Couple of questions regarding fluorescence set up (Delta Stage)

Request Help
#1

Hi,
Thank you for your great initiative that makes imaging feasible for everyone.
I am going to make a fluorescence version of the microscope and have a couple of questions. Hope someone can help me in this regard.
I am mainly focusing on GFP imaging in plants at the beginning.

  1. Can you suggest a good place and cat. #s to buy the excitation, emission filter, dichroic beam splitter in United States? I talked with Thorelab technical support, they couldn’t give me a catalog number that specifically fits with the Openflexure modules.
  2. How I can come up with a way to take images removing the chlorophyll autofluorescence that interfere with GFP in Openflexure setup. Here is the link about the issue Please note that I would like to have three images for my studies (autofluorescence, GFP and Brightfield)
  3. What is the recommended blue LED for the GFP imaging?

Sorry for asking a lot of questions. Thank you in advance.

Venura

#2

Hello,

Glad you’re planning on using the microscope.

  1. We cut our own filters for the beam splitter. We order the filters as a glass sheet and cut it ourselves to the correct dimensions. I don’t know what a suitable supplier for that is in the US, in the UK we tend to use Comar Optics. They do ship to US, but I imagine there will be a cheaper, local supplier.

  2. I’m sure if you choose your filters specifications correctly you will be able to remove the autofluorescence. You will probably need to get different filters for the autofluorescence and GFP images, depending on which wavelengths you want to keep or remove.

  3. The major excitation wavelength of GFP is 395 nm so you will want to find an LED which is as close as possible to that. For example this LZ1 star led.

Hope that helps!

Samuel

#3

Thank you Samuel,

I ordered a set of filters from Thorlabs. Hope they will do the job.
I will keep you updated. So excited!
have a great day,

Venura

1 Like