Confocal Microscopy Interactive Tutorials

Confocal Microscope Simulators

• Laser Scanning Confocal Microscope Simulator

  Explore multi-laser fluorescence and differential interference contrast (DIC) confocal imaging using the Olympus FluoView FV1000 microscope software interface.

• Comparing Confocal and Widefield Fluorescence Microscopy

  Discover and compare the differences between specimens when viewed in a confocal versus a widefield fluorescence microscope in this interactive tutorial.

• Spectral Bleed-Through (Crossover) in Confocal Microscopy

  Explore spectral bleed-through in laser scanning confocal microscopy and the mechanisms available to reduce or eliminate the artifact in this interactive tutorial.

Confocal Microscope Systems

• Laser Confocal Microscopy Scanning Modes

  Laser scanning modes enable investigators to optimize data collection for three-dimensional imaging. Examine various scanning mechanisms in confocal microscopy utilizing a cube-shaped virtual specimen.

• Galvanometer-Based Confocal Scanning Systems

  Explore how the scanning mirrors are coordinated to direct the laser beam into the objective, and then to reflect secondary fluorescence to the emission filter.

• Scanning System Basics

  Examine how the galvanometer-driven mirrors and optical system of a confocal microscope are configured to enable the objective rear aperture to be continuously filled with light during the raster scanning operation in this tutorial.

• Laser Scanning Confocal Microscopy

  (approximately a 30 second download on 28.8K modems)
  Explore imaging specimens through serial z-axis optical sections utilizing a virtual confocal microscope in this interactive tutorial.

Lasers for Confocal Microscopy

• Argon-Ion Lasers

  Learn more in this tutorial about the well-explored family of ion lasers and how the argon-ion laser operates in the visible and ultraviolet spectral regions by utilizing an ionized species of the noble gas argon.

• Diode Lasers

  Learn more in this interactive tutorial about semiconductor diode lasers having sufficient power output to be useful in optical microscopy are now available from a host of manufacturers

• Helium-Cadmium Lasers

  Explore a simplified model of the helium-cadmium laser cavity operation by using a cutaway drawing of a helium-cadmium laser illustrated in the central window in this interactive tutorial.

• Helium-Neon Lasers

  Learn more about the most widely utilized laser system with compact size dimensions, helium-neon laser, which is known for having the most superior Gaussian beam quality in this interactive tutorial.

• Krypton-Argon Lasers

  Discover and learn more in this interactive tutorial as it simulates the three major spectral lines produced by an krypton-argon mixed-gas laser used in confocal microscopy.

• Acousto-Optic Tunable Filters

  Explore more about electro-optic devices, such as the acousto-optic tunable filter (AOTF), which are increasingly being employed to modulate the wavelength and amplitude of illuminating laser light in confocal microscopes.

• Stimulated Emission in a Laser Cavity

  Discover how laser amplification occurs starting from spontaneous emission of the first photon to saturation of the laser cavity and the establishment of a dynamic equilibrium state in this interactive tutorial.

• Pockels Cell Laser Modulators

  The beam intensity of lasers can be stabilized by electronic control of the tube current or utilization of external components. Examine how the Pockels cell modulator operates to stabilize laser beam intensity.

• Laser Energy Levels

  Population inversion can be produced in a laser through two basic mechanisms. Explore metastable states for both three-level and four-level laser systems in this interactive java tutorial.

• Laser Cavity Resonance Modes and Gain Bandwidth

  Explore how varying the appropriate frequencies can alter curves describing the number of cavity modes and gain bandwidth of a laser in this featured interactive java tutorial.

Fluorophores for Confocal Microscopy

• Fluorescent Probe Excitation Efficiency

  The absorption and fluorescence emission spectral profiles of a fluorophore are important when selecting probes for laser scanning. See how this relationship can be used to match fluorophores with specific lasers.

• Optical Highlighter Fluorescent Proteins

  Explore the optical conversion of several useful highlighter probes and it simulates how these proteins would be viewed in an actual confocal microscope in this interactive tutorial.

• Colocalization of Fluorophores in Confocal Microscopy

  Analyze colocalization in a wide spectrum of specimens that were designed to demonstrate the phenomenon, or to provide examples of targets that lack any colocalization.

• Choosing Fluorophore Combinations for Confocal Microscopy

  Explore the matching of dual fluorophores with laser excitation lines, calculation of emission spectral overlap values, and determination of the bleed-through level.

• Fluorescent Protein Fluorophore Maturation Mechanisms

  The interactive tutorials in this section explore fluorophore formation in a wide variety of spectrally diverse fluorescent proteins deduced from crystallographic studies.

• Excitation Photobleaching Patterns

  Compare and contrast excitation-induced photobleaching patterns that occur near the focal region in both multiphoton and confocal microscopy systems in this interactive java tutorial.

Resolution, Contrast, and Sampling in Confocal Microscopy

• Airy Pattern Basics

  This tutorial explores how Airy pattern size changes with objective numerical aperture and the wavelength of illumination; it also simulates the close approach of two Airy patterns.

• Airy Patterns and the Rayleigh Criterion

  This tutorial examines how Airy disk sizes, at the limit of optical resolution, vary with changes in objective numerical aperture and illumination wavelength and how these changes affect the resolution of the objective.

• Airy Patterns and Resolution Criteria (3-D Version)

  Explore with a three-dimensional model how Airy disk sizes, at the limit of optical resolution, vary with changes in objective numerical aperture and illumination wavelength.

• Reflected Confocal Microscopy: Integrated Circuit Inspection

  Examine layers on the surface of integrated circuits with this tutorial as it simulates imaging of integrated circuits using a reflected light Nipkow disk Confocal Microscope.

Confocal Microscope Detectors

• Side-On Photomultipliers

  In the side-on photomultiplier tube design, photons impact an internal photocathode and eject electrons from the front face (as opposed to the rear side as in the end-on designs) as shown in this tutorial.

• Channel Photomultipliers

  Each time an electron impacts an inner wall of the channel, multiple secondary electrons are emitted. Learn and explore how electrons are multiplied within the conductive chain of a channel photomultiplier in this interactive tutorial.