The Live Cell Imaging facility

Nordic microscopy symposium – Save the date!

Please join the Nordic microscopy symposium on the 3^rd of October at the Live Cell Imaging core facility, Nikon Centre of Excellence at Karolinska Institutet. The symposium is organised by BergmanLabora, Ramcon Denmark and Inter Instrument AS. Please save the date.

The event will highlight Nordic research where light microscopy is a key tool.

Several Nikon fast super resolution systems will also be available: Nikon AX confocal with NSPARC and Nikon/CrEST V3 spinning disk with DeepSIM.

Before and after the symposium, the capability of these systems will be demonstrated during public sessions (on the 3^rd of October) and in private sessions (booking required) where you are welcome to bring your own samples.

Preliminary schedule:

Tuesday 3rd October

1000-1045 AX R NSPARC + CrEST V3 with DeepSIM public demonstration (in parallel)

1100-1145 AX R NSPARC + CrEST V3 with DeepSIM public demonstration (in parallel)

1200-1300 Lunch (requires pre-registration)

1300-1305 Welcome

1305-1345 *Keynote: TBC

1345-1410 *Nicoline Dorothea Daugaard (Dept. Biochemistry and Molecular Biology, SDU – Denmark):

Tracking T cells and their interactions with cancer cells in 3D using NIS-Elements General Analysis (GA3)

1410-1440 *LCI short talks:

Chiara Annunziata (Karolinska Institutet – Sweden)

Andrea Coschiera (Karolinska Institutet – Sweden)

Natalie Geyer (Karolinska Institutet – Sweden)

1440-1500 Coffee

1500-1525 *Super resolution system overview (Nikon): Title TBC

1525-1540 *Staffan Strömblad (Karolinska Institutet – Sweden): Multi-site assessment of reproducibility in high-content cell migration imaging data

1540-1615 *Pieta Mattila (University of Turku – Finland): Title TBC

Wednesday 4th October

0830-1400 AX R NSPARC + CrEST V3 with DeepSIM private demonstrations (bring your own sample)

Thursday 5th October

0830-1400 AX R NSPARC + CrEST V3 with DeepSIM private demonstrations (bring your own sample)

Venue (symposium): Erna Möller Lecture hall, Neo, Flemingsberg Campus, Karolinska Institutet

Venue (system demonstrations): Live Cell Imaging core facility, Neo, Flemingsberg Campus, Karolinska Institutet

Please share this event with colleagues who may be interested in attending.

Cool stuff one can do with the LCI Primo

At the LCI core facility, you can use a machine called Primo to micropattern/print proteins at the bottom of a microscopy dish and enable real time imaging of cell-protein interactions.

Now we learnt that we can even micropattern lipids and antibodies on microscopy dishes to image how cells interact with these molecules! This can be done with any pattern and even in multiwell plates! 🙂

Lipid micropatterning: Here is a cool paper showing how filopodia interact with sourrounding lipids and proteins. They image with Structured Illumination and TIRF, techniques that are also on offer at our facility. 🙂

Antibody micropatterning: A new way to analyse extracellular vesicles with multiplexed detection of proteins and RNAs at single EV resolution.

The power of Deep Neural networks

I am wowed at the images published recently in BioArxiv in the Zero-shot deconvolution networks paper. It will be interesting to see the peer-reviewed paper.

I would be interested in visualizing the amount of errors made by the network. A simple way to do it is to acquire a short time lapse (eg 10 images) of a fixed sample, run it through the network and see which structures are stably identified and which change from frame to frame. 🙂

Live broadcast of all public lectures the LCI microscopy course

Microscopy course: improve your imaging skills – from sample preparation to image analysis

Starting from next week, the Live Cell Imaging core facility at KI will run its yearly microscopy course.

All the course lectures will be broadcasted live on Zoom, free of charge and there is no need to register. On this page, you can find the course schedule (public activities are in blue) and the Zoom link to join. Scroll down to read the student testimonies! 😊

30 Jan – 17 Feb 2023

The aim for this course is to improve the microscopy skills of students and researchers who have already used a microscope to acquire digital images of fluorescent samples but feel that more knowledge could help them.

The course covers the following topics:

Optics, image formation
Fluorescence, fluorophores
Bleedthrough
Anatomy of a microscope
Objectives and refraction index
Cameras and detectors
Noise and background, Bit depth and saturation
Multichannel imaging and spectral unmixing
Resolution and contrast
Sample preparation, Immunostaining
Nyquist sampling
Confocal and wide field settings
Speed, High throughput/content
Volume imaging, deconvolution
Clearing and expansion
Live cell imaging
Fourier
AI, Super Resolution microscopy
Colocalization
Data handling, OMERO.figure

Hope you enjoy the Live Cell Imaging core facility microscopy course! 😃

Registration to the LCI microscopy course 2023 closes on Monday!

It is time to enroll to the yearly light microscopy course run by the Live Cell imaging core facility (30 Jan – 17 Feb 2023): ‘Microscopy: improve your imaging skills – from sample preparation to image analysis’ (6 credits).

Check the course schedule to see the course content and testimonies from alumni
If interested, you can enrol as a student. Please read carefully the eligibility criteria and note that the last registration date is the 15^th of November.

Additionally, all the lectures (in blue on the schedule) will be publicly broadcasted live on Zoom and accessible to anyone without registration. Even if you do not want to enrol as a student, you can have a look at the and listen to any lecture that triggers your interest. There is no need to register. The schedule and zoom link are on the course page.

The purpose of this course is to enable PhD students and researchers who have already and recently used a microscope to acquire images of fluorescent samples, to improve their microscopy 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 their OWN microscope, to enable them to:

1) prepare their sample and formulate their scientific question in a way that is suitable for data extraction from fluorescence images

2) properly use the hardware available in their lab/facility and 3) fully understand each parameter they need to set in the software in their lab/facility.

The aim is to provide the course students with the tools to acquire on ANY wide field, confocal or light sheet microscope, images of their samples that reliably answer their scientific question.

Please help us spread the word. 😃

Light-seq: Multiplexed, non-destructive spatial transcriptomics of tissues sections using light

Light-Seq is a new pretty cool technique for highly multiplexed sequencing of RNA in tissue sections using light. This technique is highly sensitive, highly spatially resolved and because it does not destroy the tissue, it can be combined with protein labelling (genetic or by immunolabelling).

On one of our single-point confocal/spinning disk/widefield system at the LCI facility, we have a device called Primo (DMD + UV laser) which can be used to run this technique! 🙂

Let us know if you would like to set up LightSeq at the LCI core facility!

Live demo of the Nikon Mizar TILT light sheet

On the 22nd of September at 10, you get a chance to see and try the latest light sheet on the market: the Nikon Mizar TILT.

- Easy to use stage top light sheet with only 1 objective
- Imaging of cell monolayer, cells in gels, organoids in gels, small organisms…
- Samples in glass bottom multiwell chamber slides
- Stage top incubator for live sample imaging
- Advantage of using the TILT light sheet
  - low phototoxicity and bleaching even with high resolution objective
  - excellent temporal and spatial resolution for live samples
- See here for more info.

This demo is only for a limited time (10 days) and in person only. Please let us know if you want to join the demo and if you want to try your own sample.

Live demo of the Crest spinning disk with DeepSIM

9^th of September, 10-12am: Live/zoom demo of the latest microscope acquired by the LCI facility: Crest spinning disk with DeepSIM (new Orion)

Ti2 microscope
Crest spinning disk (identical to the one on Tweety but with slightly different wavelengths)
DeepSIM: SIM is a fast super-resolution technique which is gentle for the sample (less bleaching, less photodamage for live samples) and offers double the resolution of a confocal microscope (down to 120nm resolution), even thick in samples. It is flexible and can be used with any objectives, even low magnification air objectives.
No specific sample preparation requirements
More information here

For the best experience, join the demo in person at the LCI facility (please apply by replying to this email) but if you cannot make it, you can also listen on Zoom.

The system has been purchased by the LCI so it is here to stay! 🙂

Please let us know if you want to join the demo in person and mention if you would like to try imaging your own sample with DeepSIM.

mCherry-XL: brigther, more stable and with better spectra!

mCherry is a very popular red fluorescent protein. However it has several disadvantages:

It is shifted towards far red (ex peak 585 nm) so it often is not imaged optimally with the illumination sources and filters commonly available.
It bleaches fast

mCherry has now been evolved into mCherry-XL with several improvements:

This variant is shifted back towards green (ex peak 560 nm) therefore being very well excited with popular 561nm lasers.
It is 3 times brighter than mCherry
There is also a clear improvement in the lifetime for FLIM
Together the 2 points above means that less excitation power is required so it should help with the bleaching problem

Here is the paper. Therefore you should consider mCherry-XL for your future tagging with red fluorescence proteins.