Computer screen photo-assisted detection of complementary DNA strands using a luminescent zwitterionic polythiophene derivative

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Abstract

The computer screen photo-assisted technique (CSPT), a practical method for the evaluation of assays using computer screens as light sources and web cameras as detectors, has been used to detect the attachment of complementary DNA strands (20-mer, 5′-CAT GAT TGA ACC ATC CAC CA-3′) to a complex of single DNA strands and a polythiophene derivative (poly(3-[(S)-5-amino-5-carboxyl-3-oxapentyl]-2,5-thiophenylene hydrochloride (POWT)). The complex is a highly sequence specific indicator, based on non-covalent coupling of DNA to a water-soluble, zwitterionic, electroactive and photoactive polymer able to produce a combined absorption-emission signal readable by CSPT.

The observed CSPT signal retains key spectral features of the complex spectrum, distinguishing the DNA attachment, as well as other stimuli, such as pH regulation at concentrations of 30 μM POWT and 15 μM DNA. A CSPT time resolved (linked to temperatures between 8 and 18 °C) approach is demonstrated as a complementary source of discrimination and for testing the robustness of the achieved classification.

Introduction

The computer screen photo-assisted technique (CSPT) is a practical method for the evaluation of colorimetric or fluorescent assays, which only demand a standard computer set and a web camera as measuring platform. Naturally, within the potential applications of CSPT [1], [2], [3], [4], are the remote evaluation of medical diagnostic or environmental tests at homes, using the already available computers and Internet connections.

Different existing sensing technologies are suitable with CSPT, as it has been demonstrated so far [1], [2], but indeed one of the most attractive alternatives is the use of assays based on the recognition of specific DNA strands, in a similar way as complementary DNA (cDNA) microarrays perform for tracing human diseases to its very genetic cause.

Although CSPT is a versatile method, it does not compete with dedicated analytical methods in detection limit or accuracy. For exploiting its own advantages, CSPT requires the aid of a suitable chemistry providing a response readable by the platform. In this way, a very promising selective fluorometric DNA hybridization detection method has been recently demonstrated [5], in this case giving highly sequence-specific information, based on non-covalent coupling of DNA to a water-soluble, zwitterionic, electroactive and photoactive polythiophene derivative [6]. This alternative offers a novel way to create DNA assays without using covalent attachment to a receptor (or labeling of the analyte).

In this work, we demonstrate the feasibility of CSPT for detecting the attachment of a 20 mer DNA strand to its complementary chain complex with the referred polythiophene derivative. The robustness of the detection is verified by a time resolved experiment linked to a temperature change between ∼8 and 18 °C.

Section snippets

Experimental

The CSPT light source used in this work was a 130 × 80 mm2 area of a standard computer screen (a LCD monitor, Sony Vaio PCG-FX 505, operating at XGA resolution) displaying a 50-color illuminating sequence at a rate of 1 color/s. This particular color sequence, which resembles the human perception of the visible spectrum, is repeated continuously during 30 min (36 times) until the end of the experiment. Along the experiment, the screen illuminates the whole array of samples at the same time (in this

Results and discussion

Fig. 1a shows the absorption and emission characteristics of the present samples (for narrowband excitation at 400 nm). The absorption and emission spectra of POWT were recorded after diluting samples S1, S2, S3 and S4 four times; this gives a polymer concentration of 0.025 mg/ml (7.5 μM).

The S2 sample shows an absorption maximum around 440 nm and a shoulder in the region at 550 nm, associated with a planar polymer and aggregation of polymer chains, respectively [7]. When the polymer is associated

Conclusions

The ability of CSPT to detect the subtle spectral features associated with the attachment of a 20 mer complementary DNA strand to a single-stranded DNA–POWT complex has been demonstrated. This result corroborates the flexibility of CSPT for evaluating different kind of assays (colorimetric, fluorescent, absorbing, etc.) [1], [2], [4]. Thus, the sensitivity and purpose of the measurements is given by the assay itself (e.g. cell viability, hormone detection, DNA attachment), the measuring

Acknowledgments

Financial support from the Swedish Research Council and the Swedish Agency for Innovation Systems (VINNOVA) is gratefully acknowledged by the authors.

Daniel Filippini was born in 1968 in Buenos Aires, Argentina. He received his electrical engineer degree (MSc) from the National Technological University, Buenos Aires, Argentina, in 1993. In 1998, he obtained MSc in biomedical engineering, at Favaloro University, Argentina. From 1998 to 2000 he worked on his PhD, as a DAAD scholar, at the Institute of Physical and Theoretical Chemistry, University of Tübingen, Germany, obtaining his PhD from the University of Buenos Aires in 2000. Since 2003

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Daniel Filippini was born in 1968 in Buenos Aires, Argentina. He received his electrical engineer degree (MSc) from the National Technological University, Buenos Aires, Argentina, in 1993. In 1998, he obtained MSc in biomedical engineering, at Favaloro University, Argentina. From 1998 to 2000 he worked on his PhD, as a DAAD scholar, at the Institute of Physical and Theoretical Chemistry, University of Tübingen, Germany, obtaining his PhD from the University of Buenos Aires in 2000. Since 2003 he is an assistant professor in applied physics at the Department of Physics and Measurement Technology at Linköping University, Sweden. Filippini has been appointed docent in 2005. His current research interest relates to controlled light assisted sensor systems for chemical or biochemical applications, and the development of new concepts for chemical image generation.

Peter Åsberg was born in Nässjö, Sweden, on December 1, 1973. He recieved his MSc degree in engineering biology, with specialization towards microsystem technology and biosensors, from Linköping University, Sweden, in 2001. At present he is working as a PhD student at the Department of Physics, Measurement Technology, Biology and Chemistry, Linköping University, Sweden. His research interests are development of biosensors and biochips based on luminescent conjugated polyelectrolytes.

Peter Nilsson was born in Linköping, Sweden, on February 7, 1970. He recieved his MSc degree in Chemistry from Kalmar University, Sweden, in 2000. At present he is working as a PhD student at the Department of Physics, Measurement Technology, Biology and Chemistry, Linköping University, Sweden. His research interests is to develop biosensors based on luminescent conjugated polyelectrolytes.

Olle Inganäs, Professor of Biomolecular and organic electronics, IFM, Linköpings Universitet. Born in Huddinge 1951, received a MSc in technical physics from Chalmers University of Technology 1977, a BSc in philosophy and economics from Göteborg University in 1978, and a PhD in applied physics at Linköping University in 1984. He was appointed docent in applied physics in 1989 and a professor in 1999. Inganäs has focused on studies of the class of conjugated polymers (conducting and emitting polymers) throughout areas of polymer physics, electrochemistry, electronics and optics, and covering topics of chromism in soluble conjugated polymers, electronic devices with semiconducting polymers, (transistors, diodes, light emitting diodes and photodiodes/solar cells) and electrochemical devices (energy storage in batteries and supercapacitors, smart windows and actuators). He has published ca. 280 papers and holds 20 patents and patent applications, and has contributed to the creation of 3 technology companies.

Ingemar Lundström was born in 1941 in Skellefteå, Sweden. He received his PhD (in 1970) in electrical engineering (solid-state electronics) from Chalmers University of Technology, Göthemburg, Sweden. He was an assistant professor at the Research Laboratory of Electronics, Chalmers, until 1978, when he was appointed as a professor at the technical faculty of Linköping University, Linköping, Sweden, where he now heads the Laboratory of Applied Physics. The laboratory, which has an interdisciplinary research staff, conducts research on chemical sensors and biosensors, catalysis, thin films, conductive polymers, surface modifications, biomaterials and interface biology. Lundström is presently involved, in the research and development of high temperature chemical sensors, electronic noses and tongues, surface oriented biospecific interaction analysis and natural nanosystems such as pigment containing cells for biosensing purposes. Lundström has published about 450 scientific papers.

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