Highly Efficient Selection, Enumeration, Enrichment, and Molecular Profiling of Low-Abundance Biological Cells

• Main Author: Dharmasiri, Udara R. Dharmasiri Rasika
• Other Authors: Francis, Joseph
• Format: Others
• Language: en
• Published: LSU 2010
• Subjects: Chemistry
• Online Access: http://etd.lsu.edu/docs/available/etd-10292010-133201/

After brief overviews of low-abundance cell selection techniques in chapter 1 and circulating tumor cells in chapter 2, this dissertation initially focuses on the development of aptamer incorporated high-throughput microfluidic techniques to select rare circulation prostate cancer cells (LNCaP) directly from whole blood with subsequent quantification of these rare cells using a non-labeling approach. Then, I extended the technology to environmental samples in an effort around time, sensitivity, and portability of traditional groundwater assessment. As a model bio- pathogen, E. coli O157:H7 was chosen due to its toxicity and its adverse impact on recreational waters. Low-abundance (<100 cells mL-1) E. coli O157:H7 cells were isolated and enriched from environmental water samples using a microfluidic chip that its capture beds were covalently decorated with E.coli O157:H7 specific polyclonal antibodies. The selected cells were enumerated using RT-qPCR technique. Finally, I have integrated HTMSU with electrokinetic enrichment microfluidic unit for performance of single recombinant low-abundance CTC cell-based assay. A series of analytical processes were carried out, including immunoaffinity selection of rare CTCs, quantification of selected cells via conductivity impedance and electrophoretic enrichment of selected cells for PCR/LDR/CE interrogation for detection of low-abundance point mutations in genomic DNA.