■　Protein crystallization with batch method

〇 Features

  • Small amount of solution （1 µL～）　
  • Easy to observe crystals in a capillary
  • Crystals are stable in capillaries.
  • Concentration of the crystallization reagent is not changed.

    • 〇 Protocol for Lysizyme crystallization

       —A protein crystallization kit for the batch method (C-Kit ABE) is used.

       —Suitable for STEM education and hands-on science learning.

       Click here to purchase the kit:  info@confsci.co.jp.

       ***When students conduct experiments, please provide guidance to the instructor.***

    1. Keep the solutions (buffer solution, sodium chloride solution, and lysozyme solution) to room temperature (20°C).
    2. Shake the solutions gently to make them homogeneous.
    3. Make the sample solutions by placing the solutions into microtubes in the following order.

      ・Load the buffer solution, add the sodium chloride solution, and then the lysozyme solution, in this order.

      ・Gently mix the solution with a micropipette to ensure uniform concentration (be careful not to whisk).

    4. Using capillary action, fill the capillary with the sample solution.

      5. ・If the solution has difficulty rising up the capillary, tilt the microtubes to help the solution rise.

      ・The amount of liquid to be filled should be about 10 mm shorter than the length of the capillary, taking into account that the ends of the capillary will later be sealed with sealing compound.

      　　　　

      ※If the capillary cannot be easily raised by capillary action, a micropipette can be used to suck up the solution.

           ・Attach a silicon tube to the tip of the pipette tip, attach a capillary to the tip, and turn the dial of the micropipette to suck up the sample solution. Finally, suck in about 5 mm of air.

           ・Seal the end of the capillary with sealing compound, and then remove the capillary from the silicone tube. (How to seal with sealing compound is 6)

           ・After this, proceed to 6.

      　　　　

    5. Add air to both ends of the capillary.

      ・Hold the capillary with the side containing the solution up to the end in your hand, and lightly strike your hand against the lab bench to inject air about 5 mm from the end.Be careful not to strike the capillary too hard, as the solution will escape from the capillary.

      （The reason for adding air to both ends is to prevent the sealing compound from coming in contact with the sample solution. If the sealing compound comes in contact with the sample solution, it may enhance the formation of crystals, resulting in a variation in the reproducibility of the number of crystals obtained.）

    6. Seal both ends of the capillary with sealing compound.

      ・The sealing compound should be heaped onto a flat surface such as a spatula and pressed lightly against the ends of the capillary to ensure that it enters the capillary properly.

    7. Place the capillary vertically in a conical tube and allow it to stand at room temperature (20°C).

      8. ・・・・・

    8. Observe the crystals every few days for 1-2 weeks.

      ・Turn on the touch light, place two polarizing plates on top and put the capillary between the polarizing plates. Rotate the upper polarizer horizontally to change the way light passes through it. The crystal appears to glow.