Heat shock protein 70 (HSP70) is an important health related biomarker, being implicated as an early stage cancer marker and as an indicator of cardiac health. It also has important implications in wildlife environmental monitoring, as its levels can be affected by food deprivation, elevated temperatures, and pollution. Therefore, the use of HSP70 as a biomarker is highly desirable, yet the current methods of quantifying HSP70 are time consuming, expensive, and require dedicated labs. In order to facilitate widespread use of the HSP70 biomarker, a quantification tool that can be used at the point-of-care is needed. This implies the development of a simple and inexpensive HSP70 biosensing technique that is highly sensitive and selective. Therefore, in this work a label-free HSP70 biosensor has been designed based on the optical properties of gold nanoparticles (NPs). Gold NPs exhibit a large absorbance peak in the visible spectrum due to localized surface plasmon resonance (LSPR). The peak position is dependent on the local refractive index, which can be employed as a biosensor by selectively capturing the target analyte to the NP surface. To design an LSPR HSP70 sensor, optical and fluidic simulations were developed to determine optimal NP geometries and microchannel dimensions. The results showed optimal response when using 100nmx5nm gold nanotriangles inside of a 100μmx100μm microchannel. Simulations of the sensor performance showed HSP70 detection from 0.92-4000ng/ml with a resolution of 1.1ng/ml, all of which satisfied the design requirements. An LSPR sensor was experimentally tested at the benchtop scale to prove the concept. Gold NPs were fabricated by electron beam lithography and enclosed in a polymer flow cell. For initial testing of the LSPR sensor, the NPs were functionalized with biotin for selective capture of streptavidin. Streptavidin was detected in real time over the range 55-500,000ng/ml. The use of bovine serum albumin (BSA) was shown to be necessary to block non-specific binding sites to ensure a streptavidin-specific response. The LSPR sensor was then demonstrated to detect salmon HSP70 at 4600ng/ml using its synthetic antibody. Overall, these results demonstrate that LSPR can be used to realize an HSP70 biosensor suitable for point-of-care applications.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OWTU.10012/6818 |
| Date | 18 June 2012 |
| Creators | Denomme, Ryan |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Thesis or Dissertation |
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