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Application Notes

At Nikon we are committed to providing resources to researchers for performing cutting-edge imaging techniques and assays on our various research microscope platforms.  Provided here is a collection of Application Notes – short but detailed works focusing on modern applications in the constantly growing and evolving field of biological imaging.

Questions, comments, or concerns?  Please feel free to contact us at nikoninstruments@nikon.net or 1-800-52-NIKON.

A1+

The A1+ is Nikon's powerful, galvano scanner-based confocal system with GaAsP detectors for high-resolution, high-sensitivity imaging.

Reflectance imaging for visualization of unlabeled structures using Nikon A1R and N-SIM

download: English (3.43 MB)

Reflectance imaging for visualization of unlabeled structures using Nikon A1 and N-SIM

Reflectance imaging allows the user to form an intensity image from light backscattered by the sample.  Highly reflective markers, including a variety of nanoparticles, allows for imaging with very high signal-to-noise and virtually free of photobleaching, ideal for both confocal and structured illumination microscopies.


A1R HD

The A1R HD is a high-performance confocal system with high-definition resonant scanner for beautifully clear images at high speeds. Ultra-resolution galvano scanner provides incredible resolution over a large field of view. Hybrid scanner enables simultaneous photo stimulation and imaging while GaAsP detectors provide exceptional sensitivity.

Reflectance imaging for visualization of unlabeled structures using Nikon A1R and N-SIM

download: English (3.43 MB)

Reflectance imaging for visualization of unlabeled structures using Nikon A1 and N-SIM

Reflectance imaging allows the user to form an intensity image from light backscattered by the sample.  Highly reflective markers, including a variety of nanoparticles, allows for imaging with very high signal-to-noise and virtually free of photobleaching, ideal for both confocal and structured illumination microscopies.


A1R MP+ Multiphoton

Nikon’s A1R MP+ confocal microscope is a unique multiphoton imaging system featuring a fast, high resolution galvanometer scanner and an ultra-high speed resonant scanner.

Nikon’s large-format multiphoton system for intravital imaging

download: English (0.27 MB)

Nikon’s large-format multiphoton system for intravital imaging

Nikon’s A1R MP multiphoton confocal microscope system provides a choice platform for deep tissue imaging.  Researchers at the Allen Institute for Brain Science have applied the A1R MP towards fast dual-color intravital imaging of the mouse brain.  The open architecture instrument design allows for the addition of custom stages for large samples, external photostimulation devices, and more – perfect for constantly evolving needs.


BioStation CT Cell Culture Observation

The BioStation CT is a self-contained, automated cell culture monitoring system that allows users to manage and observe multiple live cell imaging studies locally or by remote operation over a public or private network.

Survival Analysis of Human In Vitro-derived Neurons Using New Live Cell Extended Time-lapse Imaging Technology

download: English (2.65 MB)

Survival Analysis of Human In Vitro-derived Neurons Using New Live Cell Extended Time-lapse Imaging Technology

In this application note we demonstrate the Nikon BioStation CT for long-term live-cell imaging of stem cell-derived motor neurons – tracking key attributes such as cell body size and neurite length before, during, and after treatment with different stressors. This approach provides a standardized method for drug discovery studies in neurodegenerative disease models.


Development of an Integrated Bioprocess for Production of NK-92 Cells for Immunotherapy

download: English (5.69 MB)

Development of an Integrated Bioprocess for Production of NK-92 Cells for Immunotherapy

Anti-cancer NK-92 lymphocytes are promising for application in immunotherapies.  This application note explores a comprehensive workflow for producing clinical-grade NK-92 cells, using Nikon’s BioStation CT to monitor NK-92 phenotype and potency.


Embryonic Stem Cell Responses to Commercially Available Substrates for Stem Cell Maintenance

download: English (1.78 MB)

Embryonic Stem Cell Responses to Commercially Available Substrates for Stem Cell Maintenance

In collaboration with the Centre for Commercialization of Regenerative Medicine (CCRM) we compare the performance of 8 common substrates for stem cell growth and maintenance by developing an assay utilizing the Nikon BioStation CT high content screening microscope and incubation system. Fluorescence imaging of Oct4 is used as a marker for pluripotency.


Establishment of an Algorithm for Automated Detection of iPS/non-iPS Cells Under a Culture Condition by Non-Invasive Image Analysis

download: English (2.68 MB)

Establishment of an Algorithm for Automated Detection of iPS/non-iPS Cells Under a Culture Condition by Non-Invasive Image Analysis

In this application note we detail an automated high throughput method for observing and classifying stem cell colonies with a standardized algorithm and using the Nikon BioStation CT. Automated image analysis discriminates stem cell colonies from non-stem cell colonies using phase contrast imaging and subsequent analysis of distinguishing morphological features.


Automated, Non-Invasive Culture, and Evaluation System for iPS Cells under Neural Differentiation Process

download: English (2.99 MB)

Automated, Non-Invasive Culture, and Evaluation System for iPS Cells under Neural Differentiation Process

This application note details an integrated workflow for the culture, observation, evaluation, and differentiation of induced pluripotent stem (iPS) cells into neurons, introducing image processing methods for detecting neural rosettes and neurite outgrowth – two important morphological checkpoints in neural differentiation. Automated imaging for this assay is performed using a Nikon BioStation CT.


Generation of a Growth Curve for iPS Cells in a Feeder-Free Culture by Non-Invasive Image Analysis

download: English (2.61 MB)

Generation of a Growth Curve for iPS Cells in a Feeder-Free Culture by Non-Invasive Image Analysis

Long-term time-lapse observation of feeder layer-free stem cell cultures, along with automated detection of stem cell colonies, is demonstrated here using a Nikon BioStation CT system.  Growth curves for both differentiated and un-differentiated cells were generated for a time period of about 1 week.


A Non-Invasive Method for Counting Human Pluripotent Stem Cell Numbers by Live Cell Imaging

download: English (1.67 MB)

A Non-Invasive Method for Counting Human Pluripotent Stem Cell Numbers by Live Cell Imaging

Presented here is a method for counting live pluripotent stem cells using the fluorescent nuclear stain SYTO24 and phase contrast imaging on the Nikon BioStation CT high content screening microscope. The results show that colony coverage area deduced from phase contrast images can be used as an accurate estimator of stem cell count alone, correlating well with haemocytometer-based measurements.


Automated Haematopoietic Colony Forming Cell (CFC) Assay

download: English (0.6 MB)

Automated Haematopoietic Colony Forming Cell (CFC) Assay

Nikon’s BioStation CT high content live cell screening system is applied in an assay for automated enumeration and identification of haematopoietic colony-forming cells.  This assay allows for the identification of multiple colony types, helping distinguish between lineages.


Antitumor Lymphocyte Kinetic Cytotoxicity Assay (Collaboration with CCRM)

download: English (0.69 MB)

Antitumor Lymphocyte Kinetic Cytotoxicity Assay (Collaboration with CCRM)

In this application note we explore the use of the fully robotic Nikon BioStation CT high content screening microscope platform towards quantifying the death rate of a model human leukemia cell line upon co-incubation with anticancer NK-92 lymphocytes.  This assay integrates automated imaging of Calcein-AM fluorescence to perform live/dead cell analysis.


Screening Assay for PSC Responses to Substrates

download: English (0.47 MB)

Screening Assay for PSC Responses to Substrates


Stem Cell Research with Nikon

download: English (0.86 MB)

Stem Cell Research with Nikon


Outline of Stem Cell applications (written in both English and Japanese)

download: English (2.72 MB)

Outline of Stem Cell applications (written in both English and Japanese)


Eclipse Ti2 Inverted Microscope

The Eclipse Ti2 is Nikon's all-new inverted research microscope featuring never-before-achieved FOV imaging capacity.

Application of Patterned Illumination Using a DMD for Optogenetic Control of Signaling

download: English (0.22 MB)

Application of Patterned Illumination Using a DMD for Optogenetic Control of Signaling

Digital micromirror devices (DMDs) are powerful tools for photostimulation applications, including photoconversion and optogenetic manipulation, owing to their robust ability to produce novel illumination patterns with high spatiotemporal resolution. In this Application Note we showcase recent work describing how DMD technology integrated into a Nikon system can be applied toward light-gated optogenetic control of intracellular signaling.


Hardware Triggering: Maximizing Speed and Efficiency for Live Cell Imaging

download: English (0.31 MB)

Hardware Triggering: Maximizing Speed and Efficiency for Live Cell Imaging

Live cell imaging experiments now require higher speeds and more data throughput than ever before. Nikon Instruments has robust tools that enable hardware triggering of imaging devices in microscopy via direct signaling between hardware. This minimizes delays, synchronizes devices, and reduces the exposure of specimens to light. This Application Note explains how Nikon’s NIS-Elements hardwaretriggering workflow operates, and details its benefits for common time-lapse acquisition routines.


Application Note Preview

download: English (0.41 MB)

Robotic Microscopy with the Nikon Ti2 for High-Content Analysis Applications

Robotic Microscopy—a combination of high-content screening methods—enables multivariate experimental approaches with large cell populations and member-level sensitivity. Here we explore how the new Nikon Ti2 line of inverted research microscopes is uniquely suited to Robotic Microscopy applications, focusing on work utilizing induced pluripotent stem cells (iPSCs) as disease models in drug screening.


HCA High Content Analysis

Nikon’s High Content Screening Microscope is an ultra-high-speed imaging system with 7 available excitation wavelengths that can meet most high-content imaging requirements.

Application Note Preview

download: English (0.41 MB)

Robotic Microscopy with the Nikon Ti2 for High-Content Analysis Applications

Robotic Microscopy—a combination of high-content screening methods—enables multivariate experimental approaches with large cell populations and member-level sensitivity. Here we explore how the new Nikon Ti2 line of inverted research microscopes is uniquely suited to Robotic Microscopy applications, focusing on work utilizing induced pluripotent stem cells (iPSCs) as disease models in drug screening.


Identification of astrocytes derived from neural progenitor cells by high-throughput screening using a live cell mRNA detection technology

download: English (2.13 MB)

Identification of astrocytes derived from neural progenitor cells by high-throughput screening using a live cell mRNA detection technology

This application note details an automated experimental approach for identifying astrocytes through detection of GFAP mRNA using the live-cell SmartFlareTM fluorescence-based RNA detection technology in combination with Nikon’s High Content Analysis (HCA) software.  Nikon’s HCA software provides a platform for performing both automated image acquisition and analysis in high throughput screening applications.


N-SIM and N-SIM E Super-Resolution

Nikon’s N-SIM and N-SIM E super-resolution systems double the resolution of conventional optical microscopes by combining “Structured Illumination Microscopy” technology with renowned Nikon objective lenses.

Reflectance imaging for visualization of unlabeled structures using Nikon A1R and N-SIM

download: English (3.43 MB)

Reflectance imaging for visualization of unlabeled structures using Nikon A1 and N-SIM

Reflectance imaging allows the user to form an intensity image from light backscattered by the sample.  Highly reflective markers, including a variety of nanoparticles, allows for imaging with very high signal-to-noise and virtually free of photobleaching, ideal for both confocal and structured illumination microscopies.


N SIM Application Note

download: English (2.87 MB)

N-SIM for Quantitative Ultra-Structural Analyses of the Nuclear Lamina

Super-resolution Structured Illumination Microscopy (SIM), available from Nikon via the N-SIM and N-SIM E systems, allows for the observation of details inaccessible to traditional microscopes, such as confocal and widefield. In this application note we see how the N-SIM system enables quantitative multi-color evaluation of the distribution of different nuclear lamin proteins and the structures they form.


N-STORM Super-Resolution

Nikon N-STORM super-resolution microscopes offer an order of magnitude improvement in resolution compared to a conventional light microscope.

Quantitative Analysis Tools and Correlative Imaging Applications for N-STORM

download: English (0.45 MB)

Quantitative Analysis Tools and Correlative Imaging Applications for N-STORM

Stochastic Optical Reconstruction Microscopy (STORM) has had a significant impact on our understanding of nano-scale biological processes. In this application note, we see how Nikon’s N-STORM system, combined with quantitative analysis tools, enables single molecule level studies of cannabinoid signaling in the brain. Correlative 3D-STORM, confocal microscopy, and patch-clamp electrophysiology is also explored.


Quantitative Cluster Analysis Applications for N-STORM

download: English (2.66 MB)

Quantitative Cluster Analysis Applications for N-STORM

Nikon’s N-STORM super-resolution microscope is applied to mapping the distribution of nucleosomes on chromatin fibers, shining a light on the mechanisms of chromatin folding, gene expression, and pluripotency. Multicolor STORM is used to explore how nucleosome distribution differs between differentiated and pluripotent stem cells and correlates with the distribution of other important factors in replication, such as RNA Polymerase II.


N-STORM with DNA-PAINT for Reliable Multicolor & 3D Single Molecule Localization Imaging

download: English (1.73 MB)

N-STORM with DNA-PAINT for Reliable Multicolor & 3D Single Molecule Localization Imaging

DNA-PAINT is a technique for single molecule localization microscopy.  The sample is immunolabeled with specialized secondary antibodies conjugated to short DNA oligomers known as docking strands.  Complimentary oligomers conjugated to fluorophore, known as imaging strands, are introduced in the imaging buffer and will transiently bind to docking strands – allowing single molecules to be localized.  Herein multicolor and 3D DNA-PAINT is demonstrated on the Nikon N-STORM 4.0 system.


NIS-Elements

Nikon's NIS-Elements revolutionizes imaging software for the microscopy market by combining automated intelligence to microscopes, cameras, components and peripherals with powerful archiving, analysis and visualization tools. Its intuitive interface simplifies workflow and speeds up image acquisition times while providing powerful features such as image stitching, object counting and volume views.

Hardware Triggering: Maximizing Speed and Efficiency for Live Cell Imaging

download: English (0.31 MB)

Hardware Triggering: Maximizing Speed and Efficiency for Live Cell Imaging

Live cell imaging experiments now require higher speeds and more data throughput than ever before. Nikon Instruments has robust tools that enable hardware triggering of imaging devices in microscopy via direct signaling between hardware. This minimizes delays, synchronizes devices, and reduces the exposure of specimens to light. This Application Note explains how Nikon’s NIS-Elements hardwaretriggering workflow operates, and details its benefits for common time-lapse acquisition routines.


Ti-LAPP

The Nikon Ti-LAPP system provides modular illuminators for total internal reflection fluorescence (TIRF), photoactivation/conversion, photobleaching and epi-fluorescence. Each module can be flexibly combined to build microscope systems that are optimized for individual research needs.

Application of Patterned Illumination Using a DMD for Optogenetic Control of Signaling

download: English (0.22 MB)

Application of Patterned Illumination Using a DMD for Optogenetic Control of Signaling

Digital micromirror devices (DMDs) are powerful tools for photostimulation applications, including photoconversion and optogenetic manipulation, owing to their robust ability to produce novel illumination patterns with high spatiotemporal resolution. In this Application Note we showcase recent work describing how DMD technology integrated into a Nikon system can be applied toward light-gated optogenetic control of intracellular signaling.



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