNG, 0.01% (w/v) GDN and 0.006% (w/v) CHS (20 CVs)

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