کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
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868689 | 909812 | 2009 | 6 صفحه PDF | دانلود رایگان |
This paper discusses the design, microfabrication and use of an electrochemical biosensor based on a polymer substrate for cost effectiveness and disposability. As model analyte, amplified hsp70 mRNA from Cryptosporidium parvum was chosen. Microfluidic channels were fabricated in poly(methyl methacrylate) (PMMA) using hot embossing with a copper master. The electrochemical transducer, an interdigitated ultramicroelectrode array (IDUA) was also realized directly on the PMMA surface. First, the unstructured PMMA surface was UV functionalized. An 8 min UV treatment resulted in a carboxylic acid density of approximately 8 nmol/cm2 on the PMMA surface. The surface carboxylic acid groups were then conjugated to cystamine using water-soluble carbodiimide chemistry. Gold (200 nm) was then evaporated onto the thiol-functionalized surface. Using standard photolithography techniques, the IDUA containing 10 μm wide electrodes with 5 μm gaps was then formed followed by a gold etch. The PMMA surface containing the microchannel was subsequently bonded to the PMMA surface containing the IDUA using UV-assisted thermal bonding. The additional UV treatment also served to decrease the water contact angle of the surface from 62.5° ± 0.7° to 48.4° ± 0.2° thus, aiding with the capillary flow in the device. The hsp70 mRNA was isolated from C. parvum oocysts and amplified using nucleic acid sequence-based amplification (NASBA). The amplicon was detected in a sandwich hybridization assay with capture probe-coated superparamagnetic beads and reporter probe-tagged liposomes. The liposomes entrapped potassium ferro/ferrihexacyanide to enable amperometric quantification of the amplicon on the IDUA. Amplified mRNA from only 1 oocyst was detectable with this PMMA biosensor. The final detection device measured approximately 10 mm × 40 mm × 3 mm and contained two detection channels for dual analyses.
Journal: Biosensors and Bioelectronics - Volume 24, Issue 8, 15 April 2009, Pages 2428–2433