Elsevier

Sensors and Actuators A: Physical

Volume 216, 1 September 2014, Pages 301-307
Sensors and Actuators A: Physical

Micro patterned quantum dots excitation and imaging for cellular microarray screening

https://doi.org/10.1016/j.sna.2014.05.029Get rights and content

Highlights

  • We describe a quantum dot (QD)-based thin-film multicolor light source array for lab-on-a-chip cellular array imaging.

  • Microwell array and the multicolor light sources are created based on printing technologies.

  • Use of QDs allowed simple multicolor excitation system that covers the wide spectrum of visible wavelengths.

  • We have performed transmission mode imaging of cancer cell morphology to evaluate the nucleus–cytoplasm ratios.

  • We have also performed immunofluorescence imaging of cancer cells cultured in a microwell array.

Abstract

We developed a colloidal quantum dot (QD)-based multicolor excitation light source array designed for compact lab-on-a-chip cell screening and imaging. We have demonstrated multicolor ex vivo transmission mode microscopy to evaluate the nucleus–cytoplasm ratios of cancer cells. We have also performed immunofluorescence excitation of two types of cancer cells (MDA-MB 435 and SKBR3) that are cultured in a microwell array to quantify the disease specific protein expression. Printed array of color filters and microwells were used to perform fluorescence excitation and measurement of the biomarkers. Our method provides patterned multicolor light sources at low-cost that are suitable for high-throughput microarray cellular screening.

Introduction

Colloidal quantum dots (QDs) are 5–10 nm-sized semiconductor nanocrystals where electrons and holes are confined in the three dimensions. Due to this quantum confinement, the energy levels of a single QD are discrete and the band gap is highly related to the size and the shape [1]. QDs have demonstrated potential as fluorescent markers for bioimaging [2], [3], [4]. The tunable band gap of QDs has also been utilized for optoelectronic devices such as light emitting diodes [5] and solar cells [6]. The benefits of QDs for such applications include the emission and absorption wavelengths can be easily tailored through proper choice of materials and sizes; they are much more stable and slow to photobleaching compared to commonly used organic fluorescent dyes.

The compatibility with the advanced microfabrication techniques is another important advantage of integrating QDs on microsystems. In previous studies, we have shown micropatterning of colloidal QDs on silicon substrates [7], [8], [9]. The feature size of patterned QDs can be arbitrary chosen from millimeter scale to single molecular order [7]. The patterned QDs were also electrically excited to compose light emitting diodes (LEDs) that were used for nanoscale fluorescence excitation [10].

The unique characteristics of quantum dots as an integrated on-chip excitation source include (1) capability of multicolor emission from visible to IR wavelengths, (2) various available methods for photo/electrical excitation, and (3) compatibility with silicon fabrication and micropatterning which make QDs an ideal material to be used in an arrayed format. One example of QD arrays is high definition displays, where red, green, and blue QD pixels are patterned in a dense array to allow creation of full color images [11]. In this paper, we propose a colloidal QD-based multicolor excitation light source which is designed for arrayed lab-on-a-chip systems for cell culture, analysis and imaging. It is made on a glass slide and adds the functions of absorption and fluorescence imaging to a standard microscope.

We utilized printing-based techniques to construct the miniature array of QD light sources, chambers, and optical filters. Such integrated microscopic systems enable novel microarrays for cell culture and screening, where cells are investigated while grown in several hundreds of microwells under precisely controlled environments [12]. Another important potential application is microchip based detection of tumor cells [13], where integrated immunofluorescence imaging is much needed. Imaging of multiple biomarkers enables precise identification and quantification of cells [14]. In order to demonstrate the efficacy of our light source, we perform ex vivo transmission mode microscopy and fluorescence imaging of cancer cells with the micropatterned QD based light source. When excited by a high power UVLED, the QDs work as an illumination source suitable for high-throughput cellular screening.

Section snippets

Fabrication and characterization of QD light source

QDs we used in this study are CdSe/ZnS core–shell quantum dots, which have a core shell structure where the core is composed of one material and is coated by a shell of another material with a larger band gap. The shell increases the stability of the QDs against environmental changes, and more importantly, improves quantum yield by passivating the surface trap states. Fig. 1(a) shows the structure of the core–shell QDs.

Fig. 1(b) and (c) illustrates the fabrication procedure and the working

Transmission mode imaging

The QD light source was used for transmission-mode illumination imaging to observe cancer cell morphology (Fig. 1c). Measurement of nucleus–cytoplasm ratio is one of the important factors in cancer cell identification and characterization [15], [16], [17]. Cancer cells tend to have larger nucleus–cytoplasm ratio than other cells [16]. This criterion is often applied in computer based cancer cell analysis such as the study of cellular response to drugs [17]. We demonstrate multicolor observation

Conclusion

We have developed a micropatterned QD-based excitation light source for compact cellular screening. The setup is successfully used for ex vivo cancer cell imaging. We demonstrated transmission-mode microscopy to show that the emission wavelength can be chosen to perform high contrast imaging or observation of inner cell structures. We also conducted QD fluorescence imaging of cancer cells to demonstrate the capability of immunoassay-based cancer cell identification. Microwells and optical

Acknowledgement

The authors are grateful for the financial support from National Institute of Health (NIH) National Cancer Institute (NCI) Cancer Diagnosis Program under the grant 1R01CA139070 and the DARPA Young Faculty Award (N66001-10-1-4049).

Kazunori Hoshino is currently an Assistant Professor of Biomedical Engineering, University of Connecticut. He obtained a Ph.D. from the University of Tokyo in 2000. From 2003 to 2006,he worked for the University of Tokyo as a full-time lecturer in the Department of Mechano-Informatics, School of Information Science and Technology. From 2006 to 2013, he worked for the University of Texas at Austin as a Senior Research Associate in the Department of Biomedical Engineering, where he studied

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    Kazunori Hoshino is currently an Assistant Professor of Biomedical Engineering, University of Connecticut. He obtained a Ph.D. from the University of Tokyo in 2000. From 2003 to 2006,he worked for the University of Tokyo as a full-time lecturer in the Department of Mechano-Informatics, School of Information Science and Technology. From 2006 to 2013, he worked for the University of Texas at Austin as a Senior Research Associate in the Department of Biomedical Engineering, where he studied microfluidic separation and analysis of circulating tumor cells (CTCs). His research interests include (1) Detection and analysis of cancer cells, and (2) Nano/micro-electro-mechanical systems(NEMS/MEMS) based mechanical sensing and optical imaging. He has more than 100 peer reviewed publications, and is the inventor of 6 US patents and 12 Japanese patents.

    Gauri Bhave received her Bachelor's degree in Instrumentation & Control Engineering from the University of Pune, India in 2007 and the Masters degree in Bioengineering from The University of Texas, Arlington in 2010. She is currently pursuing a PhD degree at the Biomedical Engineering department at the University of Texas, Austin. Her research interests include Quantum Dot nanophotonics, optimization of quantum dot LEDs with applications in lab-on-chip systems for biological imaging.

    Elaine Ng earned her Bachelor of Science degree with Special Honors in Biomedical Engineering from the University of Texas at Austin in 2013. From 2009 to 2013, she conducted research as a National Science Foundation (NSF) Research Experiences for Undergraduates (REU) and Undergraduate Research Fellow on microfluidic integrated platforms in combination with micro- patterning and fabrication techniques for immunoassay-based protein and cancer cell detection. She is currently a NSF Graduate Research Fellow pursuing a Ph.D. in Bioengineering at Stanford University. Her research interest continues to be focused on designing point-of-care technologies for biological and disease detection applications.

    Xiaojing Zhang is currently an Associate Professor at the University of Texas of Austin (UT Austin), USA. He received his Ph.D. from Stanford University, California, and was a Research Scientist at Massachusetts Institute of Technology (MIT), Cambridge, before joining the faculty at UT Austin. Zhang's research focuses on exploring bio-inspired nanomaterials, scale-dependent biophysics, and nanofabrication technology, towards developing new diagnostic devices and methods on probing complex cellular processes and biological networks critical to development and diseases. Both multi-scale experimental and theoretical approaches are combined to investigate fundamental force, flow and energy processes at the interface of engineering and biomedicine. In particular, his laboratory is leading the development of integrated photonic microsystems (MEMS, micro-electro-mechanical systems), semiconductor chips and nanotechnologies critical to healthcare, defense and environmental applications. He has published over 120 peer reviewed papers and proceedings, presented over 45 invited seminars worldwide, and filed over 15 US patents (5 patents issued). His research findings have been highlighted in many public media, and were licensed to two companies: CardioSpectra, Inc. (acquired by Volcano, Nasdaq: VOLC) and NanoLite Systems, Inc. He has co-organized many major conferences in the area of MEMS/BioMEMS, nanotechnologies and biomedical engineering. In addition to being the Principle Investigator of many major grants from U.S. federal agencies such as NIH, NSF and DARPA, Dr. Zhang was also recipient of many prestigious awards, including: the Wallace H. Coulter Foundation Early Career Award for Translational Research in Biomedical Engineering, the British Council Early Career RXP Award, NSF Faculty Early Career Development Program (NSF CAREER) Award, DARPA Young Faculty Award, and an invitee to attend U.S. National Academy of Engineering, Frontiers of Engineering (NAE-FOE) program in 2011, the NAE Frontiers of Engineering Education (NAE FOEE) program in 2012, and subsequently China-America Frontiers of Engineering Symposium (CAFOE) program in 2013. He is currently an Associate Editor for Biomedical Microdevices, IEEE/ASME Journal of Microelectromechanical Systems, and has published a textbook titled “Molecular Sensors and Nanodevices: Principles, Designs and Applications in Biomedical Engineering”.

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