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Title Microscopy: Improve Your Imaging Skills - From Sample Preparation to Image Analysis
Course number 2870
Programme Utveckling och regeneration (DevReg)
Language English
Credits 6.0
Date 2022-01-24 -- 2022-02-11
Responsible KI department Institutionen för biovetenskaper och näringslära
Specific entry requirements The students must have an on-going microscopy project for which they must know how to prepare their own fluorescent sample for microscopy. They must have been trained on and have used (minimum 3 months prior to the course start) a microscope able to acquire images of fluorescent samples.
Purpose of the course The aim for this course is to improve the microscopy skills of students and researchers who have already and recently used a microscope to acquire digital images of fluorescent samples and want to improve their skills.
The course is NOT aimed at training people to use the LCI facility microscopes. The focus is instead on providing the students with enough theoretical and practical knowledge about their OWN sample and microscope, to enable them to 1) properly use the hardware available in their lab/facility and 2) fully understand each parameter they need to set in the software. The aim is to provide them with the tools to acquire on ANY wide field, confocal or light sheet microscope, images that exactly match their samples and reliably answer their scientific questions.
Intended learning outcomes At the end of the course, the participants will be able to:
1- Describe the differences (theory and hardware) between wide field, confocal and light sheet microscopes as well as the different types of confocal microscopes and choose which system is most suited to their experiments
2- Pick the best combination of fluorophores for their experiment by matching the fluorophore spectra with the light source and filters on their microscope, identify and eliminate bleed-through and cross-excitation problems
3- Explain objective specifications and limitations and choose the appropriate objective for their own experiments
4- Describe how to fix, mount and handle their sample in a way that is optimal for imaging
5- Find the area of interest in their sample and without bleaching it
6- Adjust the condenser for proper bright field imaging
7- Explain how to set the following parameters on a wide field, a confocal or a light sheet system to best match the requirements of their sample and reliably answer their scientific question: resolution, pixel size, averaging, scan speed, illumination power, detector gain and offset, camera readout rate, exposure time and binning
8- Explain how to easily assemble a figure for publications
Contents of the course Aside from the points described in the Learning Outcomes, the participants will learn the following:
- Many practical tricks about fixation, mounting and handling of their sample in a way that is optimal for imaging
- How to deal with the challenges of imaging fluorescent volumes
- The advantages in using the automation of a microscope system to collect multidimensional data
- What hardware or software autofocus, spectral detector, resonance scanner, two-photon or super resolution microscopy are used for
- Many personalized tips on how to improve the preparation and imaging of their own sample on their own microscope (through the workshop where we will image their own sample)
- The ethics of handling scientific images for publication
- The imaging requirements for automated image analysis
- Where to get help to create an image analysis pipeline on freeware (ImageJ/FIJI, Cell Profiler) designed for their own images and scientific question
Teaching and learning activities Lectures, videos, workshops, peer review, image troubleshooting in groups, project presentations, quizzes.
Compulsory elements Attendance to all sessions is compulsory. Any absence must be reported to the course leader in advance by e-mail. Absence from any part of the course (lectures, laboratory sessions, discussion sessions and exam) is generally not accepted but could in exceptional cases be compensated by a written additional assignment to ensure the learning outcomes of the day have been reached. If it is not possible to compensate, the student will be given a chance to complete the course by attending the missing sessions the following year.
Examination The final mark (pass or fail) will depend on the results of
1) The weekly assignments;
2) The skills shown at each workshop;
3) The written examination at the end of the course.
The student must show that all intended learning outcomes of the course have been reached.
Literature and other teaching material Reference literature: Handbook of Biological confocal microscopy, James Pawley Springer Editions 2006 (PDF file available at the KI library)
Number of students 12 - 20
Selection of students The selection will be based on the applicants' potential to follow the course and fulfill the assignments (see More Detailed Information) and the usefulness of the course to their current research project.
More information The students should a) have an active microscopy project where they acquire digital images of fluorescently-labelled samples; b) be able to bring their sample to the course; c) have access to their microscope during the course (some assignments require submitting new images). The application must contain the following 4 parts: 1) the confirmation that they fulfill the a, b and c conditions listed above; 2) a brief description of the scientific aim of their imaging project; 3) a brief description of their past microscopy experience, when they started their current microscopy project (should be at least 3 months) and which microscope(s) they currently use; 4) fluorescent images of their sample acquired by them and uploaded in the original microscopy format (please write to Anna Wallén (Anna.Wallen@ki.se) to ask for a link to upload your original images). Researchers who have not yet used microscopy or do not have an active microscopy project are advised to get trained at their local imaging facility, acquire images for at least 3 months then apply to the LCI course next year.
Additional course leader In case of problem with the application process, please contact Anna Wallén (anna.wallen@ki.se).
Latest course evaluation Course evaluation report
Course responsible Sylvie Le Guyader
Institutionen för biovetenskaper och näringslära

Sylvie.Le.Guyader@ki.se
Contact person -