How to Freeze Protein Crystals

Freezing proteins is done in a science lab.

Protein crystals are frozen in order to better study their structure in science laboratories. Protein crystals are very fragile and freezing them provides more strength and protection while they are being tested. Proteins are an essential part of the human body and are being studied in order to further understand the concept of protein folding as well as other protein functions. This could eventually be helpful in the field of medicine to create better medicines and treatments for problems in the human body.

Things You'll Need

• Protein crystals grown for experimentation
• X-ray tube
• Cryo-agent (ethylene glycol or Paratone-N)
• Freezer

Instructions

1. Use protein crystals that have been grown and collected for the purpose of experimentation. Check that the crystals diffract at room temperature by generating x-rays of the crystal in an x-ray tube. Interpreting the data involves assigning indices to each reflection to see the pattern of diffraction. Crystals can be labeled as multiple mosaic, mosaic, multiple or single.

2. Select a cryo-agent, such as Paratone-N or ethylene glycol, to chemically freeze the proteins. Determine the correct solution concentration to freeze the proteins. A 100% solution of ethylene glycol is a common cryo-agent in many labs.

3. Soak crystals in this solution. If the crystals are stable then test to see how long they remain stable. Crystals that remain stable for longer periods of time, such as multiple days, are more useful than crystals that become unstable quickly. If the crystals are unstable, flash freeze immediately in the freezer and test diffraction using the x-ray method discussed earlier. Crystals that develop cracks, bends, dips, or other variations in appearance are not stable.

4. Test the diffraction of the stable crystals. Check to see that diffraction of all frozen crystals is as good as at room temperature. If the diffraction is the same quality then the crystals can be used immediately or stored in a freezer.

5. Soak for a shorter period of time if crystals did not diffract well. Try another cryo-agent if the shorter soaking time does not work. Once the diffraction is of good quality the crystals can be flash frozen for later use.

References

• Structural Medicine: Protein Crystallization Course
• Carleton College: Single-crystal X-ray Diffraction
• Structural Medicine: Crystal Flow Chart