Use of self assembled magnetic beads for on-chip protein digestion.
The use of grafted trypsin magnetic beads in a microchip for performing protein digestion is described. The PDMS device uses strong magnets to create a magnetic field parallel to the flow with a strong gradient pointing through the center of the chip channel. This allows for the formation of a low-hydrodynamic resistance plug of magnetic trypsin beads that serves as a matrix for protein digestion. This device represents an inexpensive way of fabricating a multi open-tubular-like column with an appropriate pore size for proteins. Kinetics studies of the hydrolysis of a model peptide show a 100-fold increase in digestion speed obtained by the microsystem when compared to a batch wise system. This system also offers the great advantage of easy replacement, as the bead matrix is easily washed out and replaced. High performance and reproducibility for digesting recombinant human growth hormone are confirmed by analysing the digest products in both CE and MALDI-TOF MS. Similar sequence coverage (of about 44%) is obtained from MS analysis of products after 10 minutes on-chip and 4 h with soluble trypsin in bulk.
Slovakova
Marcela
M
Laboratoire Physicochimie-Curie, UMR/CNRS 168, Institut Curie, 75248 Paris Cedex 5, France.
Minc
Nicolas
N
Bilkova
Zuzana
Z
Smadja
Claire
C
Faigle
Wolfgang
W
Fütterer
Claus
C
Taverna
Myriam
M
Viovy
Jean-Louis
JL
D004129:Dimethylpolysiloxanes / Q000737:chemistry; D019382:Human Growth Hormone / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008280:Magnetics; D008838:Microcomputers; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation / Q000379:methods*; D009757:Nylons / Q000737:chemistry; D016062:Porosity; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D015203:Reproducibility of Results; D014357:Trypsin / Q000737:chemistry* / Q000378:metabolism
D004129:Dimethylpolysiloxanes; D009757:Nylons; D011506:Proteins; D011994:Recombinant Proteins; C085707:poly(dimethylsiloxane)-polyamide copolymer; D019382:Human Growth Hormone; D014357:Trypsin