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Method and apparatus for performing peptide digestion on a microfluidic device

  • US 9,097,723 B2
  • Filed: 04/03/2006
  • Issued: 08/04/2015
  • Est. Priority Date: 04/01/2005
  • Status: Active Grant
First Claim
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1. A method of processing a protein sample in a microfluidic device, the method comprising the steps of:

  • providing a microfluidic device comprising a first fluid network that includes a sample reservoir, a waste reservoir, a plurality of parallel channels connecting the sample reservoir to the waste reservoir, the plurality of parallel channels defining a first region proximate the sample reservoir, a second region adjacent the first region, and a third region proximate the waste reservoir, and a plurality of reagent reservoirs that introduce reagents at various points along each of the plurality of parallel channels, wherein each of the reagent reservoirs is in fluid communication with at least two of the plurality of parallel channels;

    maintaining the first region at 50°

    C. and the third region at a temperature below 50°

    C. but above room temperature;

    placing immobilized trypsin in a portion of each of the plurality of channels in the third region;

    placing the protein sample into the sample reservoir;

    placing the reagents required to carry out reduction, alkylation, and quenching into the reagent reservoirs;

    applying a single negative pressure driving force to the waste reservoir that generates a continuous flow of fluid from the sample reservoir, through the plurality of channels, to the waste reservoir, wherein the flow sequentially exposes the protein sample to the reduction reagent in the first region, and the alkylation and quenching reagents in the second region before the protein sample flows through the immobilized trypsin in the third region; and

    collecting the fluid including the protein sample in the waste reservoir.

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