This is an ad to a job as facility manager in Braga, Portugal. 🙂
Month: October 2019
Seminar on how to directly label your primary antibodies: Skip the secondary, part 3!
The LCI will host a seminar about GlyCLICK, a new way to directly label primary antibodies and stop using secondaries: 6th of November at 13:00 in the Lipid seminar room in Neo, KI Flemingsberg. The seminar will be broadcasted live. Here is how to find us and here how to follow the seminar online.
Everyone agrees that it would be great to be able to label our samples without using secondary antibodies.
- less animals killed
- shorter and cheaper protocols
- no problem with isotype cross-reaction
- no problem with secondary species when using many antibodies at once
There are many kits to label primary antibodies directly with fluorophores. The main disadvantage compared to primary/secondary stainings is that direct labels are often weaker because the final primary/fluorophore ratio is too low. Using an amplification method like TSA (Tyramide Signal Amplification) leads to a loss of resolution.
Over the years users at the LCI have tried this kit, this kit and this one. They gave mixed results depending on the antibody but we keep looking! Come to the LCI seminar about a new direct labelling technique that uses Click chemistry.
Apply now to the LCI microscopy course 2020 :)
It is now time to register to the LCI intensive microscopy course (Jan/Feb 2020). Check out the course schedule.
Loads of fun workshops, informative lectures, intense discussions and our popular Student Imaging Challenge workshop where students get direct feedback on how to improve their own sample preparation/experimental design.
We always run two courses in parallel:
- the full course (#2870, 6 points, apply here) where students attend all activities
- the theory only course (#2871, 4.5 points, apply here) for students who only attend the lectures
As usual, all lectures are public and broadcasted live so you are welcome to just show up (how to find us) or watch remotely (how to connect) without registration. Check the program as it may be updated in case of (unlikely) last minute changes.
We welcome your feedback about the quality of the webinar and the content of the lectures.
How to precisely measure the volume of a cell?
Measuring the volume of a cell is often done by labelling the cell membrane or its cytoplasm. Analysing large flat cells this way is easy but it is much harder for tiny cells like blood cells, yeast or bacteria.
Another way to measure volumes is to use a negative stain, i.e. where the medium is made fluorescent with a dye that does not go into the cell. The cell appears as a black hole in fluorescent images and unlike lipid-based membrane labelling, borders are even and easy to segment.
While many dyes can be used for live cells, one must choose large dyes when negatively imaging cells that have been fixed and permeabilized.
This paper and this one use high molecular weight (2000 KDa) Dextran to achieve these results and measure the size of bacteria.
This recent paper optimizes the technique.
Free genetic and image analysis tools
Here are a few image analysis tools (constructs for FRET, software…) that might be useful to you.
- Construct for fluorescence biosensors and optogenetic tools
- Free Image analysis software
http://www.hahnlab.com/tools/index.html
- Free Image analysis software
https://www.utsouthwestern.edu/labs/danuser/software/
- Free Image analysis software
Party time at the LCI! :D
On the 10th of October, the Live Cell Imaging facility will have an open house and a little party to celebrate loads of great stuff:
- The LCI turned 10 years old this year! 😊
- We got a wonderful light sheet system earlier this year. It is high time to give it a name and splash it with a bit of champagne!
- Gisele Miranda from Scilife/BII has joined the LCI team to help our users with image analysis
- We got a great server/analysis capacity set up by the KI IT department
Wow! What a year! 😃
Please come and celebrate with us! If you do not know what our microscopy facility has to offer, it is time to be curious and pay us a visit.
- Open house: drop in between 9:00-11:00
- Baptising of the light sheet system and celebration of Gisele and our shiny new server: at 11:00
All this will happen on the 10th of October (next Thursday) at the LCI facility, on the 7th floor of Neo at KI Flemingsberg. Here you can see how to find us.
Please help us spread the news! 😊
Deep red fluorescent proteins
The microscopy field is moving away from blue dyes. This is because red light, used to excited far red and deep red fluorophores, is less damaging to live cells than near UV light which is used to excite blue fluorophores.
On top of that, red light penetrates deeper into thick samples.
So as the trend in microscopy is to move to thicker samples and use more live samples, far red and deep red fluorophores are becoming more attractive.
Here is an article describing 3 new fluorescent protein in the far red to deep red range. One can excite them with 640 nm or a 685 nm lasers or LEDs.