Abstract 2721: Real-time and label-free aptasensor of VEGF for cancer diagnostics

Abstract

Abstract Early detection of cancer markers is a critical demand for clinical research, particularly in preventing cancer cells from establishing themselves in a metastatic stage. We have developed a stable, sensitive, and selective detection method utilizing a nanoplasmonic ‘aptasensor’ (aptamer-based sensor) of vascular endothelial growth factor (VEGF) for breast cancer diagnostics. Nanoplasmonic aptasensors are prepared by fixing gold nanoplasmonic particles (GNPs) on an amino-silane coated glass slide and coating poly-L-lysine (PLL) on top of GNPs, which provides the GNP with a strong positive-charged surface and a spacing layer avoiding quenching. To accomplish high selectivity, VEGF-specific aptamers are immobilized on the PLL-GNP surface by an electrostatic force. To monitor the event of target binding, the aptamer is conjugated with a fluorescent molecule, Cy-3, which has an absorption peak wavelength (λ = 552 nm) that matches with a localized surface plasmon resonance wavelength of the 80 nm in diameter GNP (λ ≈ 550 nm). In the absence of target VEGF molecules, Cy-3 is spaced in a relevant coupling distance from the GNP surface for the surface plasmon resonance to amplify an incident light and enhance the fluorescent signal of Cy-3. However, once VEGF molecules bind to the aptamer-Cy-3, the conformational change of aptamer occurs, leading to its subsequent detachment from the GNP surface, which results in no signal enhancement and, thus, a loss in the fluorescence intensity. Therefore, the concentration of VEGF molecules can be estimated by measuring the amount of signal decrease in the fluorescence intensity. With high affinity and specificity, the aptasensor of VEGF requires a small amount of sample (10 μL) for a single detection and incubation time of 1 hour at a room temperature. Also, the aptasensor achieves VEGF detection from 100 nM (2 μg/mL) to 100 pM (2 ng/mL) with a signal decrease up to 50%. In contrast, there is no discernible signal change in a non-specific binding protein solution (100 nM bovine serum albumin) or a strong ionic solution (1.4 M NaCl). Furthermore, the aptasensor is applicable to VEGF detection in human fluids, owing to the biological stability of a fully 2′-O-methyl nucleotide-based aptamer. We can detect VEGF molecules present in saliva and serum of breast cancer patients by observing signal decrease corresponding to the concentration of 100 nM (2 μg/mL) VEGF and a consistent decrease with concentration variation from 1 to 100 % in saline. The aptamer-based nanoplasmonic sensor may be useful for not only basic studies on angiogenesis but also clinical research in evaluating the effectiveness of drug candidates that inhibit VEGF secretion. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 2721.