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Freeze-etching, the practice of removing excess surface water from a sample through sublimation into the vacuum of the analysis environment, has been extensively used in conjunction with electron microscopy. Here, we apply this technique to time-of-flight secondary-ion mass spectrometry (ToF-SIMS) imaging of cryogenically preserved single cells. By removing the excess water which condenses onto the sample in vacuo, a uniform surface is produced that is ideal for imaging by static SIMS. We demonstrate that the conditions employed to remove deposited water do not adversely affect cell morphology and do not redistribute molecules in the topmost surface layers. In addition, we found water can be controllably redeposited onto the sample at temperatures below −100 °C in vacuum. The redeposited water increases the ionization of characteristic fragments of biologically interesting molecules 2-fold without loss …