Using Micromounts™ and MicroLoops™ for Macromolecular Crystallography

Click here for a printable pdf file of these instructions.

MicroMounts and MicroLoops are used in much the same way as nylon loop mounts, and are much easier to use than glass fibers and other mounting methods traditionally used in small-molecule crystallography. Their nonmagnetic stainless steel rods are compatible with all existing goniometer bases (caps) and mounting hardware. Our suggested procedures follow.

Select a mount with a sample aperture size slightly smaller than the crystal size. In most cases you don't want to trap the crystal inside a liquid meniscus spanning the aperture, common practice when using nylon loop mounts. Instead, the crystal should rest on top of the aperture, so as to minimize excess liquid and background X-ray scatter. Choosing a slightly smaller aperture also makes it much easier to fish the crystal out of its solution.

Cut the stainless steel rod (if necessary) to a desired length using our cutters or other cutters designed for spring-temper steel. Use the millimeter scale on the label of the product box, lining up the end of the pin with the right end of the scale.

Insert the pin into a standard goniometer base using a small amount of, e.g., Dow-Corning #976V high vacuum grease or Duco Cement hold it in place. For easier handling, attach the base to a magnetic rod. The pin can also be inserted into a 0.7 mm mechanical pencil.

Slowly and carefully insert the gold-colored polyimide into the crystal-containing drop, trying to minimize fluid disturbances and motion in the drop. Slide the sample aperture under your crystal, and then carefully remove the crystal + mount from the drop, keeping the crystal centered over the sample aperture.

To retrieve crystals that have settled to the bottom of a crystallization plate, try pressing downward to bend the tip of the mount so that it slides flat along the bottom.

If using MicroMounts, you can remove excess liquid by slowly and carefully inserting a size 15 paper wick into the large opening that connects to the sample hole. If you have lots of liquid and you insert the wick too quickly, the crystal will flow with the liquid to the wick. For viscous liquids you may need to "mop up" around the crystal with the wick.

Mounted correctly, you should have very little liquid around your crystal - far less than when using nylon loops. The flash cooling rate will be much faster than in a loop and the cryoprotectant concentrations required to prevent icing much smaller.

In general, it's bad to have a lot of any liquid - water or oil - around your crystal during flash cooling: when the liquid freezes, it will tend to crush your crystal. If you have just a thin layer of liquid around the crystal, it will shatter (like the shell of an egg) and so do little damage.

If your penetrating cryoprotectant isn't enough to keep the external liquid from crystallizing, transfer your crystal to a drop of our LV Cryo Oil , mount it and then remove excess oil by wicking it away, or by flicking it away by gently tapping on the steel rod of the mount.

Flash cool the crystal by your favorite method. We recommend using the hyperquenching protocol of Warkentin et al. (J. Appl. Cryst. (2006) 39, 805-811) , a variant on standard plunge cooling in liquid nitrogen in which you you blow away the cold gas layer that forms above the liquid nitrogen just before you plunge in your sample. You can also use liquid nitrogen that has been vacuum-pumped to near its freezing temperature (to reduce boiling), or (more dangerous) liquid propane, but the hyperquenching method is safer, easier and generally gives the best results.

Comments or suggestions? Please contact us!