• The basic layout of the light microscope as consisting of a microscope objective and an eye-piece, and their properties such as aberrations, magnification, numerical aperture, and field of view.
• Different methods for obaining contrast in a microscope, such as absorption, fluorescence, and phase-contrast methods.
• Different image-quality measures such as resolution, contrast, signal-to-noise ratio (SNR), modulation transfer function (MTF), sampling density, field of view, and depth of field.
• Layout of the illumination systems, particularly Köhler illumination for transmission and epi-fluorescence microscopy, and how the illumination system can be used to optimize image quality. 
• How properties of the microscope and illumination system affect image quality such as resolution and contrast, mainly via Fourier methods including point-spread functions (PSF) and optical transfer functions (OTF, MTF).
• How the choice of detector affects image quality measures such as signal-to-noise (SNR) and sampling. How to sample to avoid loss of information and artefacts. Some microscope photometry.
• The basic layout for confocal microscopy and hence three-dimensional imaging, including resolution and sampling in different dimensions.
• The basics of nanoscopy and imaging beyond the classical resolution limit. 

After completing the course the student should be able to:

• adjust the microscope and the illumination system to obtain optimal performance in transmission and fluorescence microscopy.
• select suitable contrast methods and microscope objectives for standard microscopic samples.
• perform and report quantitative microscopic measurements, including image computer processing and 3D visualization.
• extract and comprehensively present relevant information from a scientific publication.
• assess how different image quality measures are affected by physical limits connected to choices of microscopes and imaging parameters, and use this knowledge to choose suitable settings in new imaging experiments.

Lectures will be given in a hybrid format, so that students may attend on-campus or via zoom as they choose.

Each student will perform four 4-hour laboratory exercises. At a specific date and time, maybe two weeks into the course, the sign-up for labs will open, so that students may sign up for the specific times that suit them best.

Each student will study a scientific paper on a subject relevant to the course, and present it at a seminar. A number of suggestions for papers will be offered but you can also choose your own, if approved by the teacher.

Lecture 1. Basics of light microscopy: imaging ray-path and illumination ray-path, aberrations,   objective types, magnification, numerical aperture

Lecture 2. Contrast techniques: absorption, fluorescence, phase contrast, DIC, dark-field

Lecture 3. Fundamentals of radiometry and photometry, microscope photometry, detectors, noise

Lecture 4. Resolution, mathematical representation of the imaging process

Lecture 5. The Fourier transform and its interpretation, the optical transfer function OTF

Lecture 6. Continue OTF: for diffraction-limited optics, for 2D, for detectors, for imaging chain

Lecture 7. Sampling: sampling and aliasing, reconstruction calculations, multidimensional sampling

Lecture 8. Coherent imaging in microscopy, role of condenser numerical aperture

Lecture 9. Introduction to confocal microscopy

Lecture 10. Imaging properties of confocal microscopy

Lecture 11. Confocal microscopy: Limitations and errors, multi-channel detection

Lecture 12. Super resolution fluorescence microscopy: stimulated emission depletion microscopy (STED)

Lecture 13. Super resolution fluorescence microscopy: single-molecules based microscopy

Lecture 14. Problem-solving session

Lab 1. Build your own microscope with Koehler illumination (4 hours at AlbaNova)

Lab 2. Practical use of research microscopes in different imaging modes (4 hours at AlbaNova)

Lab 3A. Confocal microscopy (2 hours at AlbaNova)

Lab 3B. Super resolution fluorescence microscopy (2 hours at SCILife)

Lab 4. Analysis and visualization of results from lab 3 (4 hours via zoom)

Schema HT-2021-196

Two compendia by Kjell Carlsson available via the course canvas page.

Students at KTH with a permanent disability can get support during studies from Funka:

Funka - compensatory support for students with disabilities

A, B, C, D, E, FX, F

• LAB1 - Laboratory Experiments, 2.0 credits, Grading scale: P, F
• SEM1 - Seminar, 1.5 credits, Grading scale: P, F
• TEN1 - Examination, 4.0 credits, Grading scale: A, B, C, D, E, FX, F

Based on recommendation from KTH’s coordinator for disabilities, the examiner will decide how to adapt an examination for students with documented disability.

The examiner may apply another examination format when re-examining individual students.

Laboratory Experiments ( LAB1 )

Complete all four labs (active participation) and submit a report of sufficient quality to pass pefore deadline. If needed you will get a chance to improve your report according to comments.

Seminar ( SEM1 )

There will be several seminars and you should attend one, where you present a paper and participate in the discussion of the other papers. The specific times, and which seminar to attend, will be determined after discussion with the students.

Examination ( TEN1 )

The examination will happen in two steps:

1. Oral exam via zoom, 30 min per student. If you pass you get the grade E on the course.

2. If you pass the oral exam, you will be offered the chance of a written home exam to improve your grade to D, C, B, or A. The home exam will be three hours long and distributed via Canvas.

As we cannot do all orals and written exam in one day, the examination wil be extended over several days and hence not necessarily happen during the scheduled exam time. The exact time distribution will be determined after discussions with the students. Students will sign ut for time slots for the orals.

Written examination (TEN1; 4 hp, grading scale A/B/C/D/E/Fx/F), completed laboratory course (LAB1; 2 hp, grading scale P/F) and seminar presentation (SEM1; 1.5 hp, grading scale P/F).

Learning outcomes 1-3

(P) complete all labs and report and evaluate the results in a comprehensive way

Learning outcome 4 [SK2501 only]

(P) present a research paper and actively participate in the seminar

Learning outcome 5

(Fx) almost fulfil the requirements for (E)

(E) asses how some of the image quality measures are affected by microscopy parameters, and choose suitable microscopy systems and parameters in some cases.

(D) fulfil the requirements for (E) and partly those for (C)

(C) asses how most of the image quality measures are affected by microscopy parameters, and choose suitable microscopy systems and parameters in most cases

(B) fulfil the requirements for (C) and partly those for (A)

(A) asses how the image quality measures are affected by microscopy parameters, and choose suitable microscopy systems and parameters

• All members of a group are responsible for the group's work.
• In any assessment, every student shall honestly disclose any help received and sources used.
• In an oral assessment, every student shall be able to present and answer questions about the entire assignment and solution.