title: Digital Microfluidics for Immunoprecipitation.
abstract: Immunoprecipitation (IP) is a common method for isolating a targeted protein from a complex sample such as blood, serum, or cell lysate. In particular, IP is often used as the primary means of target purification for the analysis by mass spectrometry of novel biologically derived pharmaceuticals, with particular utility for the identification of molecules bound to a protein target. Unfortunately, IP is a labor-intensive technique, is difficult to perform in parallel, and has limited options for automation. Furthermore, the technique is typically limited to large sample volumes, making the application of IP cleanup to precious samples nearly impossible. In recognition of these challenges, we introduce a method for performing microscale IP using magnetic particles and digital microfluidics (DMF-IP). The new method allows for 80% recovery of model proteins from approximately microliter volumes of serum in a sample-to-answer run time of approximately 25 min. Uniquely, analytes are eluted from these small samples in a format compatible with direct analysis by mass spectrometry. To extend the technique to be useful for large samples, we also developed a macro-to-microscale interface called preconcentration using liquid intake by paper (P-CLIP). This technique allows for efficient analysis of samples >100× larger than are typically processed on microfluidic devices. As described herein, DMF-IP and P-CLIP-DMF-IP are rapid, automated, and multiplexed methods that have the potential to reduce the time and effort required for IP sample preparations with applications in the fields of pharmacy, biomarker discovery, and protein biology.
authors.affiliation: Department of Chemistry, University of Toronto , 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.
authors.forename: Brendon
authors.initials: B
authors.lastname: Seale
authors.affiliation: Department of Chemistry, University of Toronto , 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.
authors.forename: Charis
authors.initials: C
authors.lastname: Lam
authors.affiliation: Department of Chemistry, University of Toronto , 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.|Donnelly Centre for Cellular and Biomolecular Research , 160 College Street, Toronto, Ontario M5S 3E1, Canada.
authors.forename: Darius G
authors.initials: DG
authors.lastname: Rackus
authors.affiliation: Department of Chemistry, University of Toronto , 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.|Donnelly Centre for Cellular and Biomolecular Research , 160 College Street, Toronto, Ontario M5S 3E1, Canada.
authors.forename: M Dean
authors.initials: MD
authors.lastname: Chamberlain
authors.affiliation: SCIEX , 71 Four Valley Drive, Concord, Ontario L4K 4V8, Canada.
authors.forename: Chang
authors.initials: C
authors.lastname: Liu
authors.affiliation: Department of Chemistry, University of Toronto , 80 St. George Street, Toronto, Ontario M5S 3H6, Canada.|Donnelly Centre for Cellular and Biomolecular Research , 160 College Street, Toronto, Ontario M5S 3E1, Canada.|Institute of Biomaterials and Biomedical Engineering, University of Toronto , 164 College Street, Toronto, Ontario M5S 3G9, Canada.
authors.forename: Aaron R
authors.initials: AR
authors.lastname: Wheeler