Problem: trying to image Cy7.5 with Zeiss AxioObserver but fluorescence illumination light path appears to cut light above approx 700 nm.

We are using a pE-4000 LED illuminator to deliver light to the microscope fluorescence input at the rear. We adjusted the LED powers such that 635, 660, 740, and 770 nm all have 52 mW at the end of the liquid light guide.

We have removed the IR filter for the autofocus system that is in the objectives nosepiece. However, the light at 740 and 770 nm is blocked from arriving at the objective lenses.

Can this be corrected somewhere else in the lightpath or are we unable to use this microscope for illumination for Cy7.5?

The next question is that Zeiss says that their camera will image out to 850+ nm, but does anyone know whether the optics will pass this?

(Power less at back aperture with no lens because some misses power meter probe.)

Because we were using a 50/50 mirror probably purchased over 20 years ago for this test, the following readings bouncing the light off the mirror with no other optics shows that the mirror is not the source of the loss of power.

Lightpath where we think IR is being blocked: