Wide Field-of-View Fluorescence Imaging with Optical-Quality Curved Microfluidic Chamber for Absolute Cell Counting
"> Figure 1
<p>(<b>a</b>) Optical prescription for prototype curved-substrate imaging system, designed for a wavelength of 560 nm. A single commercial lens was used for imaging. All dimensions are in millimeters. (<b>b</b>) Schematic of experimental setup.</p> "> Figure 2
<p>(<b>a</b>) 3D model of curved sample chamber composed of an upper window (gray color) and a lower substrate (black color). A sample solution is introduced into the gap of 100-μm thickness between those two separable parts through one of the holes in the lower substrate. The other three holes are air vents. (Scale bar = 10 mm). (<b>b</b>) Curved sample chamber fabricated by plastic injection molding technique. The upper window is optically clear for image acquisition and the lower substrate is colored black to minimize light reflection at the surface.</p> "> Figure 3
<p>(<b>a</b>) A schematic of the cross-sectional view of the curved sample chamber and suspended cells in the gap between upper window and lower substrate; and (<b>b</b>) loading of cell suspension with a micropipette is facilitated by air vent holes made through the lower substrate as shown in (<b>a</b>).</p> "> Figure 4
<p>Ray tracing simulation of astigmatism field curvature. Ray tracing of a simple biconvex lens for: (<b>a</b>) flat substrate; and (<b>c</b>) curved substrate. The grayscale images in (<b>b</b>,<b>d</b>) were acquired using the flat and the curved substrates, where the image plane was kept flat. The images in (<b>b</b>,<b>d</b>) represent 0, 3, and 6 mm sample substrate height. The field curvature graphs for the flat and the curved substrates are shown in (<b>e</b>,<b>f</b>), respectively, where the blue and red lines denote tangential and sagittal surfaces.</p> "> Figure 5
<p>Analytical characterization of simple imaging system for flat and curved substrates. The spot size diagrams include a circle showing the Airy disk (9.87 μm in radius) for the flat and the curved substrates in (<b>a</b>,<b>d</b>), respectively. (<b>b</b>,<b>e</b>) MTF and (<b>c</b>,<b>f</b>) PSF of flat and curved substrates, respectively. In the MTFs, the black line shows the diffraction limit, and the red and pink lines represent the on- and off-axis image fields, respectively.</p> "> Figure 6
<p>Change in surface power and object curvature with change in curvature radius of lens. These radii of the lens consider both sides of biconvex lens, and have the same focal length as the radius.</p> "> Figure 7
<p>Imaging performance of single-lens fluorescence microscopy with curved sample chamber. Fluorescence images taken for beads of diameter 10 μm for: (<b>a</b>) planar; and (<b>b</b>) curved sample chambers. The fluorescent microbeads are distributed uniformly over the entire FOV. The intensity profiles of the bead images are shown in (<b>c</b>,<b>d</b>). As observed for the curved substrate, a wider area of the bead image is more focused in (<b>b</b>), and the aberration is lower than that in (<b>a</b>). (Scale bar = 1 mm).</p> "> Figure 8
<p>Fluorescence images of stained white blood cells in small and large field imaging. (<b>a</b>) Microscopic image of flat chamber. The inset white boxes in (<b>b</b>,<b>c</b>) are of the same size in the microscopy images of the flat and curved chambers. (<b>d</b>,<b>e</b>) Cropped areas of the flat and curved chamber images, where (ii) denotes the center part of an image. All cropped images are of the same size. The cell counts in the center (ii) were similar for both chambers. The bars in (<b>b</b>,<b>c</b>) represent a scale of 1 mm, while those in the other figures represent a scale of 200 µm.</p> "> Figure 9
<p>(<b>a</b>) WBC count obtained using planar chamber is remarkably different from the reference cell count obtained with conventional microscopy; and (<b>b</b>) WBC concentrations of 10%–70% with an interval of 10% and 100% were compared for absolute counting obtained by microscopic, flat, and curved chamber imaging. The count obtained with the curved chamber is close to that obtained with conventional microscopy. However, the corresponding result with the flat chamber differs considerably for large fields.</p> "> Figure 10
<p>(<b>a</b>–<b>c</b>) The reference, flat, and curved chamber images with 500 µm<sup>2</sup> white boxes inside, respectively (scale bar = 0.5 mm); (<b>d</b>–<b>f</b>) the normalized intensity profiles of a fluorescently-labeled WBC in those three images; and (<b>g</b>) the size of the fluorescent WBCs measured with microscopy, and flat and curved imaging, represented by Ref, Flat, and Curved on the <span class="html-italic">x</span>-axis of the graph, respectively. The range of WBC size is 16–20 µm. The normalized intensity profile of the fluorescence shows the size of the SYTO-stained WBC in pixels, and the inset images are cropped from the center part of the processed image.</p> ">
Abstract
:1. Introduction
2. Materials and Methods
2.1. Theory
2.2. Imaging Setup
2.3. Fabrication of Curved Sample Chamber
2.4. Sample Preparation
2.5. Image Acquisition and Cell Counting
3. Results and Discussion
3.1. Simulation Study
3.2. Validation Study
3.3. Absolute Counting of WBC
3.4. Advantages of Large FOV Imaging with Curved Sample Chamber
4. Conclusions
Acknowledgments
Author Contributions
Conflicts of Interest
References
- Jenkins, F.A.; White, H.E. Fundamentals of Optics, 4th ed.; Shirley, G., Ed.; McGraw-Hill: New York, NY, USA, 2001. [Google Scholar]
- Cybulski, J.S.; Clements, J.; Prakash, M. Foldscope: Origami-based paper microscopy. PLoS ONE 2014, 9, e98781. [Google Scholar] [CrossRef] [PubMed]
- Smith, Z.J.; Chu, K.; Espenson, A.R.; Rahimzadeh, M.; Gryshuk, A.; Molinaro, M.; Dwyr, D.M.; Lane, S.; Matthews, D.; Wachsmann-Hogiu, S. Cell-phone-based platform for biomedical device development and education application. PLoS ONE 2011, 6, e17150. [Google Scholar] [CrossRef] [PubMed]
- Breslauer, D.N.; Maamari, R.N.; Switz, N.A.; Lam, W.A.; Fletcher, D.A. Mobile phone based clinical microscopy for global health applications. PLoS ONE 2009, 4, e6320. [Google Scholar] [CrossRef] [PubMed]
- Zhu, H.; Sencan, I.; Wong, J.; Dimitrov, S.; Tseng, D.; Nagashima, K.; Ozcan, A. Cost-effective and rapid blood analysis on a cell-phone. Lab Chip 2013, 13, 1282–1288. [Google Scholar] [CrossRef] [PubMed]
- Choi, J.R.; Kim, K.J.; Kim, D.H. In situ fluorescence optical detection using a digital micromirror device (DMD) for 3D cell-based assays. J. Opt. Soc. Korea 2012, 16, 42–46. [Google Scholar] [CrossRef]
- Potsaid, B.; Finger, F.P.; Wen, J.T. Automation of challenging spatial-temporal biomedical observations with the adaptive scanning optical microscope (ASOM). IEEE Trans. Autom. Sci. Eng. 2009, 6, 525–535. [Google Scholar] [CrossRef]
- Hecht, E. Optics, 4th ed.; Addison Wesley: San Francisco, CA, USA, 2002; p. 226. [Google Scholar]
- Rim, S.B.; Catrysse, P.B.; Dinyari, R.; Huang, K.; Peumans, P. The optical advantages of curved focal plane arrays. Opt. Exp. 2008, 16, 4965–4971. [Google Scholar] [CrossRef]
- Jin, H.; Abelson, J.R.; Erhardt, M.K.; Nuzzo, R.G. Soft lithographic fabrication of an image sensor array on a curved substrate. J. Vac. Sci. Technol. B 2004, 22, 2548–2551. [Google Scholar] [CrossRef]
- Malyarchuk, V.; Jung, I.; Rogers, J.A.; Shin, G.; Ha, J.S. Experimental and modeling studies of imaging with curvilinear electronic eye cameras. Opt. Exp. 2010, 18, 27346–27358. [Google Scholar] [CrossRef] [PubMed]
- Jung, I.; Xiao, J.; Malyarchuk, V.; Lu, C.; Li, M.; Liu, Z.; Rogers, J.A. Dynamically tunable hemispherical electronic eye camera system with adjustable zoom capability. Proc. Natl. Acad. Sci. USA 2011, 108, 1788–1793. [Google Scholar] [CrossRef] [PubMed]
- Ozcan, A.; Demirci, U. Ultra wide-field lens-free monitoring of cells on-chip. Lab Chip 2008, 8, 98–106. [Google Scholar] [CrossRef] [PubMed]
- Greenbaum, A.; Luo, W.; Su, T.W.; Göröcs, Z.; Xue, L.; Isikman, S.O.; Coskun, A.F.; Mudanyali, O.; Ozcan, A. Imaging without lenses: Achievements and remaining challenges of wide-field on-chip microscopy. Nat. Methods 2012, 9, 889–895. [Google Scholar] [CrossRef] [PubMed]
- Seo, S.; Isikman, O.S.; Sencan, I.; Mudanyali, O.; Su, T.W.; Bishara, W.; Erlinger, A.; Ozcan, A. High-throughput lens-free blood analysis on a chip. Anal. Chem. 2010, 82, 4621–4627. [Google Scholar] [CrossRef] [PubMed]
- Su, T.W.; Erlinger, A.; Tseng, D.; Ozcan, A. Compact and light-weight automated semen analysis platform using lensfree on-chip microscopy. Anal. Chem. 2010, 82, 8307–8312. [Google Scholar] [CrossRef] [PubMed]
- Mudanyali, O.; Oztoprak, C.; Tseng, D.; Erlinger, A.; Ozcan, A. Detection of waterborne parasites using field-portable and cost-effective lensfree microscopy. Lab Chip 2010, 10, 2419–2423. [Google Scholar] [CrossRef] [PubMed]
- Göröcs, Z.; Ling, Y.; Yu, M.D.; Karahalios, D.; Mogharabi, K.; Lu, K.; Wei, Q.; Ozcan, A. Giga-pixel fluorescent imaging over an ultra-large field-of-view using a flatbed scanner. Lab Chip 2013, 13, 4460–4466. [Google Scholar] [CrossRef] [PubMed]
- Ulrich, W.; Shafer, D.; Bader, D.; Epple, A. Refractive Optical Imaging System, in Particular Projection Objective for Microlithography. U.S. Patent 7,511,890, 31 March 2006. [Google Scholar]
- Malacara, D.; Malacara, Z. Handbook of Optical Design, 2nd ed.; Marcel Dekker: New York, NY, USA, 2004. [Google Scholar]
- Hagen, N.; Tkaczyk, T. Foveated endoscopic lens. J. Biomed. Opt. 2012. [Google Scholar] [CrossRef] [PubMed]
- Zhu, H.; Yaglidere, O.; Su, T.; Tseng, D.; Ozcan, A. Cost-effective and compact wide-field fluorescence imaging on a cell-phone. Lab Chip 2010, 11, 315–322. [Google Scholar] [CrossRef] [PubMed]
- Lee, W.; Kwon, D.; Choi, W.; Jung, G.Y.; Jeon, S. 3D-printed microfluidic device for the detection of pathogenic bacteria using size-based separation in helical channel with trapezoid cross-section. Sci. Rep. 2015. [Google Scholar] [CrossRef] [PubMed]
- Smith, W.J. Modern Optical Engineering, 4th ed.; McGraw-Hill: New York, NY, USA, 2007; p. 90. [Google Scholar]
- Arpali, S.A.; Arpali, C.; Coskun, A.F.; Chiang, H.H.; Ozcan, A. High-throughput screening of large volumes of whole blood using structured illumination and fluorescent on-chip imaging. Lab Chip 2012, 12, 4968–4971. [Google Scholar] [CrossRef] [PubMed]
- Cho, M.O.; Chang, H.M.; Lee, D.; Yu, Y.G.; Han, H.; Kim, J.K. Selective detection and automated counting of fluorescently-labeled chrysotile asbestos using a dual-mode high-throughput microscopy (DM-HTM) method. Sensors 2013, 9, 5686–5699. [Google Scholar] [CrossRef] [PubMed]
- Shapiro, M.H. Practical Flow Cytometry, 4th ed.; Wiley-Liss: Hoboken, NJ, USA, 2003; pp. 18–20. [Google Scholar]
Lens | Inner Surface of Sphere | Outer Surface of Sphere | ||||
---|---|---|---|---|---|---|
Radius | Height | Volume | Radius | Height | Volume | |
(mm) | (mm) | (mm3) | (mm) | (mm) | (mm3) | |
LB1014 | 15.06 | 1.25 | 71.88 | 15.56 | 1.75 | 144.09 |
LB1258 | 18.08 | 1.03 | 59.114 | 18.58 | 1.53 | 132.89 |
LB1378 | 24.12 | 0.8 | 47.96 | 24.62 | 1.3 | 128.41 |
LB1844 | 30.135 | 0.605 | 34.42 | 30.635 | 1.105 | 116.10 |
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Shourav, M.K.; Kim, K.; Kim, S.; Kim, J.K. Wide Field-of-View Fluorescence Imaging with Optical-Quality Curved Microfluidic Chamber for Absolute Cell Counting. Micromachines 2016, 7, 125. https://doi.org/10.3390/mi7070125
Shourav MK, Kim K, Kim S, Kim JK. Wide Field-of-View Fluorescence Imaging with Optical-Quality Curved Microfluidic Chamber for Absolute Cell Counting. Micromachines. 2016; 7(7):125. https://doi.org/10.3390/mi7070125
Chicago/Turabian StyleShourav, Mohiuddin Khan, Kyunghoon Kim, Subin Kim, and Jung Kyung Kim. 2016. "Wide Field-of-View Fluorescence Imaging with Optical-Quality Curved Microfluidic Chamber for Absolute Cell Counting" Micromachines 7, no. 7: 125. https://doi.org/10.3390/mi7070125