Growing crystals from solution some practical advice

4 Growing crystals from solution some practical advice 

Proteins generally are available in small quantities and are often difficult to crystallize. Therefore the strategy is to use a combinatorial method in which selected combinations of different organic solvents, different pH ranges, different salt concentrations, and different precipitating agents are tested for their ability to aid in production of diffraction-quality crystals. Very small amounts of protein solution (10-30 mg/ml), for example, 2 pi (also sometimes called 2 A) are used. In this way the best conditions for crystallization can be refined. Recipes for growing crystals are usually only specific for a particular protein. 

FIGURE 2.8. Some methods for growing crystals from solution, (a) Slow solvent evaporation, primarily used for small molecules, (b) Vapor diffusion, (c) The hanging drop method, primarily used for macromolecules, (d) A crystallization plate used for hanging drops there are slightly different crystallization conditions in each well, varied systematically with respect to, for example, pH and ionic strength. 

Slow solvent evaporation is a valuable method for producing crystals of high quality it is used primarily for small molecules. The rate of growth is manipulated by adjusting conditions (initial concentration, temperature, evaporation rate) so that crystals are obtained in a few days, not months or minutes. The rate of evaporation can be adjusted by covering the solution with a watch glass or a piece of film (aluminum or Parafilm ) with a number of small holes punched in it. The solution is then left to stand undisturbed. 

Single-component solvent evaporation methods depend on the removal of solvent in order to obtain the saturated solution required for crystallization to occur. If single-solvent solutions do not yield diffraction-quality crystals, mixtures of solvents may be tried. Multi-component solvent evaporation methods depend on the difference in the 

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