Tag: Sample preparation

The LCI microscopy course 2022 is now open for registrations!

It is now time to apply to the intensive LCI microscopy course Jan/Feb 2022: Microscopy: improve your imaging skills – from sample preparation to image analysis

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 their knowledge is limited.

All the lectures at the LCI microscopy course will also broadcasted live online, free of charge and there is no need to register.

All details about the course including course schedule, how to apply, and how to follow the lectures are found here.

Scroll down to read the kind testimonies of our dear students! 😊

Hope you enjoy the LCI facility microscopy course 2022!

Brighter secondary antibodies

Fluorophores are in constant development.

There is now a new generation of Alexa Fluor fluorophores: the Alexa Fluor Plus. They can be purchased already coupled to secondary antibodies on the ThermoFisher website.

Brilliant violet antibodies are also very bright and despite their name, they come in all colours. The can be purchased from Biolegend which produces the fluorophore.

Another excellent source of bright fluorophores is the Janelia Fluor dyes. They can be purchased coupled to antibodies from the NovusBio website.

The first microscopy MOOC?

This MOOC for undergrad students by the University of Montpellier is about labeling and imaging vertebrate embryos.

All the videos are also available on their YouTube channel.

As far as I know, this is the first pure microscopy MOOC (Massive Open Online Course), dealing with sample preparation, imaging and data handling!

If you know of another microscopy MOOC (I know of some image analysis MOOC but not microscopy), please leave me a comment or sent me an email! 🙂

The LCI microscopy course 2021 starts next week! :)

As usual the lectures at the LCI microscopy course will broadcasted live online, free of charge and there is no need to register.

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

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.

Applications are closed but all lectures will be broadcasted live and open to anyone without registration.

The course covers the following topics:

  • Optics, image formation, fluorescence, fluorophores, microscope and microscopy types
  • Objectives and refraction index, Cameras and detectors
  • Noise and background, Cameras and detectors, Bit depth and saturation, Multicolour imaging
  • Resolution and contrast, Sample preparation, Immunostaining
  • Nyquist sampling, Confocal and wide field settings, Scaling up and speeding up, High throughput/content
  • Volume imaging, deconvolution, multiphoton, Clearing and expansion
  • Live cell imaging, Fourier, AI, Super Resolution microscopy
  • Data handling, OMERO.figure, Requirements for image analysis, Colocalization
  • Image processing and analysis

Check the course schedule and details of how to join the Zoom webinars. Scroll down to read the kind testimonies of our dear students! 😊

Here is the course syllabus.

Hope you enjoy the LCI facility microscopy course 2021!

Where to find microscopy jobs, forums and wikis

Where to ask microscopy-related questions?

– Image.sc and microlist
Post your microscopy, sample preparation and image analysis questions

– Confocal server
Old email-based forum that is followed by tons of people across the globe as well as most microscopy companies. Post your microscopy and sample preparation questions.

Freeware for image analysis

– Image J/Fiji
Great freeware to open images acquired on any microscope, resize, crop, change contrast, create movies, annotate, analyze and much more.

– Cell Profiler
Freeware for automated image analysis including machine learning.

– KNIME
Open source to build a custom made image analysis pipeline

– Icy
Great freeware to open images acquired on any microscope, resize, crop, change contrast, create movies, annotate, analyze and much more.

Networks, jobs and career

Light microscopy jobs, meetings, courses, networking… in Europe. Remember to mail them that you want to show up on their map of European microscopy!

Loads of information from which companies/institute works with research to how to find an apartment when moving to another lab, find work, know your rights, express your opinion about researchers’ work conditions…

Other places to look for work:

Microscopy wikis

– Kurt Thorn’s excellent microscopy blog
I highly recommend this blog even if it is not updated anymore. Monthly digest of major microscopy related articles. Loads of tips.

– Microscopy Education by Nikon

– Leica Science Lab

– Microscopy Education by Olympus

– Microscopy Education by ZEISS

– Molecular Expressions

– SVI deconvolution

– Microscopy info

– Nature Milestones Light Microscopy

– iBiology Microscopy Course

Great webinars about sample preparation at Scilife

Hans Blom from the Advanced Light Microscopy facility at Scilife organises next week (10th and 1tth of June) 3 sessions of great seminars:

I especially recommend the CUBIC seminar to anyone who wants to image samples thicker than 100 um. This will teach you how to treat your sample to make it fully transparent so you can image through many cm of tissue!

🙂

Super bright green fluorescent protein

Fluorophores are evolving fast! Here is a paper about a bunch of new fluorophores isolated from the jelly fish Aequoria. This includes the brightest fluorophore ever isolated: a new green fluorescent protein called AausFP1 that is almost 5 times brigther than GFP! Respect! 🙂

One-step non-toxic clearing protocol

Do you know that clearing is not just about light sheet microscopy? Even if you have done your job well and your sample is directly on the coverslip (not on the slide), as soon as your sample is thicker than 10 um (1 cell diameter), you will see the effect of the refraction index mismatch.

What is that? Your sample and the mounting medium around it have a certain refraction index (or likely several). The objective you are using is designed for a certain refraction index (e.g. air, water or oil). If these refraction indices do not match what happens? as soon as you image a tiny bit away from the coverslip, the sample will look elongated, the intensity and contrast will drop very fast.

Sounds familiar? If yes, changing your mounting medium to match the objective will solve the problem. It works for light sheet but it also works for wide field or confocal imaging! Just change your mounting medium and you will see an enormous difference!

Here is an article describing a one-step clearing protocol. This basically is about using a different mounting medium. Easy, cheap and non-toxic! Give it a try!

Have a look at this post for more info.

The antibody validation nightmare

Ever wondered if that antibody you used throughout your whole PhD was actually also binding to something else than its supposed target protein?

Antibody validation in tissue staining is a very difficult task!

Here is a great step-by-step validation protocol published by EuroMabNet, a network of scientists who try to improve antibody validation.

And this paper gives a useful flow chart for antibody validation.

And here is the 5 pillars of antibody validation paper which explains what can be done to validate antibodies.

 

Know your RRid!

Imagine starting a study about some cool protein.

You find some useful articles on Google Scholar. In one paper, an antibody is mentioned. The name of the company that sold it to the authors is mentioned in the Material and Method but unfortunately that company has closed down or has been swallowed by one of the Pharma giants so you cannot order. Then you realize that the company was not producing any antibodies anyway, they were buying it from another company so there is no way to trace and buy the same antibody. Nightmare…

Then imagine that instead, the paper mentions the RRid number for that antibody. You do not know about what that is but you check and find this paper that explains it all.

Now suddenly, not only you can find on the Scicrunch website which company produces this antibody and which resells it so you can buy it, but you can also search pubmed for the RRid and find all the articles that mention it, opening your eyes to lots of results about your protein that have been published specifically with using that antibody. Now you can also check if the antibody gives consistent results!

And imagine being to do this for your favorite mouse model as well. See all publications that have mentioned your mouse RRId!

But it relies on you writing the RRid of your antibody or mouse in your next publication so think about it! 😀

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