Incubator Embedded Cell Culture Imaging System (EmSight) Based on Fourier Ptychographic Microscopy

Abstract

Multi-day tracking of cells in culture systems can provide valuable information in bioscience experiments. We report the development of a cell culture imaging system, named EmSight, which incorporates multiple compact Fourier ptychographic microscopes with a standard multiwell imaging plate. The system is housed in an incubator and presently incorporates six microscopes. By using the same low magnification objective lenses as the objective and the tube lens, the EmSight is configured as a 1:1 imaging system that, providing large field-of-view (FOV) imaging onto a low-cost CMOS imaging sensor. The EmSight improves the image resolution by capturing a series of images of the sample at varying illumination angles; the instrument reconstructs a higher-resolution image by using the iterative Fourier ptychographic algorithm. In addition to providing high-resolution brightfield and phase imaging, the EmSight is also capable of fluorescence imaging at the native resolution of the objectives. We characterized the system using a phase Siemens star target, and show four-fold improved coherent resolution (synthetic NA of 0.42) and a depth of field of 0.2 mm. To conduct live, long-term dopaminergic neuron imaging, we cultured ventral midbrain from mice driving eGFP from the tyrosine hydroxylase promoter. The EmSight system tracks movements of dopaminergic neurons over a 21 day period. "Pharmacological chaperoning of nicotinic acetylcholine receptors reduces the endoplasmic reticulum stress response," Mol. Pharmacol. 81(6), 759-769 (2012). 7. http://www.zeiss.com/microscopy/en_us/products/microscope-components/incubation.html. 8. http://www.nikoninstruments.com/Products/Live-Cell-Screening-Systems/BioStation-CT. 9. http://www.essenbioscience.com/essen-products/incucyte/. 10. A. W. Lohmann, R. G. Dorsch, D. Mendlovic, Z. Zalevsky, and C. Ferreira, "Space-bandwidth product of optical signals and systems," J. Opt. Soc. Am. A 13(3), 470-473 (1996). 11. G. Zheng, X. Ou, and C. Yang, "0.5 gigapixel microscopy using a flatbed scanner," Biomed. Opt. Express 5(1), 1-8 (2014). super-resolution in portable lensless on-chip microscopy using a fiber-optic array," Lab Chip 11(7), 1276-1279 (2011). 17. A. Greenbaum, U. Sikora, and A. Ozcan, "Field-portable wide-field microscopy of dense samples using multiheight pixel super-resolution based lensfree imaging," Lab Chip 12(7), 1242-1245 (2012). 18. A. F. Coskun, I. Sencan, T. W. Su, and A. Ozcan, "Wide-field lensless fluorescent microscopy using a tapered fiber-optic faceplate on a chip," Analyst (Lond.) 136(17), 3512-3518 (2011). 19. A. F. Coskun, I. Sencan, T. W. Su, and A. Ozcan, "Lensless wide-field fluorescent imaging on a chip using compressive decoding of sparse objects," Opt. Express 18(10), 10510-10523 (2010). 20. G. Zheng, R. Horstmeyer, and C. Yang, "Wide-field, high-resolution Fourier ptychographic microscopy," Nat.Photonics 7(9), 739-745 (2013). 21. X. Ou, R. Horstmeyer, C. Yang, and G. Zheng, "Quantitative phase imaging via Fourier ptychographic microscopy," Opt. Lett....

Development of a Low-Cost System for Simultaneous Longitudinal Biological Imaging

“…To date, few 3D printed microscopes are designed to function inside incubators 24, 36 . We have run the picroscope in the incubator for three weeks.…”

“…Other systems have taken advantage of one camera attached to a gantry system to perform imaging of multiple experimental replicates [33][34][35] . Few 3D-printed microscopes have been developed that perform multi-well imaging with medium throughput 34,36 . Several biological applications exist that would greatly benefit from multiwell, multi-week simultaneous imaging, as it allows for concurrent interrogation of different experimental conditions and the inclusion of biological replicates.…”

UC2 – A Versatile and Customizable low-cost 3D-printed Optical Open-Standard for microscopic imaging

“…Alongside different imaging modalities, long-term observations of living organisms with the least possible bias on their behaviour became an important aspect in microscopy. Keeping the cells in a well-controlled environment and state pose additional constraints like imaging inside an incubator [4,5] with appropriate microscopes [6,7,8] or on-microscope incubator units [9]. Assembling, maintaining and improving such setups as well as analysing and verifying the produced data very often requires a specialist dedicated to the respective instrument, separating microscope engineers from microscope users.…”

Imaging of dense cell cultures by multiwavelength lens‐free video microscopy

“…A phase retrieval algorithm without support mask is used to recover the high contrast phase images at a resolution of approximately 1 mm with a large FOV [2.1 mm 3 1.8 mm, (2)]. Recently such a system has been designed to fit inside an incubator, and offers an even larger FOV (5.7 mm 3 4.3 mm) (3). An alternative to maximize the available field of view together with a small footprint is lens-free microscopy (4)(5)(6).…”