Casein probe-based fast plasmin determination in the picomolar range by an ultra-high frequency acoustic wave biosensor

SENSORS AND ACTUATORS B-CHEMICAL

卷:275

Article

Romanszki, Lorand^1,2  Tatarko, Marek^1,3  Jiao, Mengchi¹  Keresztes, Zsofia²  Hianik, Tibor³  Thompson, Michael¹ 

[1] Univ Toronto, Dept Chem, Lash Miller Labs, 80 St George St, Toronto, ON M5S 3H6, Canada

[2] Hungarian Acad Scie, Res Ctr Nat Sci, Inst Mat & Environm Chem, Funct Interfaces Res Grp, Magyar Tudosok Korutja 2, H-1117 Budapest, Hungary

[3] Comenius Univ, Fac Math Phys & Informat, Dept Nucl Phys & Biophys, Mlynska Dolina F1, Bratislava 84248, Slovakia

关键词: Plasmin;    beta-casein;    Electromagnetic piezoelectric acoustic sensor;    Inverse Michaelis-Menten equation;   

DOI  :  10.1016/j.snb.2018.08.025

来源: Elsevier

【 摘 要 】

Detection of residual plasmin activity in milk represents a difficult challenge for the dairy industry. Conventional methods are either too expensive or incapable of providing enough data from UHT treated milk. Acoustic wave-based biosensors operated in the thickness shear mode (TSM) showed potential for the detection of proteolysis of beta-casein, a milk protein by protease plasmin. An ultra-high frequency device, the electromagnetic piezoelectric acoustic sensor (EMPAS), designed to enhance the sensitivity of TSM, was tested for detection of plasmin at low concentrations. 5-casein layers immobilised on the hydrophilic or hydrophobized surfaces of EMPAS quartz discs served as substrate for the enzyme. In contrast with conventional TSM devices, the shearing oscillations in EMPAS are induced contactless, by a magnetic coil located 30 mu m below the quartz crystal. This configuration allows the registration of unusually high harmonics (up to the 49th-53rd), thus enhancing the sensitivity of detection. On both surface types, the adsorbed beta-casein mass and the stability of the layer was compared, with the result that hydrophobic surfaces provide superior conditions for immobilisation than the hydrophilic case. Consequent proteolysis measurements of these substrate layers were carried out in a broad plasmin concentration range (32 pM-10 nM) in flow mode. Initial reaction rates measured at different enzyme concentrations have been used to construct a calibration curve based on an inverse Michaelis-Menten type equation. The sensitivity of the EMPAS allowed measurements of as low as 32 pM concentration of plasmin, reaching (and often exceeding) levels comparable to state of the art techniques like ELISA. The presented method however, unlike ELISA, is effective on a timescale of minutes.

【 授权许可】

Free