Effect of freezing conditions on distances and their distributions derived from Double Electron Electron Resonance (DEER): A study of doubly-spin-labeled T4 lysozyme

► DEER with very rapid freezing yielded broader distributions than regular freezing. ► Neither method of freezing altered the distance maxima. ► Rapid freezing yielded more uniform distribution of spin-labels and enhanced DEER S/N. ► Rapid frozen samples require less cryoprotectant than regularly frozen samples. ► Bromo-substituted spin-label produced narrower distance distributions..