title: Blotting-free and lossless cryo-electron microscopy grid preparation from nanoliter-sized protein samples and single-cell extracts.
abstract: We present a sample preparation method for cryo-electron microscopy (cryo-EM) that requires only 3-20nL of sample to prepare a cryo-EM grid, depending on the protocol used. The sample is applied and spread on the grid by a microcapillary. The procedure does not involve any blotting steps, and real-time monitoring allows the water film thickness to be assessed and decreased to an optimum value prior to vitrification. We demonstrate that the method is suitable for high-resolution cryo-EM and will enable alternative electron microscopy approaches, such as single-cell visual proteomics.
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland; Swiss Nanoscience Institute, University of Basel, Switzerland.
authors.forename: Stefan A
authors.initials: SA
authors.lastname: Arnold
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Stefan
authors.initials: S
authors.lastname: Albiez
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Andrej
authors.initials: A
authors.lastname: Bieri
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Anastasia
authors.initials: A
authors.lastname: Syntychaki
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Ricardo
authors.initials: R
authors.lastname: Adaixo
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Robert A
authors.initials: RA
authors.lastname: McLeod
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Kenneth N
authors.initials: KN
authors.lastname: Goldie
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland.
authors.forename: Henning
authors.initials: H
authors.lastname: Stahlberg
authors.affiliation: Center for Cellular Imaging and NanoAnalytics (C-CINA), Biozentrum, University of Basel, Switzerland. Electronic address: thomas.braun@unibas.ch.
authors.forename: Thomas
authors.initials: T
authors.lastname: Braun
mesh_terms: D002457:Cell Extracts; D020285:Cryoelectron Microscopy / Q000379:methods*; D044085:Microfluidics; D011506:Proteins / Q000648:ultrastructure*; D013048:Specimen Handling
chemical_list: D002457:Cell Extracts; D011506:Proteins
keywords: Cryo electron microscopy; Microfluidics; Sample preparation