How to align polarization filters in two filter cubes

Posted by John Oreopoulos on
URL: http://confocal-microscopy-list.275.s1.nabble.com/Strange-artifact-in-confocal-Z-stack-tp593216p593218.html

Search the CONFOCAL archive at http://listserv.acsu.buffalo.edu/cgi-bin/wa?S1=confocal Hello confocal listserver,

I have an unusual situation that I'm hoping someone out there might be able to advise me on. I would like to modify my lab's inverted Olympus IX70 microscope to have the ability to look at polarized light when illuminated in the transmission pathway. I want to polarize the white light that illuminates the sample, and then analyze the polarization of the image by examining the light that comes through the sample with another polarizer that can be oriented parallel and perpendicular to the direction of the polarizer in front of the light source. 

We have DIC optics for the microscope - this means that I have a polarizer that can be fitted on top of the light source condenser, and I also have a fixed position analyzer that can slide in below the fluorescence filter cube turret. My problem is that the polarizer in the analyzer slider is fixed in a direction that is crossed to the polarizer in front of the light source. What I really want to do is to be able to rotate the the polarizer in the slider to the parallel direction as well. I know that Olympus and Leeds sells rotating analyzers that can slot into these positions on the microscope, but my supervisor does not have the funds to afford these right now. So I've come up with a slightly cheaper alternative solution.

We bought two 1" round film polarizers from Edmund optics (~20$ each). These are the exact same size as interference filters that normally get fitted into the fluorescence filter cubes which go in the rotating turret below the objective nosepiece on the inverted microscope. My plan was to take two empty fluorescence filter cubes and insert a polarizer in each - one oriented in the parallel direction and one in the perpendicular direction. Here is my problem: how can I orient and lock the position of the the two polarizers in the filter cubes such that they are aligned with their polarization axis directions correctly? Ideally, since I'm trying to be quantitative with the imaging, I'd like their alignment from their relative crossed position to be within 1 degree or less. When I screw them down with the retaining ring of the filter cube, they rotate slightly in the cube before they become locked. Also, I can't tell if I'm dropping them into the filter holder in the right position since their reference axis position is marked on the edge of the filters. Can anyone out there suggest a good method to overcome this problem? How do the microscope manufacturers align their polarizers in the sliders they sell for DIC imaging, etc?

Thank you in advance for any help!

John Oreopoulos, BSc,

PhD Candidate

University of Toronto

Institute For Biomaterials and Biomedical Engineering

Centre For Studies in Molecular Imaging

Tel: W:416-946-5022