Cell-Specific Aptamer Modified Electrodes for Highly Sensitive and Specific Detection of Tumor

Cells
Author :QuLiMing
Tutor:PengZuo
School :Suzhou University
CLC :R730.4
TYPE :Masters thesis
Download the PDF Full Text:http://www.topresearch.org/showinfo-171-677769-0.html
Year:2013
Abstract:
Circulating tumor cells (CTCs) are cancer cells that are circulating in the peripheral blood or
lymphatic vessels, which originate from primary tumors or metastatic locus, travel to distant tissue
sites and eventually form new tumor deposits. Early detection of CTCs in the peripheral blood is
critical for monitoring tumor progression. However, due to the extremely limited number of CTCs in
the blood, and the complexity of blood samples, reliable detection methods with higher sensitivity to
decrease false negative rate, and higher specificity to decrease false positive rate are still highly
desirable in clinical setting.Aptamers are oligonucleic acid molecules that are usually selected from a
pool of random sequences by in vitro screening. They can bind to their specific targets with high
specificity and affinity. Over the years, a number of electrochemical detection methods have been
developed for highly sensitive and convenient biosensing. In this work, we combine the advantages of
both aptamer and electrochemistry, fabricated tumor cell-specific aptamer modified electrodes. After a
series of optimization, which led to signal enhancement and significantly lowered background, the
aptamer-modified electrodes were applied in highly sensitive and specific detection of tumor cells in
whole blood cells.We first covalently attached TLS1c, an aptamer specific for mouse hepatoma cell
line BNL1ME A.7R.1(MEAR), to the surface of a glassy carbon electrode (GCE) through a single
stranded DNA linker (T15). Binding of cells to GCE-immobilized TLS1c was analyzed by cyclic
voltammetry (CV), electrochemical impedance spectroscopy (EIS), and difference pulse voltammetry
(DPV), and showed that TLSlc modified on the surface of electrode could recognize MEAR cells
specifically.In order to enhance sensitivity and decrease the background signal, we first separated
whole blood cells (WBC) from peripheral blood by a series wash and centrifugations, decreasing the
sample complexity. Then, we converted the flexible T15DNA linker (ss-linker) into a rigid one (dslinker) by hybridizing with its complementary strand A15. Compared with ss-TLS1c-mosified GCE,
the ds-TLS1c-mosified GCE showed lower background, and further detection of increasing number of
MEAR cells using DPV showed that as low as10MEAR cells can be selectively detected
in109WBC.We further optimized the electrode with two MEAR-specific aptamers, TLS1c (ss-linker)
and TLS11a (ds-linker), to facilitate the capture of MEAR cells on the modified GCE. The rigid dslinker ensures the recognition of MEAR cells and probes happens away from the GCE surface,
decreasing steric hindrance, while the flexible ss-linker enables TLS1c to better capture MEAR cells.
Our results demonstrated that, compared to ds-TLS1c-GCE and ds-TLS1c/ds-TLS11a-GCE-GCE, this
dual-aptamer modified GCE (ss-TLS1c/ds-TLS11a-GCE-GCE) possesses the highest signal to noise
ratio, and is able to detect a single MEAR cell in109WBC, with a liner range from0-7MEAR cells,
suggesting promising potential in early CTC detection.