177 results about "Isoelectric focusing" patented technology

The invention relates generally to compositions and methods for altering the isoelectric point of an antibody, and in some cases, resulting in improved plasma pharmacokinetics, e.g. increased serum half-life in vivo.

An apparatus and method are disclosed for the precise selection and extraction of a selected analyte in a focused zone produced by isoelectric focusing performed in micro-channels. A cross-channel microfluidic device comprises a sample mixture introduction and separation channel and an extraction channel, which are in fluid communication with each other at a point of intersection. Means are provided for selectively moving the pattern of separated zones following cIEF to the intersection point, and means are provided for applying an extraction pressure to direct a single zone containing a selected analyte into and then out of the extraction channel for collection.

The invention relates generally to compositions and methods for altering the isoelectric point of an antibody, and in some cases, resulting in improved plasma pharmacokinetics, e.g. increased serum half-life in vivo.

Improved apparatus is disclosed for carrying out axially illuminated laser induced fluorescence whole-column imaging detection in the capillary isoelectric focusing of proteins. The separation capillary was made of low refractive index Teflon conditioned with methylcellulose to reduce electroosmotic flow and a small amount of high refractive index organic solvent (glycerol) was added to the sample mixture. It was found that an axially directed laser excitation beam was propagated essentially with total internal reflection, so that minimum interference arose from stray light or from scattering light originating from the wall of the capillary. With the naturally fluorescent protein R-phycoerythrin, a concentration detection limit LOD 10⁻¹¹ M or mass LOD 10⁻¹⁷ Mo was obtained.

A separation module operates to fractionate or separate an analyte into fractions according to pI, i.e., pI bands, utilizing capillary isoelectric focusing (“CIEF”) within a first microchannel. The fractions are stacked to form plugs, the number of which is determined by a number of parallel second microchannels integrally connected to the first microchannel, into which the fractions are directed according to the buffer characteristics found in each of the individual microchannels. Within the microchannels the plugs are separated into proteins according to a different chemical property, i.e., “m/z,” utilizing capillary electrophoresis (“CE”).

A method and apparatus are provided for performing capillary isoelectric focusing followed by mobilization of the focused zones by induced hydrodynamic flow or chemical mobilization. These two dimensions of separation are integrated with real-time whole-channel electrophoresis detection and automatic sample injection to achieve a separation resolution superior to that obtainable using known orthogonal capillary two dimensional arrangements.

Disclosed herein is an apparatus for the separation of proteins, comprising a capillary isoelectric focusing unit (2) for primarily separating protein samples on the basis of pi; and a hollow fiber flow field flow fractionation unit (4), connected to one side of the capillary isoelectric focusing unit (2), for secondarily separating the protein samples. The apparatus allows proteins to be separated on the basis of pi and molecular weight without denaturation and can further be applied for the identification of proteins in conjunction with nanoflow liquid chromatography-electrospray ionization-tandem mass spectrometry after enzymatic digestion of the protein fractions.

The invention provides a novel solution isoelectric focusing device and method that can reproducibly fractionate charged molecules into well-defined pools. This approach can be applied to mixtures of charged molecules, such as eukaryotic proteome samples where reproducible resolution and quantitation of greater than 10,000 protein components is feasible.

The microflow control chip for protein analysis features that it consists of four basic units connected successively with the sample outlet of the upper basic unit being connected to the sample inletof the next basic unit. They are the first protein isoelectric focusing separation unit; the second protein screening electrophoresis unit with organic polymer selected from cellulose and agarose asscreening medium; the third protein proteinase enzymolyzing unit with specific proteinase bonded to the inner wall or specific proteinase stuffing filled; and the fourth interface unit to convey the enzymolysis product of protein to mass spectral analysis. Using the chip of the present invention in protein analysis can obtain great amount of information in short time and through simple operation.

Methods and apparatus are provided to resolve analytes within a fluid path using isoelectric focusing, gel electrophoresis, or other separation means. Materials within the fluid path that are compatible with these separation means are used to attach resolved analytes to the wall of the fluid path. Attachment results from a triggerable event such as photoactivation, thermal activation, or chemical activation. In accordance with a further aspect of the present invention, the material in the capillary may also be disrupted, by either the triggerable event or a subsequent event such as melting or photocleavage. Thus, an open lumen or porous structure may be created within the fluid path, allowing unbound analyte materials to be washed from the fluid path, and detection agents to be washed into the fluid path. The separation-compatible materials may be polymerizable monomers, gels, entangled polymers or other materials.

A self-assembled engineered lattice of nanometer-scale silica particles, or other suitable particles generally resembling regularly-sized spheres, is configured in a separation bed for electrophoresis, isoelectric focusing, chromatography, or other voltage-induced separation of analytes. After separation, the analytes are immobilized on the separation bed and then ionized using matrix-assisted laser desorption/ionization (MALDI) for use with a mass spectrometer. The nanoparticles can be coated with polymers that activate to immobilize the analytes or assist with MALDI. The separation can occur in two dimensions.

The invention discloses a clam bioactive peptide and its extraction method and use. The clam bioactive peptide has an N-tail end sequence of TRGIDEIQNT GGGGTRR, an isoelectric point of 8.79 and molecular weight of 18KDa. The clam bioactive peptide can promote macrophage phagocytosis activity and splenic lymphocyte propagation capability of an immunization system, has pharmacological effects of improving immunity, can be used for preparation of a drug or a health food for improving immunity and has potential of exploitation of novel immunopotentiation drugs.

Provided herein are IEF focusing methods for determining the number of drug molecules present in at least one antibody-drug conjugate (ADC) species subpopulation. In one embodiment, the method comprises performing free solution isoelectric focusing on a sample comprising at least one ADC species, to obtain a focused sample. The absorbance of the sample at two different wavelengths is then measured, for example, over a range of pI values. Absorbance values at the two different wavelengths are compared at at least one corresponding pI value, where the at least one corresponding pI value is the pI of the ADC subpopulation. The number of drug molecules in the at least one ADC species subpopulation is then determined based on the comparison. The methods provided herein can also be used to determine the number of specific binding pair members bound to its target specific binding pair member.

The invention relates to a method for separating, enriching and identifying macromolecular weight protein, belonging to the field of biochemical analysis. In the invention, a mixed gel of low-melting point agarose and polyacrylamide is used as an electrophoresis supporting medium, and an analysis path of gel slab electrophoresis combined with efficient liquid-phase chromatography - electrical spraying mass spectrum identification and another analysis path of solid-phase gradient pH dry adhesive tape isoelectro-focusing combined with gel slab electrophoresis separation - matrix auxiliary laser desorption ionization mass spectrum identification are collectively used to separate and identify the macromolecular weight protein. The gel provided by the invention has the advantages of big aperture, high mechanical strength, high resolution ratio, good repeatability and good mass spectrum compatibility; the macromolecular weight separation scope can be adjusted based on different component proportions; and protein with the molecular weight interval above 100kDa can be selectively separated with repeatability RSD up to 7.6%, thereby obviously improving the identification ratio. The method is an effective tool for separating the macromolecular weight protein, which can be applied to proteomics study field.

The invention discloses a dielectrophoresis method for proteins of malus plants. The steps of the method are as follows: 1) plant materials are ground in liquid nitrogen and are added with an acetone solution which is pre-cooled at a temperature of 20 DEG C below zero and contains a trichloroacetic acid with the concentration of 10g/100ml and beta-mercaptoethanol with the concentration of 0.07ml/100ml and continuously ground to be the homogenate, and the homogenate stands at a temperature of 20 DEG C below zero for 1 hour and centrifuged by the rotary speed of 12000rpm for 15 minutes, and the precipitation is kept; 2) the precipitation is added with an acetone solution which contains the beta-mercaptoethanol with the concentration of 0.07ml/100ml and the volume of which is 4 times of the volume of the precipitation, and the mixed solution is evenly mixed and stands at a temperature of 20 DEG C below zero for 1 hour, and centrifuged by the rotary speed of 12000rpm for 15 minutes, and the precipitation is kept; 3) the step 2 is repeated for 6 times, and total proteins are obtained; 4) total proteins obtained from the step 3 undergo the electric focusing electrophoresis, and the sample volume of proteins is of 300ug/170mm adhesive tape, and the polyacrylamide gel electrophoresis is conducted, and the silver dyeing is conducted after the electrophoresis.

A plurality of particles of from about 5 nm to 100 μm possessing predetermined isoelectric points in the pH range from about 2.5 to 11 is used in a method of detection of a plurality of analytes, wherein the isoelectric particles of each isoelectric point further contain a label and a member of a binding pair capable of interacting with a selected analyte. The particles that formed specific binding pairs are recovered and separated by isoelectric focusing, followed by the detection of the labels associated with the particles. A flow cytometer may be used as a detector of the isoelectric particles.

The invention discloses the main sensitization protein of humulus pollen which can be separated and obtained from the humulus pollen and combined with over 89 percent of serological specificity IgE of patients allergic to humulus pollen. The apparent molecular weight of the protein measured by SDS-PAGE is about 12kDa, the isoelectric point is 4.7 and the N-terminal sequence is NH4<+>-MDNPFENGMKA-COO<->. The separation and purification of the protein lay the foundation for realizing the standization of the allergenic preparations of the protein.

Disclosed is an agent for digesting a protein highly resistant to denaturation and degradation, comprising as an active ingredient an enzyme exhibiting an activity of digesting a protein highly resistant to denaturation and degradation and having the following properties: (a) activity and substrate specificity: hydrolyzing a peptide bond of a protein highly resistant to denaturation and degradation; (b) molecular weight: 31,000 (determined by SDS-polyacrylamide gel electrophoresis using a homogeneous gel having a gel concentration of 12%); (c) isoelectric point: pI 9.3 (determined by polyacrylamide gel isoelectric focusing electrophoresis); (d) optimum pH: pH 9.0 to 10.0; and (e) optimum temperature for activity: 60 to 70° C.