Microfluidic platforms for single-cell analyses
Abstract/Contents
- Abstract
- This dissertation describes several microfluidic platforms for single-cell analysis. They represent the continuing advance of analytical technologies to provide ever increasingly detailed information of cellular populations. Microfluidics offers several advantages for working with samples at the single-cell level, including the ability to manipulate nanoliter volumes and enhancement of interactions in confined volumes. In order to demonstrate the scientific utility of these new platforms, the devices are used to process and study various strains of cyanobacteria, which represent an important class of organism of interest to the environmental microbiology community. A microfluidic device that incorporates a membrane of conical nanopores is described and used to capture hundreds of single cells, one per pore. The conical nanopores are created using simple chemical etching of a polycarbonate membrane containing heavy-ion damage tracks. The geometries can be tuned to create nanopores capable of capturing single cyanobacteria cells based on size. Capture and observation of single cells, as well as size-based separation of a cellular mixture, is demonstrated. A second method for capture of single cells using a molecular-imprinted film integrated into a microfluidic device is also described. Films are imprinted with target strains of cyanobacteria and used to selectively capture single cells of the same strain as the imprint template. The imprints are shown to be selective even between different strains of the same genus of cyanobacteria. It is thought that this selectivity derives from diverse chemistries present on these organisms, allowing the imprints to prefer template-matching individuals. The separation efficiency of microfluidic molecularly imprinted films, as well as how that efficiency can be tuned with pH, is demonstrated. Microfluidic single-cell genetic amplification is extended to a new class of organisms through the development of chemical lysis protocols for cyanobacteria that remain sufficiently compatible with the amplification reaction. Previously, such methods were limited to organisms susceptible to simple alkaline lysis. Issues of contaminating genetic material and complications of the MDA reaction, such as coverage variance, are also characterized. The utility of the technique is demonstrated by amplifying a heterogeneous sequence of interest, CRIPSR, and sequencing it.
Description
| Type of resource | text |
|---|---|
| Form | electronic; electronic resource; remote |
| Extent | 1 online resource. |
| Publication date | 2012 |
| Issuance | monographic |
| Language | English |
Creators/Contributors
| Associated with | Hall, Eric William |
|---|---|
| Associated with | Stanford University, Department of Chemistry |
| Primary advisor | Zare, Richard N |
| Thesis advisor | Zare, Richard N |
| Thesis advisor | Andersen, Hans, 1941- |
| Thesis advisor | Fayer, Michael D |
| Advisor | Andersen, Hans, 1941- |
| Advisor | Fayer, Michael D |
Subjects
| Genre | Theses |
|---|
Bibliographic information
| Statement of responsibility | Eric Hall. |
|---|---|
| Note | Submitted to the Department of Chemistry. |
| Thesis | Thesis (Ph.D.)--Stanford University, 2012. |
| Location | electronic resource |
Access conditions
- Copyright
- © 2012 by Eric William Hall
- License
- This work is licensed under a Creative Commons Attribution Non Commercial 3.0 Unported license (CC BY-NC).
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