Crystallization cassette for the growth and analysis of macromolecular crystals and an associated method
AssigneeUniversity of Alabama in Huntsville
InventorsJoseph D. Ng, Juan-Manuel Garcia-Ruiz, Jose A. Gavira-Gallardo, Mark Wells, Greg Jenkins
Life of this patent
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Abstract
The invention is a crystallization cassette and associated method for growing and analyzing macromolecular crystals in situ by X-ray crystallography. The cassette allows proteins (as well as other macromolecules) to be crystallized by the counter-diffusion method in a restricted geometry. Using this procedure, crystals can be adequately prepared for direct X-ray data analysis such that the protein's three-dimesional structure can be solved without crystal manipulation.
Claims62
A crystallization cassette comprising:
- a support member having a top, middle, and bottom portion;
- a housing member joined to the top portion;
- a stabilizing member joined to the middle portion having a plurality of capillary passageways;
- a precipitating reservoir member joined to the bottom portion, whereby the capillary tubes are in fluid communication with the precipitating reservoir;
- and
- a plurality of capillary tubes each having a proximal and distal end, wherein the capillary tubes' proximal ends are each joined to the housing member, and wherein each capillary tube extends downwardly from the housing member through a capillary passageway.
A crystallization cassette comprising:
- a shaft having a top, middle, and bottom portion;
- an upper member joined to the top portion of the shaft;
- a middle member proximate to the middle portion of the shaft including:
- a first channel for receipt of the shaft;
- and
- a plurality of capillary passageways that are substantially parallel to the shaft and extend longitudinally through the middle member;
- a plurality of capillary tubes each having a proximal and distal end, wherein the capillary tubes extend downwardly from the upper member, and wherein each capillary tube extends downwardly through a capillary passageways;
- and
- a lower member proximate to the lower portion of the shaft having
- a first surface having a plurality of cavities located on the surface, wherein each cavity is in alignment and corresponds to a capillary tube,
- a second surface, opposite the first surface, and
- a second channel for receipt of the shaft, whereby the capillary tubes are in fluid communication with the lower member.
A crystallization cassette comprising:
- a shaft having a top, middle, and bottom portion;
- a capillary housing member joined to the top portion of the shaft;
- a capillary stabilizing member joined to the middle portion of the shaft having a first channel that is adapted for slidably receiving the shaft, and a plurality of passageways that are parallel to the vertical axis of the shaft and extend longitudinally through the stabilizing member;
- a plurality of capillary tubes having a proximal and distal end, wherein the capillary tubes extend downwardly from the housing member, and wherein each capillary tube extends downwardly through one of the passageways;
- and
- a precipitation reservoir member joined to the lower portion of the shaft having
- a surface facing the stabilizing member,
- a plurality of cavities located on the surface, wherein each cavity is in alignment and corresponds to a capillary tube, and
- a second channel that is adapted for slidably receiving the shaft.
A cassette positioning system for rotating a crystallization cassette about an x-axis and y-axis comprising:
- an X-ray source;
- a turntable for rotating the cassette about the y-axis of rotation;
- an eccentricity correction stage disposed below the turntable;
- a capillary scan stage disposed under the eccentricity stage;
- and
- a cassette rotation stage for rotating a the cassette about the x-axis.
A method of growing and analyzing macromolecules comprising:
- providing a plurality of capillary tubes each having a first and second end;
- contacting the second end of each capillary tube to a solution having a solvated macromolecule therein;
- diffusing the solution into the capillary tubes;
- inserting the second end of each capillary tube into a corresponding cavity having one or more precipitating agents therein;
- allowing the one or more precipitating agents to counter-diffuse against the solution in each capillary tube;
- allowing macromolecule crystals to grow in the capillary tubes;
- and
- analyzing the crystals in situ via x-ray diffractometry.
Dependencies are inferred from the claim text.