A blog about health economics, public health issues, free software, and probably everything else too …
Tag: gel
I’ve just released the version 0.2 of Picklist Editor. Now you have a table of all the proteins on the right of the gel. If you double-click on a cell, you can edit it (note this is not a recommended behaviour). After revalidating the table, your new spot will be included in the gel (and saved to your picklist if you like it). For me, this version is stable and fully functional 🙂
This morning, I released Picklist Editor 0.1 with a text introduction … Hmmm … on my photos on Flickr you can see the hardware side of the picking process … (click on pictures to see details).
On the photo on the left, you can see a gel on a low-fluorescent glass plate. This plate is in part in a tray that firmly holds it when the robot is doing its job. The holes everywhere result from the picking process but there are proteins everywhere and you can’t see them in visible light since they are labelled with fluorescent Cy dyes. You can see two white round stickers on each side of the gel: these are the picking references.
Here, the picker head is in the process of taking a part of the gel with some proteins inside. Exact positions were computed according to the fluorescent images, revealing the proteins. As you can see again: the gel is perfectly transparent for our non-bionic eyes.
Finally, you can see the spot picking robot in action. The picking head is moving following two axis thanks to the horizontal bar at the back and the perpendicular arm holding the picking head and camera. On the left, you have a pumping station: in addition to some jazz when the picking head is on the gel, the station is aspiring water through the head in order to help getting a plug out of the gel. After that the arm moves to the right of the photo where you have two 96-wells plates to collect samples. When the head is above a well, the pumping station is “blowing” water into the head in order to eject the plug into the well. Everything is under control of a computer and software that is on the right, outside of the camera angle.
This is the GE Healthcare “Typhoon 9400” scanner used to scan fluorescent gels. It’s a huge beast but it doesn’t make a lot of noise (well, I don’t want to stay the whole day next to it!). And this unit only has the red and green lasers inside. There is a second (smaller) unit below with only a blue laser source in it! You can see two gels ready to be scanned (the upper door has to be closed before!).
Next is the analysis “workstation” where images of the gels are analyzed (after scanning and before spot picking). The software (DeCyder) helps to create pick lists.
Other photos from my labs can be seen in my laboratory photostream.
When you work with 2D gel electrophoresis in proteomics, you end dealing with "pick lists". For this purpose, I wrote Picklist Editor, a tool to help visualize and modify this pick list.
As usual, software and source code are available here. Feel free to use it and report any bug or your wish list 🙂
(and if you didn’t understand everything above because you are not in the proteomics field, just go to the page too because I also wrote a small introduction)
I should change this blog title to “Version 0.1” since I recently released version 0.1 of many software tools 😉
Last week, our proteomic group did a 2D of a purified protein but, unfortunately, it seemed we had a contamination. So, this week-end, we performed blank tests in order to see if the contamination came from our experiment (or from the purification). We tested our fluorescent markers and different 1D strip holders. At a normal gain, the image looks fine: contamination doesn’t come from the 2D
But when I increase the gain, the image (and thus our gel) is really contaminated! 😦 We really have to look where did the contamination happen! Look at this:
In France the person fooled is known as poisson d’avril. This has been explained as arising from the fact that in April the sun quits the zodiacal sign of the fish. The French traditionally celebrated this holiday by placing dead fish on the backs of friends. Today the fish has been replaced with paper cut-out.
(Explanation from Wikipedia)
The whole day, I was busy working with DeCyder, a software to analyse spots of proteins in 2D gels. I hope I won’t dream of small red/yellow/green dots tonight:
The software use a 3D representation of the spot. If we change this view too fast (either by moving it or by quickly browsing through spots), the graphic diplay freezes and we fall back to an ugly VGA mode (screenshot below). Since the morning, the computer crashed 7 times! I reduced the screen resolution to 1024x768pixels in 24bits colours and it seems to work correctly now. 🙂
This week is very stressful because I am doing a 2200+ euros 2D-DIGE experiment (*) on samples from a rat organ we never studied before and from which I cannot obtain any more new samples.
We found a new pattern of proteins dispersion (compared to our previous experiments on other organs) and, more importantly, we found a clear difference in protein expression in at least 2 spots. In the image below, all the whitish spots mean proteins in these spots are found in equal amounts in the 3 conditions. But spots in red or green mean proteins expressed at different levels (even on/off) between conditions!
Now, we’ll have to perform a more mathematical/quantitative exploration of these expression patterns and hopefully identify these interesting proteins by mass spectrometry.
(*) 5nmol of Cy dyes (proteins labelling molecules) cost 2215 euros and weight something like 2.75*10-6g! I used nearly all that amount for my experiment. And I don’t take into account the 3-lasers scanner and common lab reagents
Jean-Etienne's blog 