In isoelectric focusing (IEF) separation, the resolution and focusing time is positively correlated to physical length of the separation channel. Thus, an increase in IEF resolution by means of increasing the length of the separation channel results in increase in separation time since each analyte must migrate a longer distance to be separated. The present disclosure provides a method for improved IEF resolution for a given amount of focusing time by using at least two isoelectric focusing sections of similar volume, but different diameters and linear lengths.
The present disclosure provides a method and apparatus for improvements of sample throughput in proteome analysis by mass spectrometry, by combining multiple non-overlapping isoelectric focusing separations. The method for performing an analysis of a plurality of protein samples, comprises: (a) Adding a proteolytic enzyme of a given specificity to a first protein sample to digest proteins to peptides; (b) Separating the peptides obtained in step (a) by isoelectric focusing; (c) Collecting those peptides which have their isoelectric point value within a first isoelectric point range; (d) Adding a proteolytic enzyme of a given specificity to a second protein sample to digest proteins to peptides; (e) Separating the peptides obtained in step (d) by isoelectric focusing; (f) Collecting those peptides which have their isoelectric point value within a second isoelectric point range, where said second isoelectric point range is different and non-overlapping compared to said first isoelectric point range; (g) Combining the peptides collected in steps (c) and (f) into a single sample and subjecting said sample to mass spectrometry analysis; (h) Deconvoluting signals/data obtained from the mass spectrometry analysis by calculating the isoelectric point of each peptide, and assigning a peptide to the first protein sample if its isoelectric point value matches the isoelectric point range selected in step (c) or to the second protein sample if its isoelectric point value matches the isoelectric point range selected in step (f); and (i) Obtaining quantitative information for proteins of each sample according to magnitude of the signal obtained from each peptide.
Methods and apparatus are provided that facilitate drug screening by using a multi-dimensional ligand multiplexing in combination with hydrogen deuterium exchange mass spectrometry (HDX MS). The present invention is based on ligand convolution, binding detection assay and subsequent deconvolution via signal processing. The method uses a plurality of wells arranged into rows and columns, where each well contains at least one unique ligand. Ligand convolution is performed by generating a pooled samples from each column and row. Each pooled sample is mixed with the target protein and subjected to HDX MS. The invention provides the means to deconvolute the signals in order to individualize a positive binding signal. The apparatus provides the means to perform the method and the use of a plurality of parallelized valves, columns and pumps to improve sample throughput. Therefore, a significant increase in the throughput of HDX MS is provided.
The present invention relates to a method for enriching and/or separating and/or immobilizing an analyte of interest comprising bringing an analyte of interest into contact with a derivatizing agent; incubating said analyte with said derivatizing agent, thereby incorporating a sulphonic acid group or an analogue thereof into the molecular structure of the analyte of interest; bringing the analyte of interest into contact with a molecularly imprinted polymer with selective affinity for a sulphonic acid group or an analogue thereof; and enriching and/or separating and/or immobilizing the analyte of interest by use of the molecularly imprinted polymer. Further disclosed is a kit comprising a derivatizing agent, which contains a sulphonic group or an analogue thereof and a reactive group for creating a covalent bond between said derivatizing agent and an analyte of interest, and a molecularly imprinted polymer with selective affinity for a sulphonic acid group or an analogue thereof.
B01D 15/38 - Selective adsorption, e.g. chromatography characterised by the separation mechanism involving specific interaction not covered by one or more of groups , e.g. affinity, ligand exchange or chiral chromatography
Apparatus and methods to perform hydrogen/deuterium exchange using semipermeable membranes are described. The system has two channels separated by a semipermeable membrane. One channel comprises a flow carrying the analyte of interest, and the second channel comprises a solution comprising a deuterated solvent (e.g. deuterium oxide). The system does not require an external electric field gradient across the membrane to perform the hydrogen-deuterium exchange procedure. The present invention facilitates sample and reagent handling as well as simplifies manufacture of devices and/or instrumentation related to deuterium exchange. Further described is a chemical analysation device for analysing chemical compositions and/or compounds, and a method for analysing chemical compounds and a computer program product for inducing a computer to perform steps in the method. Also described is a method for analyzing interactions between analytes and charged molecules, and calculating binding coefficients of the analytes with respect to the charged molecules.
H01J 49/04 - Arrangements for introducing or extracting samples to be analysed, e.g. vacuum locksArrangements for external adjustment of electron- or ion-optical components
C01B 4/00 - Hydrogen isotopesInorganic compounds thereof prepared by isotope exchange, e.g. NH3 + D2 → NH2D + HD
C01B 5/02 - Heavy waterPreparation by chemical reaction of hydrogen isotopes or their compounds, e.g. 4ND3+7O2→ 4NO2+6D2O, 2D2+O2→ 2D2O
G01N 33/68 - Chemical analysis of biological material, e.g. blood, urineTesting involving biospecific ligand binding methodsImmunological testing involving proteins, peptides or amino acids
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