Tools and Reagents for Cryo-EM

The past few years have been revolutionary for the field of single-particle electron cryo-microscopy (Cryo-EM), with over 50% of the total deposited structures being determined since 2014. Currently, there are over 1,300 unique (<95% sequence identity) Cryo-EM structures deposited in the PDB, over 150 of which are membrane proteins.  Here, we have curated all of the commonly used tools from Anatrace and Molecular Dimensions in Cryo-EM experiments.
 

Detergents for Cryo-EM

The following detergents have been sucessfully used in the Cryo-EM studies of membrane proteins.  Want to learn more?  Check out our compilations of membrane protein strucutures for 2016, 2017, and 2018.
 

GDN101 - GDN

 

A835 - Amphipol A8-35

 

P5008 - Amphipol PMAL-C8

Digitonin is commonly used for Cryo-EM, but there are many drawbacks including batch-to-batch variability and solubility. GDN has been shown to be an effective drop-in substitute for Digitonin which is being used in a number of recent structures.
First described in 1996 by Jean-Luc Popot, amphipols are a class of polymers that can stabilize membrane proteins in a detergent-free, aqueous solution. To date, there have been over 20 Cryo-EM structures of membrane proteins determined using Amphipol A8-35.
In recent years, PMAL-C8 has been gaining traction for use in Cryo-EM with a number of unique structures published. PMAL amphipols are zwitterionic, and contain repeating units of a carboxyl, ammoniumamidate, and alkyl chain.

D310 - DDM


NG310 - LMNG

Lipid Nanodiscs

The most commonly used detergent in membrane protein crystallization, Dodecyl Maltoside (DDM), has also been used in the Cryo-EM structures of a number proteins. DDM is also often used as a mixture with Cholesteryl Hemisuccinate (CHS).
Due to its very low CMC, the concentration of LMNG in the buffer can often be reduced to low concentrations, reducing the amount of free detergent micelles, and reducing background. Like DDM, LMNG is often used as a mixture with Cholesteryl Hemisuccinate (CHS).
Lipid nanodiscs allow for the reconstitution of a detergent solubilized membrane protein into a lipid environment, and are being increasingly used in Cryo-EM. Anatrace offers a full selection of the lipids commonly used in nanodisc reconstitution.

Screens to Improve Sample Preparation

As with any strucutral biology method, sample preparation is paramount to success. The following screens are designed to quickly optimize your protein buffer conditions and increase overall thermostability, allowing you to spend more time on the things that matter, like solving structures and publishing papers!
 

The Durham Screens

The RUBIC Screens

Developed by Emily Cardew and Ehmke Pohl at the University of Durham, The Durham pH Screen and the Durham Salt Screen provide greater depth of analysis of the effect of individual buffers, pH, and salts.
Developed by Stephane Boivin and Rob Meijers at EMBL Hamburg, the RUBIC Buffer Screen provides 96 unique buffer formulations, while the RUBIC Additive Screen analyzes the effect on stability of common additives.

Fluorinated Surfactants for Improving Vitrification

The main sample preparation step in a Cryo-EM experiment is dispensing a protein solution onto a carbon grid followed by plunge freezing, creating a thin layer of vitreous ice. There is increasing evidence that the addition of fluorinated surfactants in the sample buffer can improve the vitrification process. Want to learn more about fluorinated surfactants? Read our May 2017 newsletter.
 

F300F - Fluorinated FC-8

O310F - Fluorinated OM

Fluorinated Fos-Choline-8 has been used to improve the vitrification of a number of membrane protein structures including the CFTR, and TRPV5. The typical concentration used is 3 mM.
Fluorinated Octyl Maltoside has been used to improve the vitrification of a number of membrane protein strucutres including TcdA1 and RyR1. The typical concentration used is 0.01% - 0.2%.

Tools for Cryo-EM

We are excited to offer a full range of sample management tools for your Cryo-EM experiments, including Grid Boxes, Grid Box Storage Pucks, and Long Term Storage & Shipping Dewers.
 

Grid Box Storage Pucks

Cryo-EM Grid Boxes

Cryogenic Shippers & Dewers

Our storage system allows you to easily track your grid-boxes for storage and shipping. Each grid-box is uniquely located by chamber number and puck identifier. Researchers and microscope users can quickly and reliably identify samples of interest.
This new grid-box from Swissci has been developed in collaboration with Dr Jan Löwe at the MRC Laboratory of Molecular Biology in Cambridge. Backed by years of experience in Cryo-EM, it includes many improvements on current models.
Molecular Dimensions offers a full range of Dry Shippers, Cryogenic Refrigerators, and Cryo Tools for the safe storage and transfer of Cryo-EM Pucks.