Why so blue?

Donw · Post by **Donw** » Fri Jan 17, 2014 7:15 pm

This is a new objective for me.
Reichert (Austria) 1727

10X/.30
Plan Fluor
Infinity/-

Condensor is AO NA 1.40
Aplanatic-Achromatic

Microscope is AO H20, 2.8X Relay lens, 100W Halogen

NO FLASH

Camera is Canon T3i, Custom WB

This photograph is just what I see in the eyepiece.
While it has diminished the purple halo of my achromats, the subject itself is blue. This is also true of transparent subjects like rotifers and copepods.

Is there something I'm doing wrong, or is it supposed to look like this?

rjlittlefield · Post by **rjlittlefield** » Thu Jan 23, 2014 1:41 pm

I'm popping this to the top of the list in hopes that one of our experienced microscopists can provide a better answer.

For myself, this looks like an optical effect but I'd hesitate to say you're doing anything "wrong". I'm wondering if you're seeing a form of CA in the condenser that was not obvious with your previous achromats.

--Rik

Ken Ramos · Post by **Ken Ramos** » Thu Jan 23, 2014 7:22 pm

Well I am not an experienced microscopist but I would agree with Rik, it looks like an optical effect, and a nice one too in my opinion. Seems like I have seen something like this before.

Franz Neidl · Post by **Franz Neidl** » Fri Jan 24, 2014 4:57 am

Hello Don;

I dont know the AO-microscope, but I would controll if the infinity-objective is a good objective for your microscope (which probably is not infinity?).

The second thing I would do is to "play around" with the White balance of your camera and see how the things are changing.

Good luck!

Franz

René · Post by **René** » Fri Jan 24, 2014 5:31 am

I've come across a similar phenomenon with a specialized Olympus series, the dplanapo UV 20x/0.80 oil immersion. Everything gets surrounded with a bluish halo. Good example to show this is something slightly opaque like these Microcystis cells (4um diameter):

S-PlanApo 20x/0.70 (dry)

D-PlanApo 20x/0.80 (oil)

With monochromatic light (550nm) and the camera set to B/W, the situation looks better for the dplanapo, as you would expect with the slightly higher NA of 0.80 vs. 0.70:

S-PlanApo 20x/0.70 (dry)

D-PlanApo 20x/0.80 (oil)

(Olympus IMT-2 inverted, 0.55NA condenser, Imaging Source 5Mp cmos cam via 0.75x relay. Images at 100%, unadjusted for contrast/gamma)

Now this DPlanApo objective is a specialized series, produced some 35 years ago. The NA was enormous for this kind of magnification, so I suspect this abberation is a trade-off. It was originally ment to be used for fluorescence, not brightfield. This effect hardly matters for fluorescence work.

What it is I do not know for sure, I regard it as axial chromatic abberation. Why your Reichert lens shows the same effect, I do not know, as it seems otherwise bog standard. Did it perform OK on a Reichert stand?

Best wishes, René