VitroEase™ Buffer Screening Kit and Detergents
| Catalog Number | Description |
|---|---|
| A49856 | |
| A65372 | |
| A65373 | |
| A65374 | |
| A65375 | |
| A65376 | |
| A65377 |
The Thermo Scientific VitroEase Buffer Screening Kit provides pre-formulated buffers and detergents and a comprehensive screening strategy to identify optimal protein conditions for single particle cryo-EM sample analysis.
Features of the VitroEase Buffer Screening Kit include:
• Ease of use—14 buffers and 6 detergents pre-formulated at 10X concentration ready to be spiked in with your protein sample with minimal sample dilution
• Convenient—buffers and detergents provided with color-coded caps that correlate with color-coded screening strategy
• Comprehensive—optimized screening strategy encompassing two rounds of screening with 24 grids with trouble-shooting suggestions
• Flexible—wide range of common buffers and detergents for cryo-EM sample preparation sufficient for sample optimization of 400 grids
• Validated—components and screening strategy validated with single particle cryo-EM analysis
Cryo-EM sample screening evaluates protein behavior in various buffers and additive conditions with the end goal of identifying the optimal conditions in which the protein is stable, randomly oriented, and equally distributed in ice. Kit buffers are provided with pHs from 3.6 to 8.9 pre-mixed with various salts and divalent cations. Detergents are provided at 10X concentrations with an easy-to-use table to convert volume to CMC concentration.
The comprehensive sample screening strategy uses two rounds of screening and 24 grids, taking approximately two days to process (figure below). In the first round, optimal protein concentration, buffer, and pH conditions are determined. In the second round, detergents are further optimized. A condensed strategy is also provided using 8 grids and taking one day to process, which can be used when little sample is available or for use with side entry or semi-automated sample loading systems, like the Tundra Cryo-TEM.
The recommended screening strategies are designed for use with the Thermo Scientific Vitrobot Mk4 and Krios, Glacios, Arctica microscopes with Autoloader systems and EPU multigrid.
- Buffer 1 (10X): C2H3O2 (0.5 M), Cl (1.5 M); pH 3.6 400 μL
- Buffer 2 (10X): C2H3O2 (0.5 M), KCl (3 M); pH 3.6 400 μL
- Buffer 3 (10X): MES (0.5 M), Cl (1.5 M); pH 5.5 400 μL
- Buffer 4 (10X): MES (0.5 M), KCl (3 M); pH 5.5 400 μL
- Buffer 5 (10X): Tris-HCl (0.5 M), Mg (CH2COO)2 (0.1 M), Cl (1.5 M); pH 7.2 400 μL
- Buffer 6 (10X): Tris-HCl (0.5 M), MgCl2 (0.1 M), CH3CO2K (1.5 M); pH 7.2 400 μL
- Buffer 7 (10X): Tris-HCl (0.5 M), Mg(CH3COO)2 (0.1 M), KCl (3 M); pH 7.2 400 μL
- Buffer 8 (10X): HEPES (0.5 M), Cl (1.5 M); pH 7.4 400 μL
- Buffer 9 (10X): HEPES (0.5 M), KCl (3 M); pH 7.4 400 μL
- Buffer 10 (10X): HEPES (0.5 M), Mg(CH3COO)2 (50 mM), CH3CO2K (1.5 M); pH 7.4 400 μL
- Buffer 11 (10X): HEPES (0.5 M), MgCl2 (50 mM), CaCl2 (50 mM), Cl (1.5 M); pH 7.4 400 μL
- Buffer 12 (10X): PBS (1.37 M Cl, 270 mM KCl, 43 mM 2 HPO); pH 7.4 400 μL
- Buffer 13 (10X): Bicine buffer (0.5 M), Cl (1.5 M); pH 8.5 400 μL
- Buffer 14 (10X): CAPSO (0.5 M), KCl (3 M); pH 8.9 400 μL
- Detergent A (10X): CTAB (0.3%) 100 μL
- Detergent B (10X): CHAPS (4.9%) 100 μL
- Detergent C (10X): OG (2.7%) 100 μL
- Detergent D (10X): Tween™-20 Detergent (0.1%) 100 μL
- Detergent E (10X): DM (1%) 100 μL
- Detergent F (10X): FOM (0.7%) 100 μL