title: Preconcentration and detection of the phosphorylated forms of cardiac troponin I in a cascade microchip by cationic isotachophoresis.
abstract: This paper describes the detection of a cardiac biomarker, cardiac troponin I (cTnI), spiked into depleted human serum using cationic isotachophoresis (ITP) in a 3.9 cm long poly(methyl methacrylate) (PMMA) microfluidic channel. The microfluidic chip incorporates a 100× cross-sectional area reduction, including a 10× depth reduction and a 10× width reduction, to increase sensitivity during ITP. The cross-sectional area reductions in combination with ITP allowed visualization of lower concentrations of fluorescently labeled cTnI. ITP was performed in both "peak mode" and "plateau mode" and the final concentrations obtained were linear with initial cTnI concentration. We were able to detect and quantify cTnI at initial concentrations as low as 46 ng mL(-1) in the presence of human serum proteins and obtain cTnI concentrations factors as high as ~ 9000. In addition, preliminary ITP experiments including both labeled cTnI and labeled protein kinase A (PKA) phosphorylated cTnI were performed to visualize ITP migration of different phosphorylated forms of cTnI. The different phosphorylated states of cTnI formed distinct ITP zones between the leading and terminating electrolytes. To our knowledge, this is the first attempt at using ITP in a cascade microchip to quantify cTnI in human serum and detect different phosphorylated forms.
authors.affiliation: Gene and Linda Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-2710, USA.
authors.forename: Danny
authors.initials: D
authors.lastname: Bottenus
authors.forename: Mohammad Robiul
authors.initials: MR
authors.lastname: Hossan
authors.forename: Yexin
authors.initials: Y
authors.lastname: Ouyang
authors.forename: Wen-Ji
authors.initials: WJ
authors.lastname: Dong
authors.forename: Prashanta
authors.initials: P
authors.lastname: Dutta
authors.forename: Cornelius F
authors.initials: CF
authors.lastname: Ivory
mesh_terms: D053000:Analytic Sample Preparation Methods / Q000379:methods*; D015415:Biomarkers / Q000032:analysis / Q000097:blood; D006801:Humans; D057625:Isotachophoresis / Q000379:methods*; D046208:Microchip Analytical Procedures / Q000379:methods*; D009206:Myocardium; D010750:Phosphoproteins / Q000032:analysis / Q000097:blood / Q000302:isolation & purification; D010766:Phosphorylation; D019904:Polymethyl Methacrylate / Q000737:chemistry; D019210:Troponin I / Q000032:analysis* / Q000097:blood / Q000302:isolation & purification*
chemical_list: D015415:Biomarkers; D010750:Phosphoproteins; D019210:Troponin I; D019904:Polymethyl Methacrylate