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Functional magnetic nanoparticle-based trapping and sensing approaches for label-free ﬂuorescence detection of DNA
Ju-Yu Lin, Yu-Chie Chen ∗
Department of Applied Chemistry, National Chiao Tung University, Hsinchu 300, Taiwan
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In this study, a label-free ﬂuorescence detection method for DNA was designed using functional magnetic nanoparticles (MNPs) as afﬁnity probes. With the advantage of magnetic feature, MNP-based afﬁnity probes can be easily manipulated for trapping and sensing target species. Two types of MNP-based nanoprobes for trapping and detecting target DNAs were fabricated. The basicstrategy for this approach is the use of trapping probes to concentrate target DNAs selectively from complex samples. The detection probes are then used as ﬂuorescence reporters to explore the level of the target species. Trapping probes were constructed by covalently immobilizing probe DNA molecules complementary to the target DNA. Detection nanoprobes were made by linking a ﬂuorescent dye,riboﬂavin-5 -monophosphate (RFMP), onto the surface of the core/shell Fe3 O4 @Al2 O3 MNPs via Al-phosphate chelation. The ﬂuorescence derived from RFMP molecules became invisible when molecules were attached onto the MNP surface. However, after phosphorylated species (e.g., DNA molecules) replaced RFMP from the surface of the RFMP-Fe3 O4 @Al2 O3 MNPs under microwave heating for 15 s, the RFMP moleculesreleased from the MNPs enhanced the ﬂuorescence intensity in the solution. Based on the measurement of the ﬂuorescence intensity, the level of target DNA in the samples was determined. The remaining DNA molecules on the RFMP-Fe3 O4 @Al2 O3 MNPs were characterized by using matrix-assisted laser desorption/ionization mass spectrometry (MALDI MS). The detection limit for DNA was as low as 40 pMusing this approach. © 2011 Elsevier B.V. All rights reserved.
Article history: Received 18 June 2011 Received in revised form 26 August 2011 Accepted 29 August 2011 Available online 17 September 2011 Keywords: DNA Label free sensing Fluorescence MALDI-MS Magnetic nanoparticles
1. Introduction DNA carries informative genetic codes. Thus, it has been used as the main target in biodiagnosticsand biochemical analyses [1–3]. Electrochemistry and optical spectroscopy have been used as analytical tools for DNA detection [1–6]. For high throughput DNA analysis, DNA chips are the typical choice [7–11]. However, ﬂuorescence-labeled nucleic acid probes are usually required [9,10]. Furthermore, amplifying DNA prior to chip analysis may be necessary because of its insufﬁcient sensitivity.Although real-time polymerase chain reaction (PCR) can provide the required sensitivity for DNA detection , it is generally unsuitable for high throughput analysis because of its time-consuming experimental steps and the high cost of reagents. Alternatively, analytical methods based on label-free ﬂuorescence detection [8,13–15], and molecular beacons [9,16,17] have recently attracted muchattention. For example, intercalating dyes like ethidium bromide  have been demonstrated as highly sensitive toward double strand DNA. Combining the ability to concentrate traces of target DNA
∗ Corresponding author. Tel.: +886 3 5131527; fax: +886 5723764. E-mail address: email@example.com (Y.-C. Chen). 0039-9140/$ – see front matter © 2011 Elsevier B.V. All rights reserved.doi:10.1016/j.talanta.2011.08.061
from sample solutions and eliminating unwanted interference are desirable. Developing new methods to improve analysis performance can be beneﬁcial for DNA analysis. The rapid growth of nanotechnology has opened new avenues for progress in DNA-sensing methods. Functionalized gold nanoparticles (NPs) immobilized with DNA probes have been used extensively in the detection of...