Growing your first crystal
Who said crystallizing a protein was easy?
Here are our tips for starting out...
First of all, we suggest optimizing your protein expression. There are some great tips and suggestions in the following video presented by Dr Sheldon E. Broedel:
You can screen your protein expression with the Media Optimization kit.
Make sure your protein is:
- ~95% pure.
- Folded (CD)
Screen your sample's thermal stability using Differential Scanning Fluorimetry (DSF, or Thermofluor). Read more
Most crystallization is carried out with protein at a concentration of at least 10mg/ml, but lower can work fine too. We recommend not to use a phosphate buffer as you'll find that most of your crystals will turn out be be salt - the most popular buffers are HEPES and Tris. For concentration or buffer exchange of your protein or complex, we have a full range of ultrafiltration and dialysis devices suitable for different sample volumes, and molecular weight cut-offs.
Wondering where to start with your screens? Our global product manager Dr. Jeanette Hobbs is here with some advice:
Starting with JCSG-plus™ and PACT premier™ can give an excellent hit rate in just 192 conditions:
We also recommend the SG1™ Screen developed by Janet Newman to represent the most successful, non-redundant frequently reported crystallization conditions from all the early commercially available crystallization screens.
If you are setting up your screens by hand, we recommend either a hanging drop or a sitting drop option:
Hanging drop - we suggest 24-well XRL plate with cover slips
Sitting-drop - you can use sitting-drop bridges with the XRL plate, or you can try the 48-well MRC-Maxi sitting drop plate, which can be sealed with ClearVue sheets.
Your crystal trays need to be stored at a constant temperature, ideally with as little vibration as possible (trust us, this is important for reproducing those all important crystals!). Most people start with room temperature (17 - 21°C) with an additional temperature of 4°C depending on the protein, and quantity of material. If you don't have access to such equipment, we can recommend this range of benchtop incubators
You'll also need a microscope to check those crystal trials, we recommend using one with a polarizer to help visualize crystalline material in the drop. Check out our full range of low-cost microscopes here.
As always, if you have any questions at all, please feel free to contact email@example.com
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