Cp3 freezer

About 3 μl aliquots of the emptied mature T4 capsids prepared as above were frozen onto Lacey carbon EM grids using a Gatan CP3 freezer with a blotting time of 6 s. The EM grids were then loaded on an FEI Titan Krios EM operated at 300 kV and equipped with a Gatan K2 Summit detector. The data collection was performed using the Leginon program^{46 (link)} with a magnification of 14,000  (physical pixel size: 2.08 Å) in the “super-resolution” mode, which resulted in a pixel size of 1.04 Å per pixel. The dose rate was ~10 e⁻/(pixel·s). A total of 4377 movies, each composed of 40 frames, were collected. Each frame has an exposure time of 250 ms.

Samples of the T/F100 Env trimer–8ANC195 Fab complex were prepared for cryo-EM by mixing dodecyl maltoside (DDM) with the complex to give a final DDM concentration of 0.085 mM^{8 (link)}. Aliquots of 2.7 µl of either the unliganded T/F100 trimer or the T/F100 trimer–8ANC195 Fab complex were applied to Lacey carbon EM grids. The grids were blotted for 6 s and flash frozen in liquid ethane using a Gatan CP3 freezer. The frozen grids were loaded into an FEI Titan Krios EM, operated at 300 kV. The microscope was equipped with a Gatan K2 Summit detector. The data were collected with Leginon^{52 (link)} using a nominal magnification of ×29,000 with a dose rate of ~8 e⁻/(pixel·s) in the “super-resolution” mode, resulting in a pixel size of 0.5 Å. For the data set of the T/F100 trimer–8ANC195 Fab complex, each micrograph was recorded as a movie composed of 40 frames with an exposure time of 200 ms per frame and a total dose of ~64.0 e⁻/Ų. A total of 844 such movies were collected. For the data set of the unliganded T/F100 trimer, each micrograph was recorded as a movie composed of 25 frames with an exposure time of 200 ms per frame and a total dose of ~40.0 e⁻/Ų. A total of 2356 such movies were collected.