Mass spectrometric analysis of biopolymers
AssigneeGenencor International, Inc.
InventorsDavid A. Estell, Grant C. Ganshaw, Christian Paech, Sigrid Paech
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Abstract
The present invention makes use of unique tags of a specific biopolymer that can be exploited for determining the concentration the biopolymer in crude solutions. In preferred embodiments the biopolymer is either a protein or a polynucleotide. Particularly, the invention provides a method for the determination and quantitation of biomolecules in crude mixtures by way of a separation technique in combination with mass spectroscopy. In one general embodiment, a target biomolecule is selected for analysis and an analog thereof is generated. Peak area integration of the peptide pairs provides a direct measure for the amount of target protein in the crude solution.
Claims16
A method for determining the absolute quantity of a target biopolymer in a crude solution, comprising the steps of:
- (a) adding a known quantity of a calibrated analog of said target biopolymer to said crude solution, wherein said analog is the target polypeptide, a unique segment or a fragment thereof, and wherein one of said analog and said target biopolymer is isotope labeled;
- (b) treating the target biopolymer and analog with a fragmenting activity to generate a plurality of corresponding biopolymer and analog fragment pairs in said crude solution;
- (c) fractionating the crude solution produced in step (b) by a chromatopraphic technique to resolve said plurality of fragment pairs produced in step (b);
- (d) determining by mass spectrometric analysis of a fraction in step (c) the ratio of a selected target biopolymer to its corresponding analog;
- and
- (e) calculating, from said ratio and said known quantity of said analog, the absolute quantity of the target biopolymer in the mixture.
Dependencies are inferred from the claim text.