156 results about "Protein crystallization" patented technology

Protein crystallization droplet actuators, systems and methods are provided. According to one embodiment, a droplet actuator for providing an array of crystallization conditions is provided and includes: (a) two or more processing reservoirs; (b) two or more dispensing reservoirs collectively comprising two or more crystallization reagents; and (c) a port for introducing a sample for crystallization analysis. Systems including the droplet actuator, methods of providing an array of crystallization conditions, and methods of identifying crystallization conditions are also provided.

Protein crystallization screening and optimization droplet actuators, systems and methods are provided. According to one embodiment, a screening droplet actuator is provided and includes: (a) a port for introduction of one or more crystallization reagents and/or one or more protein solutions; and (b) a substrate including: (i) an array of two or more mixing wells; and (ii) electric field mediated microfluidics for moving droplets comprising the crystallization reagents and protein solutions into the mixing wells. Optimization droplet actuators, systems including screening droplet actuators, methods of screening protein crystallization conditions, and methods of testing conditions for growing a crystal are also provided.

The use of microfluidic structures enables high throughput screening of protein crystallization. In one embodiment, an integrated combinatoric mixing chip allows for precise metering of reagents to rapidly create a large number of potential crystallization conditions, with possible crystal formations observed on chip. In an alternative embodiment, the microfluidic structures may be utilized to explore phase space conditions of a particular protein crystallizing agent combination, thereby identifying promising conditions and allowing for subsequent focused attempts to obtain crystal growth.

Embodiments of the present invention relate to methods and apparatuses for the discretization and manipulation of sample volumes that is simple, robust, and versatile. It is a fluidic device that partitions a sample by exploiting the interplay between fluidic forces, interfacial tension, channel geometry, and the final stability of the formed droplet and/or discretized volume. These compartmentalized volumes allow for isolation of samples and partitioning into a localized array that can subsequently be manipulated and analyzed. The isolation of the discretized volumes along with the device's inherent portability render our invention versatile for use in many areas, including but not limited to PCR, digital PCR, biological assays for diagnostics and prognostics, cancer diagnosis and prognosis, high throughput screening, single molecule and single cell reactions or assays, the study crystallization and other statistical processes, protein crystallization, drug screening, environmental testing, and the coupling to a wide range of analytical detection techniques for biomedical assays and measurements. The minimal fluid interconnects and simple flow geometry makes the device easy to use and implement, economical to fabricate and operate, and robust in its operations.

A method and devices for rapid cooling of small biological samples by plunging them in a cryogenic liquid, such as liquid nitrogen, or contacting them with a cryogenic metal surface, reduce or eliminate the cold gas layer that forms above the liquid cryogens or cryogenic surfaces, producing an abrupt transition from ambient (e.g., room) temperature to the cryogen temperature as the sample enters the liquid or contacts the surface. To reduce or eliminate the effects of the cold gas layer, a flow of warm dry gas can be directed along the plunge path, for example. By removing this cold gas layer, cooling times for a 10 micron sample (the size of single cells and the smallest protein crystals now used protein crystallography) will decrease to ˜0.001 s.

Crystallization Photoresist (PR) apparatus and methods which allow for fast screening and determination of protein crystallization conditions with small protein quantities and rapid crystallization. The apparatus comprise a first region comprising a first nucleation catalyst material and a second region comprising a second nucleation catalyst material, with the first and second regions positioned adjacent to each other and configured to support at least one crystal, and with the first region having a variation in a nucleation property of the first nucleation catalyst material in the first region. The crystal may be supported at an interface of the adjacent regions. The methods comprise providing a first region of a first nucleation catalyst material and a second region of a second nucleation catalyst material adjacent said first region, with the first region having a variation in a nucleation property of the first nucleation catalyst material, exposing the first and second regions to a solution of a selected molecule, and growing at least one crystal of the molecule in association with the first and second regions.

The invention provides a semi-contact under-oil continuous droplet sample applying and liquid adding method, which is suitable for sequentially operating a droplet array system. According to the method, the distance between the pointed end of a capillary tube sample applying needle and the lower surface of a micro-hole (or a generated droplet) is controlled accurately, and the affinity or interface tension interaction between the droplet and the surface (or the generated droplet) is utilized, so that rapid and reliable continuous droplet sample applying or continuous liquid adding is realized, and the problem of cross infection during sample applying is solved effectively. The method is suitable for biochemical analysis screening researches such as high-flux medicament screening, protein crystallization condition screening, enzyme kinetics research and drug toxicity determination.

Biomolecule arrays on a substrate are described which contain a plurality of biomolecules, such as coding nucleic acids and/or isolated polypeptides, at a plurality of discrete, isolated, locations. The arrays can be used, for example, in high throughput genomics and proteomics for specific uses including, but not limited molecular diagnostics for early detection, diagnosis, treatment, prognosis, monitoring clinical response, and protein crystallography.

The present invention relates to methods and apparatus for promoting rapid formation of biomolecule crystals from a solution of biomolecules, preferably proteins, wherein the protein solution undergoes rapid concentration according to its isoelectric point in an electric field. Protein crystallization according to the methods of the present invention takes place within a period of hours or less.

The invention discloses a gradient micro-droplet array forming method based on sequential injection and microfluidic technology. The gradient micro-droplet array forming method comprises following steps: step 1, one of a diluent and a sample I is taken using a capillary tube; step 2, the other one of the diluent and the sample I is taken using the capillary tube, and direct contact of the diluent with the sample I is ensured so as to form a sample zone belt with axial concentration gradient; and step 3, the sample zone belt is injected into certain areas of a microporous array chip via the capillary tube so as to obtain sample droplet array with different concentration on the microporous array chip. Advantages of the gradient micro-droplet array forming method are that: concentration gradient generation throughput is high, concentration information is abundant, automatic degree is high; sample consumption is low, and devices and principles are simple. The gradient micro-droplet array forming method can be used for biochemical analysis and screening such as high throughput drug screening, compound toxicity determination, protein crystallization condition screening, catalyst screening, and enzyme dynamic analysis.

The invention relates to an expression, purification, crystallization method, structure elucidation and the field of application technology of forulic acid decarboxylase, which belongs to the field of molecular biology and microbiology application. The invention obtains the coding area sequence of the key enzyme forulic acid decarboxylase which decomposes forulic acid to generate 4-vinyl guaiacol in Enterobacter sp.Px6-4 by the gene clone method, expresses the enzyme in a large amount in the colon bacillus ( E.coli BL21 ) by the heterogenous expressing technology, obtains the forulic acid decarboxylase pure products by the purification method and the forulic acid decarboxylase protein crystallization by the hanging-drop crystallization process, and grasps the structure and active centre of the enzyme. The invention can obtain a large amount of highly active forulic acid decarboxylase, and provide theoretical guidance for reconstructing engineering bacteria and improving the output of 4-vinyl guaiacol by analyzing the active centre of the enzyme, thus having good potential application.

The present disclosure provides an effective method for the crystallization of proteins at high pressure. A preferential excluding agent, and optionally other reagents may be incorporated into the method. The method is applicable to substantially all proteins.

The present invention relates to a method of precipitating protein crystals from a protein-containing sample. The present invention also relates to a novel microarray and a novel device for screening for protein crystallization condition. Furthermore, the present invention relates to a method of conveniently and quickly screening for protein crystallization conditions using the microarray or the device having highly integrated and held crystallization conditions even with an extremely small quantity of a sample.

An automated biological sample analysis system and method for use in incubating and analyzing multiple samples for protein crystallization. A temperature controlled cabinet houses sample storage, sample transport, and sample imaging systems. The operation of the system is automated and can be controlled by software, which can be reconfigured remotely. An array of storage shelves includes multiple shelf columns arranged around a core. Multiple banks of removable shelves arranged as magazines are accessed through a door on the cabinet. Each shelf stores a multi-well plate and different sizes can be stored in different shelves. The core houses a sample transport system that includes a multi-axis robot that rotates about a vertical axis to access the shelves in the shelf array. The transport system retrieves and replaces the multi-well plates in the shelves and can move plates from the shelves to an imaging system where each sample can be automatically imaged.

A high-density high-throughput microplate and methods for simultaneously screening a plurality of protein crystallization solutions and for producing diffraction quality protein crystals in a vapor-diffusion environment are disclosed. The microplate has defined side-by-side paired chambers of equal size, wherein the side-by-side paired chambers have a maximum volume of about 8 μl, and wherein the paired chambers have a vapor channel, therein providing vapor exchange between the side-by-side paired chambers. The microplate further includes a membrane to seal the surface of the microplate. The microplate is adapted to receive a crystallization solution in one of the side-by-side paired chambers and a protein solution in the other of the side-by-side paired chambers, wherein the protein solution and the crystallization solution interact via a vapor diffusion process, which enables the formation of protein crystals within the chamber that contains the protein solution.

The use of microfluidic structures enables high throughput screening of protein crystallization. In one embodiment, an integrated combinatoric mixing chip allows for precise metering of reagents to rapidly create a large number of potential crystallization conditions, with possible crystal formations observed on chip. In an alternative embodiment, the microfluidic structures may be utilized to explore phase space conditions of a particular protein crystallizing agent combination, thereby identifying promising conditions and allowing for subsequent focused attempts to obtain crystal growth.

The invention discloses a protein crystallization screening 96 condition kit using polyethylene glycol as a precipitator and aims to solve the technical problem existing in the conventional 96 condition screening kit that the success rate of protein crystallization condition screening is low. The PEG adopted in the technical scheme is the top eight types PEG which are most frequently used according to the statistical data of a BMCD database rather than selected randomly, so that the crystallization success rate of biological macromolecules is obviously improved. For example, the success rate of canavaline is increased from 4.2 percent in background technology to 81.2 percent, the success rate of lysozyme is increased from 29.2 percent in background technology to 46.9 percent, the success rate of protease K is increased from 58.3 percent in background technology to 66.7 percent, and the success rate of catalase is increased from 15.6 percent in background technology to 30.2 percent. In addition, the kit uses the PEG as the precipitator, a few types of reagents are used, and compared with the background technology kit, the kit of the invention is lower in cost.

Herein is described and optimum solubility screen in which a panel of buffers and many additives are provided in order to obtain the most homogeneous and monodisperse protein condition for protein crystallization. The present methods are useful for proteins that aggregate and cannot be concentrated prior to setting up crystallization screens. A broad range of buffers is intended for use in this screen. A high-throughput method using the hanging-drop method and vapor diffusion equilibrium and a panel of twenty-four buffers is further provided. After monitoring precipitation, the conditions leading to clear drops are selected for evaluation, preferably dynamic light scattering (DLS) characterization. If the DLS results are not optimal, a series of additives are tested in the presence of the best buffer selected from the initial screen and again DLS is used to determine the best condition. Using the present methods, 14 poorly behaving proteins have been screened, resulting in 11 of the proteins having highly improved DLS results allowing concentration of the proteins, and 9 were crystallized.

The present invention provides a method for selecting a buffer solution for protein crystallization. The method comprises: preparing a uniformly distributed buffer solution to dissolve protein; mixing a crystallization reagent and the protein solution or mixing a crystallization reagent, the buffer solution and a protein sample to prepare a protein crystallization solution; and standing in a sealed crystallization plate until crystallization. According to the present invention, the appropriate protein crystallization buffer solution pH value condition is selected, such that protein solubility is changed in a wide range, and protein crystal nucleation probability is increased so as to increase crystallization efficiency.

The invention provides a method for preparing recombinant human insulin crystal, and belongs to the technical field of protein crystallization. According to the method, zinc ions are subjected to three times of crystallization, the obtained insulin crystal is large in particle size and good in homogeneity, phenols or organic solvents used in normal methods are not used in the method, so that residues of harmful substances in the product are avoided and the quality of the insulin crystal is improved; moreover, the method is stable in preparation process and beneficial to industrial production.