Another tip of the month: When plunge-freeze your grid, either using Vitrobot or other apparatuses, blot off liquid ethane with a small piece of filter paper before dipping the grid into liquid Nitrogen. If you want to look at your grid right away, this prevents column vacuum crash by ethane evaporation inside the column. By the way, I learned this from Yoshi.

This one is for real! How to have a quick and easy estimation of the ice thickness of your cryo grid in the microscope? The easiest way is to ask 3DEM mailing list to tell you how thick is your ice! But, say, what to do if you are still confused after getting so many different answers from the mailing list? Here is another easy way to actually do it yourself: Burn a small hole in the thick ice area. Then tilt your specimen, say to 60 degree, and measure the length of the ice hole tunnel. This can tell you how thick is the ice. You can then convert this ice thickness to the ratio of CCD counts from ice and from the empty hole. Next time, you can estimate the ice thickness directly from the CCD counts.

What do you do if you couldn’t find the beam in the microscope? After making sure that the microscope is working properly and tried in dark all the tricks you have learned but still couldn’t find beam? Here is another trick: Remove the Screen Cover!