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Alternative reservoirs in vapor diffusion experiments
Alternative reservoir solutions in vapor diffusion experiments

Setting up vapor diffusion crystallization experiments against different reservoir solutions may have a profound effect on the outcome of crystallization experiments. It has been demonstrated shown that one way to increase crystallization space is to set the same conditions over different reservoirs (Newman 2005). So, rather than screen more reagents, one can screen the same reagent set over two or three different reservoirs (such as sodium chloride, ammonium sulfate and polyethylene glycol 3305) in addition to the crystallization reagent. Using the screen reagent in four sets of drops and then using the reagent in addition to three alternative reservoirs expands the crystallization space as the path of equilibration and endpoint is unique for each reservoir solution.

Recommended reservoir solutions
1.25 - 2.0 M Sodium chloride
1.0 - 1.5 M Ammonium sulfate
30-50% Polyethylene glycol 3350
20% Polyethylene glycol 3350, 0.2 M sodium chloride

Replacement reservoir solutions (approximate)
0.5 M Lithium chloride = 35% PEG 4000
1.5-2.0 M Lithium chloride = 2.5 M Ammonium sulfate

Sodium chloride should be expected to act as a simple desiccant. Ammonium sulfate can desiccate and affect the pH of the drop through the gaseous NH3 which is in equilibrium with the NH4+ in solution. PEG 3350 should desiccate, although with different kinetics compared to the salts (Luft & DeTitta 1995).

Alternative reservoir solutions have also been referred to as common dehydrants, a generic reservoir, and desiccants. The use of a single, simple, concentrated solution for the reservoir in vapor diffusion crystallization experiments has been happening for decades (Dunlop & Hazes, 2005; Hempel, 1968; Luft et al., 1994; McPherson, 1992).

For 48 and 96 well crystallization plates set up manually, to speed delivery of the reservoir, the reservoir solution can be transferred to a multichannel pipetter basin (HR3-269) and delivered to the plate using a multichannel pipette. For automated experiments the reservoir solution can be transferred to a deep well block (HR3-105) or a multichannel pipetter basin (HR3-269).

References
Expanding screening space through the use of alternative reservoirs in vapor-diffusion experiments. Janet Newman. Acta Cryst. (2005). D61, 490-493.

Single Crystals of Transfer RNA from Formylmethionine and Phenyalanine Transfer RNA's.Arnold Hempel et al. Science, New Series, Vol. 162, No. 3860 (Dec. 20, 1968), 1384-1387.

A modified vapor-diffusion crystallization protocol that uses a common dehydrating agent. Dunlop & Hazes. Acta Cryst. (2005). D61, 1041-1048.

Luft et al. (1994). J. Appl. Cryst. 27, 443-452.

Luft & DeTitta. (1995). Acta Cryst. D51. 780-785.

McPherson. (1992). J. Cryst. Growth, 122, 161-167.
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