My fluorescence microscope project and request for possible stains/filters

[flyingpingu]

Hey everyone.

I have been doing a project for a fluorescence microscope based on the UC2 platform. I started it originally in 2022 already but took breaks from it, which is why it’s probably outdated and I actually couldn’t find it in the git anymore. The basic setup is however the same as here: GitHub - openUC2/UC2_Fluorescence_microscope: This is the repository for the laser-based fluorescence microscope with infinity optics. It is capable to acquire Time-Lapse videos from living cells inside an incubator.. I am using the old xyz-stage with a twist which is that the xy-part is powered by software based on this git GitHub - koendv/xy-stage: mechanical stage (I recommend taking a look). I also made a basic python interface to control the microscope and picam. Another thing is that I don’t have a beamsplitter or dichroic mirror but simply shine the laser directly on the sample.

Since my project is for university, I want to show some examples of fluorescence with it. As shown in the picture, I had some first success with an onion stained with Eosyn. Here are my questions:

• What is another sample I could use, or rather what stain would work well? I have gotten a bit frustrated trying to find something that I know I can get an affordable filter or dichroic mirror for. It just needs to be something to show that it works, nothing fancy.
• Do you think not having a beam splitter/beam expander setup but simply shining the laser directly on the sample and using an emission filter will lead to decent results, or would you recommend getting those elements?

Thanks a lot,
Alex

[benedictdied]

Hey @flyingpingu, this is amazing! Thanks a lot for pushing the project further and showing the results. Very nice! Samples are always a bit of an issue if you don’t have any already. What we use for light-sheet microscopy as a reference is paper with yellow textmarker. You find anything that is any brighter. If you observe the fibers at an edge that you scraped away, you have a nice structure already. As for the filter-set you can use those absorption filters from Theaters (i.e. Lee). You can find them at musicstore at DinA4 sizes. As for the filters, there are not real alternatives yet. Fluorescence micorscopy is just not a mass product unfortunately… So the arrangement in “darkfield” that blocks the zeroth order looks good. Almost like a light-sheet

Any thoughts @the_alquemist et al?

[benedictdied]

as For the software, we have moved to ESP32-drive firmwares. But I think It’s a whole different story
Some information here https://youseetoo.github.io/

[the_alquemist]

We use Fluorescin (FITC) in the lab. In Chile, the cheapest option is sold for $29 in a 25G format and we use a hundredth of that (or less) for basic testing (diluted in a cuvette), and it looks awesome!. I believe it also works well with plant based samples. Some food dyes also have fluorescence properties, could be an option too.

I think what you have now is amazing! Any further fine tuning will depend on the requirements or objectives stated in your project. Additional gadgets will definitely help with the image, but again, it depends.