Depth of Field Calculator

Calculate the depth of field for popular objectives with this interactive tutorial.

Eyepiece Reticle Calibration

Explores calibration of various eyepiece reticles using a stage micrometer and demonstrates how the reticle can then be employed to determine linear specimen dimensions.

Field of View Diameter

Exploring the effect of varying the field of view size on the viewable specimen area.

Immersion Oil and Refractive Index

The refractive index is critical in determining the working numerical aperture of an objective.

Inverted Microscope Optical Pathways

Examine an animated cut-away diagram of a modern inverted microscope.

Microscope Alignment for Köhler Illumination

Learn how to adjust a microscope to examine specimens in Köhler illumination.

Microscope Conjugate Field Planes

The geometrical relationship between image planes in the optical microscope.

Numerical Aperture and Image Resolution

Explore how objective numerical aperture size influences Airy disk properties.

Objective Working Distance

Examine the range of working distance values available in modern Nikon CFI60 objectives.

Photomask Reticle Operation

Practice adjustment of the photomask reticle in a focusing eyepiece.

Proper Microscope Posture

Explore proper head and back posture for microscope observations.

Refraction of Light

Examine how refractive index varies with the dispersion of properties of different materials.

Stereomicroscope Optical Pathways

Examine the light pathways in a Nikon stereomicroscope.

Useful Magnification Range

Determine if objective-eyepiece combinations are in the useful magnification range.

Integrated Circuit Inspection

Explore real-time confocal microscopy of integrated circuits by adjusting focus depth.

Lambda Stack Basic Concepts

The lambda stack is a three-dimensional dataset that consists of an image collection using the same specimen field acquired at different wavelength bands, each spanning a limited spectral region ranging from 2 to 20 nanometers.

Lambda Stacks and Spectral Signatures

Learn how individual spectra are represented as narrow wavebands in lambda stacks.

Nikon Diffraction Efficiency Enhancement System (DEES)

The purpose of the Nikon DEES system is to increase the efficiency of light diffraction by the gratings used to separate fluorescence emission into component wavelengths.

Resonant Scanning Confocal Microscope Zoom

Examine how zoom magnification is implemented in resonant scanning laser confocal microscopy using Ronchi gratings.

Resonant Scanning in Laser Confocal Microscopy

A look at the technology behind confocal resonant scanning systems.

Spectral Imaging with FRET Biosensors

Spectral imaging is of significant advantage in separating the overlapping emission spectra of fluorescent proteins and other fluorophores in dynamic fluorescence resonance energy transfer (FRET) experiments, which are often complicated by the requirement for exceedingly fast image capture.

Spectral Imaging with Linear Unmixing

Discover how individual fluorophores can be identified within a complex mixture.

CCD Noise Sources

A review of the types of noise sources associated with digital imaging.

CCD Resolution for Optical Microscopy

Matching digital camera and microscope resolution.

CCD Signal-To-Noise Ratio

Signal-to-noise represents the ratio of the measured light signal to the combined noise.

Color Balance in Digital Imaging

White and black balance settings on a digital camera can be used to adjust color balance.

Full-Frame CCD Operation

The readout scheme of a full-frame charge coupled device (CCD).

Matching Camera to Microscope Resolution

Vary numerical aperture, magnification, and video coupler size to match camera resolution.

Proximity-Focused Image Intensifiers

Light amplification with a micro-channel plate and photocathode.

Spatial Resolution in Digital Imaging

Spatial resolution refers to the number of pixels utilized in construction of the image.

Bias Retardation Effects on Specimen Contrast

Explore the effects of varying bias retardation on specimen contrast.

DIC Microscopy with de Sénarmont Compensators

A three-dimensional tutorial illustrating the de Sénarmont DIC wavefront field.

Nomarski Prism Action in Polarized Light

Variations in prism geometry yield unique interference patterns in polarized light.

Optical Sectioning in Reflected Light DIC

Optical sectioning of reflected light specimens (semiconductors).

The de Sénarmont DIC Microscope Optical Train

Examines the relationship between the wavefront fields emerging from a de Sénarmont compensator and how they can be controlled to produce positive and negative bias retardation (contrast) effects in a DIC microscope.

Wavefront Relationships in de Sénarmont and Nomarski DIC

An interactive comparison of wavefronts in these complementary techniques.

Wavefront Relationships in Reflected Light DIC Microscopy

Observe how light waves travel through a reflected light DIC microscope.

Balancing Arc-Discharge Lamp Excitation Illumination

Excitation filter balancers can be used to fine-tune fluorophore excitation.

Blue Excitation

The Nikon blue excitation fluorescence filter combinations include bandpass and longpass sets having both broad and narrow excitation bandwidths.

Blue-Violet Excitation

Exploring how the variations in the excitation and emission filter spectral profiles, as well as those of the dichromatic mirrors, affect signal levels, overall filter performance, and image contrast in combinations designed for excitation of fluorophores in the blue-violet region.

Dual Band Excitation

Explore how the variations in the excitation and emission filter spectral profiles affect signal levels, overall filter performance, and image contrast in combinations designed for dual excitation of fluorophores in the ultraviolet and blue or blue and green regions.

Fluorescence Filter Noise Terminator

Fluorescence filter block design that eliminates the possibility of residual stray light.

Fluorescent Protein Biosensors

Descriptions of selected fluorescent protein biosensor mechanisms.

Focus and Alignment of Mercury and Xenon Arc Lamps

Explore alignment and focusing of the arc lamp in a mercury or xenon burner, which simulates how the lamp is adjusted in a real microscope.

Green Excitation

Explore how the variations in the excitation and emission filter spectral profiles affect signal levels, overall filter performance, and image contrast in combinations designed for excitation of fluorophores in the green (510-560 nanometers) spectral region.

Matching Fluorescent Probes with Nikon Fluorescence Filter Blocks

Explore the various fluorophores that can be imaged with Nikon filter sets.

Stereomicroscopy Fluorescence

Explore the light paths in Nikon's SMZ1500 stereomicroscope equipped for fluorescence illumination using an intermediate tube and external lamphouse.

Triple Band Excitation

Filter sets for DAPI, FITC, and TRITC or Texas Red.

Ultraviolet Excitation

Examine specimen contrast with longpass and shortpass filter sets.

Violet Excitation

Discussion of the properties of various fluorescence filter combinations.

Yellow Excitation

Specimen contrast variations with narrow and wide bandpass filter combinations.

Adjustment of Objective Correction Collars

Learn how to adjust an objective correction collar to minimize spherical aberration.

Choosing Filter Combinations for Fluorescent Proteins

Identification of critical filter parameters for imaging fluorescent proteins.

Choosing Fluorescent Proteins for Dual Labeling Experiments

Interactive tutorial used to optimize pairing of two fluorescent proteins.

Comparison of Phase Contrast & DIC Microscopy

Examples of the same specimen viewed in either phase contrast or DIC.

DIC Microscope Component Alignment

Examine conoscopic and orthoscopic viewfields in DIC microscopy.

Focus and Alignment of Mercury and Xenon Arc Lamps

Explore alignment and focusing of the arc lamp in a mercury or xenon burner, which simulates how the lamp is adjusted in a real microscope.

Förster (Fluorescence) Resonance Energy Transfer with Fluorescent Proteins

Use this tool to determine the optimum fluorescent protein pairs for FRET.

Laser Scanning Confocal Microscopy

A virtual microscope tutorial featuring a wide variety of specimens.

Matching Camera to Microscope Resolution

Vary numerical aperture, magnification, and video coupler size to match camera resolution.

Matching Fluorescent Probes with Nikon Fluorescence Filter Blocks

Explore the various fluorophores that can be imaged with Nikon filter sets.

Microscope Alignment for Köhler Illumination

Learn how to adjust a microscope to examine specimens in Köhler illumination.

Optical Sectioning with de Sénarmont DIC Microscopy

At high numerical apertures, DIC can be used for optical sectioning.

Perfect Focus Offset System Mechanics

Learn how the offset lens system enables the operation of the Nikon PFS.

Phase Contrast Microscope Alignment

Learn how to align a phase contrast microscope and examine variations in specimen appearance through the eyepieces (at different magnifications) when the condenser annulus is shifted into and out of alignment with the phase plate in the objective.

Phase Plate Configuration Effects on Specimen Contrast

Examine contrast variations induced by altering phase plate absorption properties.

Proximity-Focused Image Intensifiers

Light amplification with a micro-channel plate and photocathode.

Spatial Resolution in Digital Imaging

Spatial resolution refers to the number of pixels utilized in construction of the image.

The Nikon Perfect Focus System (PFS)

Examine how the Nikon PFS system operates to maintain stable focus in live-cell imaging.

Chromatic Aberration

Explore axial and lateral chromatic aberrations seen in an optical microscope with this interactive tutorial.

Condenser Image Planes

Examine the relationship between field and condenser diaphragm image planes.

Coverslip Correction Collars

Investigate how internal lens elements in a high numerical aperture dry objective may be adjusted to correct for fluctuations in coverslip thickness.

Field Curvature

Common aberration caused by the spherical surface of lens elements.

Field of View Diameter

Exploring the effect of varying the field of view size on the viewable specimen area.

Geometrical Construction of Ray Diagrams

A popular method of representing a train of propagating light waves.

Geometrical Distortion

Termed pincushion or barrel, geometrical distortion often occurs in stereomicroscopy.

Immersion Oil and Refractive Index

The refractive index is critical in determining the working numerical aperture of an objective.

Infinity-Corrected Microscope Conjugate Field Planes

Objective light is focused to infinity and the tube lens forms the image at its focal plane.

Microscope Alignment for Köhler Illumination

Learn how to adjust a microscope to examine specimens in Köhler illumination.

Microscope Conjugate Field Planes

The geometrical relationship between image planes in the optical microscope.

Modulation Transfer Function: Diffraction Effects on Image Contrast

This tutorial reviews the modulation transfer function with respect to contrast.

Numerical Aperture Light Cones

Numerical aperture is a measure of the highly diffracted light rays captured by the objective.

Objective Working Distance

Examine the range of working distance values available in modern Nikon CFI60 objectives.

Perfect Lens Characteristics

An ideally corrected lens that is free of aberration and focuses light to a point.

Perfect Two-Lens System Characteristics

Explore how off-axis oblique light waves pass through a two-lens system.

Specimen Contrast Enhancement with Apodized Phase Plates

Examine a variety of specimens using traditional and apodized phase contrast microscopy.

Tube Lens Focal Length

Infinity optical systems feature tube lengths between 200 and 250 millimeters.

Viewing and Projection Eyepieces

A discussion of projection and viewing eyepiece systems.

Birefringence in Calcite Crystals

Examine how crystals of the mineral calcite demonstrate birefringence.

Birefringent Crystals in Polarized Light

Explore how birefringent anisotropic crystals interact with polarized light in an optical microscope as the circular stage is rotated through 360 degrees.

Polarized Light Waveforms

This interactive tutorial explores the generation of linear, elliptical, and circularly polarized light by a pair of orthogonal light waves (as a function of the relative phase shift between the waves) when the electric field vectors are added together.

Polarizer Rotation and Specimen Birefringence

Discover how specimen birefringence is affected by the angle of polarizer when observed in a polarized light microscope.

Apodized Phase Contrast

Explore how specimen size affects the angle of diffracted light rays that pass through apodized phase plates.

Interaction of Light Waves with Phase Specimens

Light waves are deformed by specimen geometry, refractive index, and thickness.

Optical Pathways in the Phase Contrast Microscope

Examine the light pathways through a phase contrast microscope and learn how these systems dissect the incident electromagnetic wave into a surround (S), diffracted (D), and resultant particle (P) component.

Phase Contrast Microscope Alignment

Learn how to align a phase contrast microscope and examine variations in specimen appearance through the eyepieces (at different magnifications) when the condenser annulus is shifted into and out of alignment with the phase plate in the objective.

Positive and Negative Phase Contrast

This interactive tutorial explores relationships between the surround (S), diffracted (D), and resulting particle (P) waves in brightfield as well as positive and negative phase contrast microscopy.

Shade-Off and Halo Phase Contrast Artifacts

Explore shade-off and halo artifacts, where the observed intensity does not directly correspond to the optical path difference (refractive index and thickness values) between the specimen and the surrounding medium.

Specimen Optical Path Length Variations

Explore the effects of changes to refractive index and thickness on optical path length, and discover how two specimens can have different combinations of these variables but still display the same path length.

Focus and Alignment of Mercury and Xenon Arc Lamps

Explore alignment and focusing of the arc lamp in a mercury or xenon burner, which simulates how the lamp is adjusted in a real microscope.

Oblique Coherent Contrast Illumination

Enhancing contrast in stereomicroscopy with transmitted light.

SMZ-1500 Oblique Coherent Contrast Illumination

Introduced with the Nikon SMZ1500 stereomicroscope, Nikon's proprietary Oblique Coherent Contrast illumination system optimizes contrast in transmitted stereoscopic microscopy.

SMZ1500 Stereoscopic Microscope

Explore focus and zoom settings in a virtual stereomicroscope.

Stereomicroscope Optical Pathways

Examine the light pathways in a Nikon stereomicroscope.

Stereomicroscopy Fluorescence

Explore the light paths in Nikon's SMZ1500 stereomicroscope equipped for fluorescence illumination using an intermediate tube and external lamphouse.

Toroidal Mirrors

Explore how mirror shape affects the amount of light entering the objective in darkfield stereoscopic microscopy. This tutorial demonstrates lightpath differences between conventional and toroidal mirrors.

A Comparison of STED and STORM Super-Resolution Imaging

Contrast between raster scanning and single-molecule imaging for superresolution.

Molecular Density in Super-Resolution Microscopy

The density of localized molecules affects the ultimate image resolution.

Stochastic Optical Reconstruction Microscopy (STORM) Imaging

Review the basic steps involved in acquiring a STORM super-resolution image.

Super-Resolution Microscopy via Stochastic Individual Molecule Blinking

Utilization of generic dark states to generate single-molecule superresolution images.