Zipper Lauren E, Aristide Xavier, Bishop Dylan P, Joshi Ishita, Kharzeev Julia, Patel Krishna B, Santiago Brianna M, Joshi Karan, Dorsinvil Kahille, Sweet Robert M, Soares Alexei S
Office of Educational Programs, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
Photon Sciences Directorate, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
Acta Crystallogr F Struct Biol Commun. 2014 Dec 1;70(Pt 12):1707-13. doi: 10.1107/S2053230X14025126. Epub 2014 Nov 28.
A method is described for using plate lids to reduce evaporation in low-volume vapor-diffusion crystallization experiments. The plate lids contain apertures through which the protein and precipitants were added to different crystallization microplates (the reservoir was filled before fitting the lids). Plate lids were designed for each of these commonly used crystallization microplates. This system minimizes the dehydration of crystallization droplets containing just a few nanolitres of protein and precipitant, and results in more reproducible diffraction from the crystals. For each lid design, changes in the weight of the plates were used to deduce the rate of evaporation under different conditions of temperature, air movement, droplet size and precipitant. For comparison, the state of dehydration was also visually assessed throughout the experiment. Finally, X-ray diffraction methods were used to compare the diffraction of protein crystals that were conventionally prepared against those that were prepared on plates with plate lids. The measurements revealed that the plate lids reduced the rate of evaporation by 63-82%. Crystals grown in 5 nl drops that were set up with plate lids diffracted to higher resolution than similar crystals from drops that were set up without plate lids. The results demonstrate that plate lids can be instrumental for improving few-nanolitre crystallizations.
描述了一种在低体积气相扩散结晶实验中使用板盖减少蒸发的方法。板盖上有孔，通过这些孔将蛋白质和沉淀剂添加到不同的结晶微孔板中（在盖上盖子之前先填充储液器）。针对这些常用的结晶微孔板分别设计了板盖。该系统可最大程度减少仅含有几纳升蛋白质和沉淀剂的结晶液滴的脱水情况，并使晶体产生更可重复的衍射。对于每种盖子设计，通过测量板重量的变化来推断在不同温度、空气流动、液滴大小和沉淀剂条件下的蒸发速率。作为比较，在整个实验过程中还通过目视评估脱水状态。最后，使用X射线衍射方法比较常规制备的蛋白质晶体与在带有板盖的板上制备的蛋白质晶体的衍射情况。测量结果表明，板盖可将蒸发速率降低63%至82%。在5纳升液滴中使用板盖培养的晶体比未使用板盖培养的类似液滴中的晶体衍射分辨率更高。结果表明，板盖有助于改善几纳升结晶。
