Advanced Microscopic Techniques



Time and place

  • Thu 10:00-12:00, Room Zoom-Meeting ICS

Prerequisites / Organizational information

Registration by writing to the lecturer


In this course we will cover the following topics: 1. Confocal microscopy: Confocal microscopy is an imaging technique which provides improved resolution and contrast compared to full field imaging by using a pin hole which helps reducing the out of focus light. Confocal microscopes are backbone for most of biological labs and are used frequently to study cellular mechanics. 2. Optical coherence tomography imaging (OCT): OCT is an imaging technique which can provide axial resolution better than 1 micron using broadband low coherence light source. This has allowed to perform optical biopsies for several biological samples in vivo. 3. Raman microscopy : Raman microscopy is a technique within vibrational spectroscopy, which is based on the inelastic scattering of light. It provides information on the chemical composition of the sample based on its vibrational spectra. Since the development of the first commercial Raman spectrometer in 1953, advances in lasers and detectors and the discovery of new phenomena have expanded the use of this technique in several research fields 4. Stochastic optical reconstruction microscopy (STORM): STORM is one of the most ubiquitously employed super-resolution imaging techniques. It utilizes sequential activation and time-resolved localization of photoswitchable fluorophores to create high resolution images. During imaging, only an optically resolvable subset of fluorophores is activated to a fluorescent state at any given moment, such that the position of each individual fluorophore can be determined with high precision. 5. Stimulated emission depletion (STED): STED creates super-resolution images by the selective deactivation of fluorophores, minimising the area of illumination at the focal point, and thus enhancing the achievable resolution for a given system. 6. Multi-photon excitation (MPE): MPE microscopy is an imaging technique which operates in non linear regime that combines point scanning methods with multiphoton fluorescence to create high-resolution, three-dimensional images of biological samples. Several forms of MPE such as 2 photon, 3 photon microscopy etc, are available. MPE is particularly useful in biology because it can be used to probe delicate living cells and tissues without damaging the sample. 7. Phase contrast microscopy (PCM): Several cells offer very low contrast when visualized with standard microscope. PCM provides improved contrast and is a label-free imaging technique allowing visualization of transparent cells. The quantitative phase contrast image provides information about the optical path length change introduced by the sample because of its refractive index and thickness. 8. Polarization sensitive optical coherence tomography (ps-OCT): ps-OCT is gaining attention because of its ability to diagnose certain pathological conditions at an early stage. Several pathological conditions such as cancer can be detected at an early stage by measuring birefringent properties of the tissue. ps-OCT uses low coherence polarized light to probe the birerefregence of the tissue. 9. Brillouin Microscopy: Brillouin microscopy is an emerging optical technique that enables non- contact measurement of viscoelastic properties of a material with diffraction-limited resolution in 3D. It exploits Brillouin scattering, the interaction between light and acoustic waves intrinsically present in any material due to thermal vibration. 10. Structured illumination microscopy (SIM): Structured illumination microscopy (SIM) enhances spatial resolution by collecting information from frequency space outside the observable region. This process is done in Fourier space. The reverse Fourier transform then returns the reconstructed image to a super-resolution image.