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Ouyang, W., Mueller, F., Hjelmare, M., Lundberg, E. & Zimmer, C. ImJoy: An open-source computational platform for the deep learning era. Nat. Methods 16, 1199–1200 (2019). A thin sheet of rubber on the flat side that touches the lenses on the phone will make it steadier on the phone A single islet was loaded into the microfluidic device and incubated in KR2 buffer for 30min and at which time the supernatant was collected to serve as the basal level. After an additional 30min incubation under 14 mM glucose, the supernatant was collected again as the stimulated sample. Insulin concentrations were determined using Mouse Insulin ELISA kit (Mercodia, Winston Salem, NC, USA) using a plate reader (Biotek Synergy H1, BioTek U.S., Winooski, VT, USA). Statistical analysis
Moreover, the iMicro Q3 is cost-effective, costing about 1% of the price of a desktop microscope. This affordability, combined with its ease of use and compatibility with any camera phone, makes the iMicro Q3 an accessible tool for all. Whether for students, young learners, or hobbyists, this device offers an opportunity to explore the microscopic world without the need for expensive and bulky equipment. Taylor, A. B. & Zijlstra, P. Single-molecule plasmon sensing: current status and future prospects. ACS Sens. 2, 1103–1122 (2017).Fakruddin, M. et al. Nucleic acid amplification: alternative methods of polymerase chain reaction. J. Pharm. Bioallied. Sci. 5, 245–252 (2013). Although there are many smartphone-based fluorescence imaging system, to the best of our knowledge this is the first smartphone-based fluorescence imaging system specifically designed for islet function analysis. When compared to conventional fluorescence-microscopic microfluidic systems, the portable system allows an easier setup and a much lower cost (See Table S1 in Supplement 1 for the estimated total cost of the system). Importantly, it can achieve comparable resolution and detection of fluorescence signals when compared to standard fluorescence microscope. One limitation with the current system setup is that current setup can only work with one fluorescence spectrum at a time. However, this problem can be resolved in the future by implementing filter switch functionality into the system with the help of a motorized filter wheel.
Vietz, C. et al. Benchmarking smartphone fluorescence-based microscopy with DNA origami nanobeads: reducing the gap toward single-molecule sensitivity. ACS Omega 4, 637–642 (2019). nm silver nanoparticles (100 nm BioPure Silver Nanospheres (Citrate), nanoComposix, USA) were functionalized with T 20 single-stranded DNA oligonucleotides with a thiol modification at the 3’-end (Ella Biotech GmbH, Germany) 15. Briefly, 2 mL of 0.025 mg/mL nanoparticle solution in ultra pure water was heated to 40 °C under permanent stirring. 20 µL of 10 % Tween 20 and 20 µL of a potassium phosphate buffer (4:5 mixture of 1 M monobasic and dibasic potassium phosphate, Sigma Aldrich, USA) were added as well as 10 µL of a 2 nmol thiol-modified single-stranded DNA solution (5’-T 20-SH-3’) and incubated for 1 h at 40 °C. A salting procedure was then carried out by adding 1× PBS buffer containing 3.3 M NaCl stepwise over 45 min to the heated and stirred solution, until a final concentration of 750 mM NaCl was reached. Afterwards, the particles were mixed 1:1 with 1× PBS 10 mM NaCl, 2.11 mM P8709 buffer (Sigma Aldrich, USA), 2.89 mM P8584 buffer (Sigma Aldrich, USA), 0.01 % Tween ® 20 and 1 mM EDTA. To remove the excess thiolated single-stranded DNA, the solution was centrifuged for 15 min at 2.8 krcf and 20 °C. A pellet was formed in which the particles were concentrated. The supernatant was discarded, and the washing step was repeated six more times. After functionalization of the silver nanoparticles were diluted in 1× TE buffer (10 mM Tris, 1 mM EDTA) containing 750 mM NaCl to reach the final extinction of 0.05 (0.1 mm path length) at the extinction maxima on a UV-Vis spectrometer (Nanodrop 2000, Thermo Fisher Scientific, USA). Solution synthesis of DNA origami nanoantennas for TEM imaging Redding, B., Alam, M., Seifert, M. & Cao, H. High-resolution and broadband all-fiber spectrometers. Optica 1, 175 (2014). As shown in Figure2A, the droplet-based microfluidic device consisted of a drop inlet and an open-top islet immobilizing chamber connected by a capillary channel in between. With no external force and pressure driven, once a drop of glucose solution was loaded on top of the inlets, it would automatically flow towards the islets chamber compelled by the pressure generated by surface tension difference. By connecting this USB microscope to the compatible PC or phone via the USB connector, it allows you to see micro-object images through the screen and screenshots or screen record the video. It’s compatible with Windows, Mac OS, and Linux devices and Android smartphones with the OTG function. It is, however, not compatible with iPhone or iPad. Additionally, this microscope also features 8 built-in LEDs with adjustable brightness. It also comes with a stable alloy stand, an OTG adapter, a ruler, a driver, and a carrying bag. This microscope’s design is user-friendly, portable, and lightweight, easy to carry around with you anywhere. 5. Carson MicroFlip 100x-250x LEDJaadane, I. et al. Retinal damage induced by commercial light emitting diodes (LEDs). Free Radic. Biol. Med. 84, 373–384 (2015). As shown in Figure1B, the dichroic cube holding the excitation and emission filters was placed between the smartphone camera and the microfluidic biochip. The light emitted horizontally from the illumination source passed through the excitation filter and travelled into the dichroic cube. The dichroic mirror was placed inside of the cube at an angle of 45° to reflect the excitation light vertically to be casted upon the sample. The fluorescence signal within the sample exposed to the light radiated emission light, which returned through the emission filter and was then captured by the smartphone camera ( Figure1C). Ochmann, S. E. et al. Optical nanoantenna for single molecule-based detection of zika virus nucleic acids without molecular multiplication. Anal. Chem. 89, 13000–13007 (2017). Keiser, J., Utzinger, J., Premji, Z., Yamagata, Y. & Singer, B. H. Acridine Orange for malaria diagnosis: its diagnostic performance, its promotion and implementation in Tanzania, and the implications for malaria control. Ann. Trop. Med. Parasitol. 96, 643–654 (2002).
The smartphone frame was designed using SolidWorks (SOLIDWORKS Corp, Waltham, MA, USA) and 3D printed with polylactide resin (MakerBot ® PLA resin, MakerBot ® Industries, New York, NY, USA) using a MakerBot ® 3D printer (MakerBot ® Industries, New York, NY, USA). The smartphone sat on the top of the frame while a microfluidic biochip was inserted from the side ( Figure1A). The dimensions of the 3D frame were 100mm in height, 180mm in length, and 85mm in width. Design, fabrication, and validation of microfluidic biochip Département de Physique - Photonic Nanosystems, Université de Fribourg - Faculté des Sciences et Médicine, Fribourg, Switzerland Fluorescence microscopy produces visually attractive images that can greatly enhance the contrast of specific features of the specimen being viewed. This microscopic imaging modality has historically involved costly and cumbersome mercury arc lamps or lasers. However, in recent years, comparatively inexpensive and compact laser-emitting diode (LED) technology is replacing mercury arc lamps and lasers in the microscopy industry. Advantages include lower cost, greater longevity, and maintenance free operation. Because LEDs have been engineered to emit light of virtually any wavelength on the visible light spectrum, and can be filtered if needed, the microscopy community is widely adopting use of LEDs as an excitation light source for fluorescence microscopy. Schneider, C. A., Rasband, W. S. & Eliceiri, K. W. NIH Image to ImageJ: 25 years of image analysis. Nat. Methods 9, 671–675 (2012).
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Position the microscope lens in front of your phone’s standard camera lens and center it although I tried it with the telephoto lens and it works to yield different kinds of images Like its predecessors, the iMicro Q3 is compatible with virtually any smartphone equipped with a camera, attaching seamlessly through a reusable nano-suction pad. This compatibility ensures that the iMicro Q3 can be used with a range of devices, making it a versatile tool for microscopic observation. Cybulski, J. S., Clements, J. & Prakash, M. Foldscope: Origami-based paper microscope. PLoS ONE 9, e98781 (2014).
As shown in Figure5D, the fluorescence intensity increased to 151.8% (141.2% ± 17.0) in response to 250 μM Tolbutamide (a K ATP-channel closer). In Figure5E, when 200 μM Diazoxide (a K ATP-channel opener) was added after 14 mM glucose, the fluorescence intensity dropped to 108.8% (114.4% ± 13.2) from 133.8% (145.3% ± 14.7) at 2min and continued dropping at a relatively slower rate afterward. The iMicro Q3 offers advanced magnification capabilities and a unique optical structure for improved magnification and resolution. It boasts high-resolution power below a single micron and high magnification power up to 1200x, enabling the user to achieve resolution levels at the submicron scale. This capability overcomes the resolution constraints of traditional optical microscopes, making the iMicro Q3 a powerful tool for scientific observation. Department of Chemistry and Center for NanoScience, Ludwig-Maximilians-Universität München, München, Germany TEM grids (Formvar/carbon, 400 mesh, Cu, TedPella, Inc., USA) were Ar-plasma cleaned and incubated for 60 s with DNA origami sample (5 µL, ~ 2–10 nM). Grids were washed with 2 % uranyl formate solution (5 µL) and incubated again afterwards again 4 s with 2% uranyl formate solution (5 µL) for staining. TEM imaging were performed on a JEM-1100 microscope (JEOL GmbH, Japan) with an acceleration voltage of 80 kV. Sample preparation on the coverslip for single-molecule confocal measurements A YFP filter set (Iridian spectral technology, Ottawa, ON, Canada) was used to observe the Rhodamine-123. The filter set had an excitation band of 489-505 nm, and an emission band of 524-546 nm. The camera settings used for Rhodamine-123 was identical to the ones used for Fluo-4 and the GEFPI. Single islet insulin secretionGill, P. & Ghaemi, A. Nucleic acid isothermal amplification technologies: a review. Nucleosides Nucleotides Nucleic Acids 27, 224–243 (2008). Lepeuple, A.-S., Gilouppe, S., Pierlot, E. & De Roubin, M.-R. Rapid and automated detection of fluorescent total bacteria in water samples. Int. J. Food Microbiol. 92, 327–332 (2004). In this study, we introduced a novel smartphone-microfluidic fluorescence imaging system to study the physiology of islet beta-cells. By trapping islets in a customized surface tension driven pumpless microfluidic device, we managed to use the smartphone camera to visualize mouse islets labeled with various fluorescence indicators and detected physiological insulin stimulator-secretion coupling factors, showing decent fluorescence signaling and signal vs. noise ratio in response to different stimuli/inhibitors. Our system can also achieve adequate resolution of single islet insulin secretion. For the sandwich assay in serum clotted, whole blood, sterile and filtered human blood serum (Human Serum, (from male AB clotted whole blood), USA origin, sterile-filtered, Sigma-Aldrich, USA) was used. Before adding the serum to the NACHOS and reference samples, the serum was heat inactivated by exposing it for 30 min to 56 °C and spiked with 2 nM target DNA, 6 nM imager strand and 2 M NaCl. The fully assembled NACHOS or reference DNA origami structures were incubated with target-spiked blood serum for 2 h at 37 °C and the excess of target and imager strands was removed by washing six times with 1× TE buffer containing 2 M NaCl. NACHOS were then imaged in 1× TE buffer containing 14 mM MgCl 2. Confocal measurements and data analysis
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