| Author | Kanatip Ratanachoo |
| Call Number | AIT Diss no.EV-02-06 |
| Subject(s) | Environmental toxicology
Dielectrophoresis
|
| Note | A dissertation submitted in partial fulfillment of the requirements for the degree of Doctor ofTechnical Science, Asian Institute of Technology; Inter- University Program on Environmental Toxicology, Technology and Management, Chulabhorn Research Institute and Mahidol University |
| Publisher | Asian Institute of Technology |
| Abstract | The ultimate goal of this thesis is to develop a technology, which can
provide an appropriate model and test system to detect and evaluate toxicity of chemical
pollution in water. This technology is aimed at prouding rapid detection of chemical
toxicity in a relatively simple and easy to use manner, yielding a useful tool for toxicity
screening applications both in the laboratory and field.
Dielectrophoresis (DEP), the translational movement of particles caused by
a nonuniform AC electric field, has been used to detect changes in cell membrane
properties following a variety of biological events. Cells possessing different dielectric
properties experience different DEP forces and are caused to migrate differentially to
strong or weak electric field regions. The central hypothesis of this work is that cells
exposed to toxicants will exhibit modifications in cellular characteristics that result in
dielectric changes that can be detected by DEP.
Time and dose responses of the human cultured leukemia (HL-60) line and
fish cell line (fathead minnow, FHM) were measured following exposure to paraquat,
styrene oxide (SO), N-Nitroso-N-Methylurea (NMU) and puromycin. These toxicants
were selected to represent different predominant mechanisms of action, namely membrane
free radical attack, simultaneous membrane and nucleic acid attack, nucleic acid alkylation,
·and protein synthesis inhibition, respectively. For all toxicant treatments, the specific
membrane capacitance ( C111e111) of the cells decreased while the specific membrane
conductance (Gmem) increased in dose- and time- dependent manners. The DEP responses
correlated sensitively with alterations in cell surface morphology, especially folds,
microvilli, and blebs, observed by scanning electron microscopy. Different sensitivities of
cell responses were observed for each toxicant studied in the order paraquat > SO >
puromycin > NMU for HL-60 cells. FHM cells were more sensitive to all toxicants than
HL-60 cells and showed membrane dielectric responses at ve1y low doses and at shorter
times than HL-60. Cellular responses to paraquat and styrene oxide, which directly
damaged the cell membrane, could be detected 15 minutes after exposure, while responses
to puromycin and NMU, which acted on intracellular targets, could be detected after 30
minutes. By comparison, cell viability assessments showed that 0% of HL-60 cells and less
than 10% of FHM cells died at the doses and exposure durations at which dielectric
responses could be detected. The detection times and dose sensitivity results revealed that
the DEP method is much faster and more sensitive than conventional cell and higher
organism viability testing techniques. Moreover, all tests were conducted using cells from
higher organisms, therefore, the method can potentially provide better data pertaining to
higher organisms responses, including human exposure risks, than methods based on lower
organisms.
These results suggest the possibility of producing new technologies for
toxicity detection and screening based on cellular dielectric responses. A prototype
DEP-based instrnments for environmental toxicity applications was designed and built
using Micro Total Analysis System (~LT AS) concepts. This microfluidic device integrated a
microchip, interconnected channels and chambers. All sample manipulation steps including
cell and toxicant mixing, cell characterization in DEP-FFF system and cell detection using
AC impedance sensor, took place in this one chip under electronic control. The results
demonstrated that DEP- based microfluidic devices are potentially useful tools for toxicity
screening and the rapid detection of cellular responses to toxicants. |
| Year | 2002 |
| Type | Dissertation |
| School | School of Environment, Resources, and Development (SERD) |
| Department | Department of Energy and Climate Change (Former title: Department of Energy, Environment, and Climate Change (DEECC)) |
| Academic Program/FoS | Environmental Engineering and Management (EV) |
| Chairperson(s) | Khunying Mathuros Ruchirawat; |
| Examination Committee(s) | Chongrak Polprasert;
Gascoyne, Peter R. C.;
Jutamaad Satayavivad;
Preeda Parkpian;Becker, Frederick F. ; |
| Scholarship Donor(s) | Post- Graduate Education, Training and Research
Program in Environmental Science, Technology and
Management under Higher Education Development
Project of the Ministry of University Affairs, Thailand;
King Golden Jubilee Scholarship, The Thailand Research Fund; |
| Degree | Thesis (Ph.D.) - Asian Institute of Technology, 2002 |
