Gatan quantum ls imaging energy filter

Purified samples of bovine IRBP or IRBP + mAb5 isolated from size‐exclusion chromatography were adsorbed to GO coated Quantifoil R1.2/1.3 Cu grids at a concentration of 0.17 mg/mL and plunge frozen in liquid ethane using an FEI Vitrobot (Thermo Fisher Scientific, Waltham, MA, USA). Sample was applied to the grids under 80% humidity and 4 ºC. Grids were plunged after incubation with sample for 5 seconds. Data were acquired at C‐CINA, University of Basel. In total, six datasets were collected on an FEI Titan Krios (Thermo Fisher Scientific, Waltham, MA, USA) operating at 300 keV and equipped with a Gatan Quantum‐LS imaging energy filter (GIF, 20 eV energy‐loss window; Gatan Inc, Pleasanton, CA, USA). Movies were acquired with a K2 Summit direct electron detector (Gatan Inc) in super resolution mode using SerialEM with a physical pixel size of either 1.058 Å or 0.831 Å and a total electron dose of 60‐81 e⁻ Å⁻² per micrograph.³⁵ Micrographs were motion‐corrected and dose‐weighted using MotionCor2 implemented in FOCUS.³⁶, ³⁷ Additional collection parameters can be found in Supporting Information (Table S1).