Diffuse Köhler?

GaryB · Post by **GaryB** » Sat Mar 04, 2017 2:38 pm

I was just reading a Microscopy-UK article
'The Fly In The Ointment
An Achilles Heel in Köhler illumination?
By Paul James.

http://www.microscopy-uk.org.uk/mag/art ... j-fly.html

Discussing the merits and demerits of both Köhler and diffuse illumination and it suddenly struck me that it may be possible to mix both methods. It's something I've been doing for a while due to my limited equipment and 'making do'. It seems to me there is a good compromise for mixing the two.

The argument goes that with a diffuser in front of the light you lose the filament, or in my case, the LED, and at that point the diffuser itself becomes the light source. In my setup I have a 10 watt Jansjo IKEA lamp with a ping pong ball cutout to fit on the front. On the face of the ping pong ball I drew a small cross. I use a Nikon 50mm 1.8 camera lens as my lamp condenser and lens's internal iris as the field iris.

This setup allows me to easily focus my condenser on the Nikon field iris and step up and down as needed. I then adjust the lamp position so the cross on the ping pong ball focuses sharply on the condenser iris, and is seen clearly in the objectives back plane effectively giving me diffuse Köhler. The only difference I'm seeing as far as setup procedure goes is that I get a cross rather than lamp filament in all the right conjugate planes.

The illumination looks very good and even. Very east to setup and because they're separate elements, easy to optimize. It's essentially setup the same as Köhler but has the advantage of a wide diffuse source. I'm not seeing a downside to this from my limited perspective but I'd love to hear other opinions or methods.

Pau · Post by **Pau** » Sat Mar 04, 2017 4:56 pm

Many (most?) microscopes also do it, for example with a removable diffuser placed in front of the lamp.
This is something that has intrigued me for some time as when the same makers at their websites explain Kölher they always tell to focus the lamp filament in the classic mode.

All the (four models) Zeiss illuminators I have do it, sometimes with removable and sometimes with non removable frosted glass.

My provisional conclusion is that they use this trick as a compromise to have enough even illumination with low power objectives where strict Kölher doesn't apply (just based in my limited experience and somewhat different of the explanation at the article you linked)
Now, with my DIY LED illuminators and original focusable collimating lenses I much prefer to set up true Kölher without diffusers with 25X and upwards objectives

Gary, thanks for the link, very interesting!

GaryB · Post by **GaryB** » Sat Mar 04, 2017 11:42 pm

I think your diffuser trick conclusion is right. As the article states, doing ideal Köhler does take quite a large chunk of space and it seems that light path compression and poor condensers with a lot of in-built lighting may be why using diffusers is popular.

I guess a lot of it harkens back to older (cheaper) condenser types with a fixed higher NA like mine has. It tends to make Köhler a royal pain for every day use going from a 3.5x upto to a 44x .85.

That site is amazing, the articles there have been keeping me occupied for days

I'm currently working through index page 3 which has some great stuff on lighting of all types.

http://microscopy-uk.org.uk/mag/libindex3.html

Choronzon · Post by **Choronzon** » Mon Mar 06, 2017 5:24 pm

So then, what is the difference between diffuse Kohler and diffuse Critical Illumination?

Beatsy · Post by **Beatsy** » Mon Mar 06, 2017 6:07 pm

Choronzon wrote:So then, what is the difference between diffuse Kohler and diffuse Critical Illumination?

Same difference as when they are not diffused, though critical does usually employ a diffuser because the filament image is coincident with the sample image (conjugate planes). With Koehler, the filament image is coincident with the condenser diaphragm.

GaryB · Post by **GaryB** » Mon Mar 06, 2017 8:23 pm

Yup, critical still has the diffuser in view with the subject. It's why even with a diffuser, we mostly still setup for Köhler.

Pau · Post by **Pau** » Tue Mar 07, 2017 4:37 am

Theory aside, in practical terms for critical illumination you don't focus the light source at the specimen plane because the diffuser texture will be visible and disturbing, you defocus it a bit.
For low to moderate magnification critical I. can work very well, for high magnification Kölher is clearly superior.
Kölher with diffuser is what I name (only for myself) semi-Köhler at least with microscopes like mine where the diffuser is not opaque enough to be considered the light source; your ping-pong ball could be, although a more opaque diffuser eats much more light.

GaryB · Post by **GaryB** » Tue Mar 07, 2017 12:44 pm

Yes, the ping pong ball completely diffuses the LED, much more than a frosted glass screen, which is good for me as the LED is blindingly bright, the ping pong ball is just very bright. I don't mind what people define it as, I have no axe to grind and as long as it works as intended it's all good in my book. It's a compromise, like most things.

Hopefully the situation may be moot soon as I plan on getting a Zeiss Standard soon

GaryB · Post by **GaryB** » Tue Mar 07, 2017 4:05 pm

Just found something interesting from a Zeiss manual about their use of diffusers.. not exactly what we were thinking.

"When adjusting the equipment for Köhler illumination it may happen that a certain granularity is noted which is superimposed on the specimen over the entire field of view. This is the texture of a ground glass screen or an etched collector lens which are intended to cover up the irregularity of the source. If this plane should accidentally be in sharp focus as well, it is only necessary to alter the height of the condenser slightly"

So there we have it from the horses mouth.

ChrisR · Post by **ChrisR** » Tue Mar 07, 2017 6:05 pm

At school a youngster asked me what I was doing while setting up a microscope. I mentioned Köhler and immediately wished I hadn't. Not being a proper teacher I didn't have an answer ready for the follow-up. I said it was about getting the subject properly lit without getting images of the light source in the way. Close enough?

Beatsy · Post by **Beatsy** » Wed Mar 08, 2017 3:05 am

My favourite description of Koehler illumination (read in passing somewhere) is "setting the condenser so that each point on the subject is illuminated by every point on the light source while each point on the light source illuminates every point on the subject".

Edit: ChrisR - your description was close enough for the situation. I'm not claiming the description I quoted would have been appropriate there.

Pau · Post by **Pau** » Wed Mar 08, 2017 9:16 am

GaryB wrote:Just found something interesting from a Zeiss manual about their use of diffusers.. not exactly what we were thinking.

"When adjusting the equipment for Köhler illumination it may happen that a certain granularity is noted which is superimposed on the specimen over the entire field of view. This is the texture of a ground glass screen or an etched collector lens which are intended to cover up the irregularity of the source. If this plane should accidentally be in sharp focus as well, it is only necessary to alter the height of the condenser slightly"

Having lots of Zeiss manuals and three Standards...I really know this small problem. My comment is about if this method with diffusers can or can't be considered Kölher and about its need.

GaryB · Post by **GaryB** » Wed Mar 08, 2017 5:52 pm

Yep, I understand.
I read that and and got instant brain ache.. just randomly moving the condenser down doesn't strike me as a good way of getting Köhler... in the getting Köhler part of the instructions.