00_LSM Capabilities_1652.qxd.P:1652 7/29/11 8:26 AM Page 1652 Imaging Thorlabs’ Imaging group is a world-wide team dedicated to the development of Optical Coherence Tomography Systems, Microscopy Systems, and Adaptive Optics. The group is comprised of highly skilled engineers with expertise in optical systems design, mechanical and electrical systems integration, and software development. Through close collaboration with leading researchers, the group develops imaging systems and components to help advance the research endeavors of our customers. Many of our systems are customizable to meet various application needs. We encourage you to start a dialog about how we can best help fulfill your imaging needs. Expertise in Custom and OEM Product Development • Advanced Optical and Imaging Systems • Software Development • Modular System Designs are Easily Customizable In-House Design and Manufacturing Capabilities • Custom and OEM Opportunities • On-Site Demos, Loans, and Leasing Options Available Precision Testing of a Custom Fast Steering Mirror ThorImageLSTM Software Interface for Thorlabsʼ Laser Scanning Microscopy Systems Optical Coherence Tomography Research Lab 1652 www.thorlabs.com

Selection Guide Laser Scanning Microscopy Selection Guide Multiphoton Systems Multiphoton Accessories Confocal Systems Laser Scanning Essentials Kit

01_LSM Tutorial_1653-1659.qxd.P:1653-1659 7/26/11 2:37 PM Page 1654 Imaging M CHAPTERS Laser Scanning Microscopy Microscopy Components OCT Imaging Systems OCT Components Adaptive Optics M SECTIONS Tutorial Multiphoton Systems Multiphoton Accessories Multiphoton Essentials Kit Confocal Systems Laser Scanning Essentials Kit Laser Scanning Microscopy Tutorial (Page 1 of 6) Introduction Laser scanning microscopy (LSM) is an indispensible imaging technique in the biological sciences. In this tutorial, we will be discussing confocal fluorescence imaging, multiphoton excitation fluorescence imaging,...

01_LSM Tutorial_1653-1659.qxd.P:1653-1659 7/26/11 2:37 PM Page 1655 Imaging CHAPTERS Laser Scanning Microscopy Tutorial (Page 2 of 6) Figure 3 - Signal Generation in Laser Scanning Microscopy Linear Microscopy S1 Nonlinear Microscopy Absorptive Process (A, B): The absorption of one or more excitation photons (λEX) promotes the molecule from the ground state (S0) to the excited state (S1). Fluorescence (λEM) is emitted when the molecule returns to the ground state. S 1* λ EX λ EX λ EX λ EM λ EM λ EX S0 λ SHG,THG λ EX S0 One-Photon Excited Fluorescence Multiphoton Excited Fluorescence Harmonic...

01_LSM Tutorial_1653-1659.qxd.P:1653-1659 7/28/11 11:18 AM Page 1656 Imaging M CHAPTERS Laser Scanning Microscopy Microscopy Components OCT Imaging Systems OCT Components Adaptive Optics M SECTIONS Tutorial Multiphoton Systems Multiphoton Accessories Multiphoton Essentials Kit Confocal Systems Laser Scanning Essentials Kit Laser Scanning Microscopy Tutorial (Page 3 of 6) Point illumination, typically from a single Figure 4 - Optical Path in the Laser Scanning Microscope mode, optical fiber-coupled CW laser, is the critical feature that allows optical A. Confocal Optical Path sectioning. The light...

01_LSM Tutorial_1653-1659.qxd.P:1653-1659 7/28/11 11:21 AM Page 1657 Imaging CHAPTERS Laser Scanning Microscopy Tutorial (Page 4 of 6) wavelength and numerical aperture (NA) of the objective lens as seen in Eq. 1: SpotSize = Equation 1 1.22 . λex NA In actuality, the beam isn’t focused to a true point but rather to a bullseye-like shape called an Airy disk. The spot size is the distance between the first zero’s of the Airy disk (diameter across the middle of the first ring around the center of the bullseye) and is termed one Airy Unit (AU). This will become important again later when we discuss...

01_LSM Tutorial_1653-1659.qxd.P:1653-1659 7/29/11 8:32 AM Page 1658 Imaging M CHAPTERS Laser Scanning Microscopy Microscopy Components OCT Imaging Systems OCT Components Adaptive Optics M SECTIONS Tutorial Multiphoton Systems Multiphoton Accessories Multiphoton Essentials Kit Confocal Systems Laser Scanning Essentials Kit Laser Scanning Microscopy Tutorial (Page 5 of 6) Image Display Although we are not directly rendering an image, it is still important to consider the size of the image field, number of pixels in which we are displaying our image (capture resolution) on the screen, and the lateral...

Laser Scanning Microscopy Tutorial (Page 6 of 6) relaxation times. Furthermore, re-exciting a fluorophore before it has relaxed back down to the ground state can lead to irreversible bleaching of the fluorophore. Cells have their own intrinsic mechanisms for dealing with the cytotoxicity associated with fluorescence when it occurs slowly. One method to reduce photobleaching and the associated cytotoxicity is through fast scanning. While reducing the amount of time the laser spends on a single point in the image will proportionally decrease the amount of detected signal, it also reduces some of...

02_MPM Systems_1660-1666.qxd.P:1660-1666 7/26/11 3:21 PM Page 1660 Imaging M CHAPTERS Laser Scanning Microscopy Microscopy Components OCT Imaging Systems OCT Components Adaptive Optics M SECTIONS Tutorial Multiphoton Systems Multiphoton Accessories Multiphoton Essentials Kit Confocal Systems Confocal Accessories Multiphoton Microscopy Systems (Page 1 of 6) Multiphoton Microscopy offers several advantages over other laser scanning techniques, particularly the ability to image deeper into a sample. The modular design of Thorlabs’ Multiphoton Microscopy Systems is flexible enough to suit both researchers...

02_MPM Systems_1660-1666.qxd.P:1660-1666 7/28/11 11:25 AM Page 1661 Imaging CHAPTERS Multiphoton Microscopy Systems (Page 2 of 6) Four-Channel Multiphoton System Thorlabs’ Four-Channel Multiphoton System, an extension of our MPM200-2 Two-Channel Multiphoton System presented on the previous two pages, includes an additional transmitted light detection module (TLDM). This system provides a total of four detection channels, making it ideal for monitoring fluorescence resulting from multiphoton excitation as well as photons attributed to second and third harmonic generation.* summed with the backscattered...