Is anyone doing UV transmission microscopy?

A forum to ask questions, post setups, and generally discuss anything having to do with photomacrography and photomicroscopy.

Moderators: rjlittlefield, ChrisR, Chris S., Pau

jmc
Posts: 238
Joined: Fri Apr 17, 2020 3:14 am

Is anyone doing UV transmission microscopy?

Post by jmc »

As part of my research I'm looking at UV photography - camera sensitivity, lens transmission, filter suitability, effect if light source, etc etc? All of this is done using standard cameras (SLRs and or mirrorless cameras) and while there is some macro work, it's generally no higher magnification than 1x.

I know a lot of people work with fluorescence, but is anyone here doing much work with UV transmission microscopy, in the UVA or even the UVB region? Given my interest in UV imaging, I think it'd be a really interesting area to look at, but I've not been able to find out much work reported on it, other than a few articles online, and the occasional book chapter.

If anyone has tried this I'd be interested to hear how you got on, and what the major challenges were.
Jonathan Crowther

Ichthyophthirius
Posts: 1152
Joined: Thu Mar 07, 2013 5:24 am

Post by Ichthyophthirius »

Hi Jonathan,

Do you want to do macro (let's say down to 10:1) or microscopy?

Regards, Ichty

jmc
Posts: 238
Joined: Fri Apr 17, 2020 3:14 am

Post by jmc »

Hi Ichty,

Good question, and to be honest I am not sure yet, as I'm sure it'll be a hard thing to both acquire parts for and to build. Both reflection and transmission imaging interest me.

However initially I think anything from 4x magnification up to over 50x magnification. I have a couple of lenses suitable for UVB - a Leitz 16x UV lens, and a 52x Beck reflecting objective - so I that is at least one part covered. I've also got a 25mm Luminar which is fine for UVA imaging. I have a couple of monochrome converted cameras which are good for 300nm imaging. I have a couple of different Xe and HgXe lamps which provide a continuous high power UV source.

I'll need some form of light guide for the light sources, and for transmission work a condenser (which I think is now sorted).

Just interested to hear of any experience in the area really.

Jonathan
Jonathan Crowther

Ichthyophthirius
Posts: 1152
Joined: Thu Mar 07, 2013 5:24 am

Post by Ichthyophthirius »

Peter Höbel http://www.mikroskopie-ph.de/ occasionally shows UV images of diatoms for increased resolution in the German forum. He uses a 365 nm LED, oblique illumination, a selection of finite objectives and a monochrome C-mount camera.

High intensity UV and long aquisition times are quite incompatible with live objects. The trend in live imaging is to use at least violet illumination instead (405 nm). But for prepared diatoms it's great!

Examples:
https://www.mikroskopie-forum.de/index. ... ic=14712.0
https://www.mikroskopie-forum.de/index. ... #msg138602
https://www.mikroskopie-forum.de/index.php?topic=1096.0

jmc
Posts: 238
Joined: Fri Apr 17, 2020 3:14 am

Post by jmc »

Ok cheers, will take a look.

Any other experiences welcome.
Jonathan Crowther

Smokedaddy
Posts: 1951
Joined: Sat Oct 07, 2006 10:16 am
Location: Bigfork, Montana
Contact:

Post by Smokedaddy »

Ichthyophthirius, I read in 2014 Peter was using a Imaging Source DMK 72BUC02 (1/2.5 inch Micron CMOS sensor (MT9P031), 2,592×1,944 (5 MP), up to 6 fps ... do you happen to know what monochrome camera Peter is using now? Just thought you might know.

Ichthyophthirius
Posts: 1152
Joined: Thu Mar 07, 2013 5:24 am

Post by Ichthyophthirius »

Smokedaddy wrote:I read in 2014 Peter was using a Imaging Source DMK 72BUC02
I asked him and he's still using the DMK 72 (resolution is sufficient for that kind of work and it works well from 350 nm to 1050 nm).

Lou Jost
Posts: 5944
Joined: Fri Sep 04, 2015 7:03 am
Location: Ecuador
Contact:

Post by Lou Jost »

I just now looked at some of the links to Peter's excellent work. I see some comparisons between white light and UV light, with UV light being much better.

But his tests also show that many objectives do not behave as well as expected in UV light. It migt be more practical and less dangerous to use readily available 405nm visible light, which is easily transmitted by ordinary glass. UV light gives you a 10% improvement in resolution over 405nm light.

jmc
Posts: 238
Joined: Fri Apr 17, 2020 3:14 am

Post by jmc »

Lou, oh absolutely, when used for UV some objectives will fluoresce producing unwanted visible light, or they will have huge focal shifts, or even just not let much light through at all. For some applications 405nm will be fine, for sure.

I've been working with UV photography for a few years, and recently started to look at the possibility of UV microscopy (without spending 100s of 1000s of Pounds). I've picked up a few objectives which will be good for use down to 300nm, where my cameras lose sensitivity, and below (if I ever change cameras). However as I have found out from working with UV photography, things which didn't even need considering for visible light work start to present problems, and those problems get bigger and bigger as the wavelength reduces.

It's mainly because it's an area I find interesting that I'd like to do it, however I see it having applications for the research I do into skin, and other areas too.
Jonathan Crowther

Lou Jost
Posts: 5944
Joined: Fri Sep 04, 2015 7:03 am
Location: Ecuador
Contact:

Post by Lou Jost »

Jonathan, it does sound very interesting! I've begun doing a bit of tinkering with it. Like Peter H, I have a cooled-sensor monochrome camera for this and other monochrome work.

I bought a crazy UV photolithography lens that transmits deep into the ultraviolet. But it has been too heavy to bring back home to Ecuador in my luggage!

Lou Jost
Posts: 5944
Joined: Fri Sep 04, 2015 7:03 am
Location: Ecuador
Contact:

Post by Lou Jost »

May I ask what objectives have worked well for you? And what light source do you use for such short wavelengths? Thanks in advance.

jmc
Posts: 238
Joined: Fri Apr 17, 2020 3:14 am

Post by jmc »

Lou Jost wrote:May I ask what objectives have worked well for you? And what light source do you use for such short wavelengths? Thanks in advance.
Lou, let me just start by saying my microscopy journey has only recently started, so I have yet to start making the UV microscope. However I have been collecting parts, and I have been doing UV photography for a few years (including macro work, down to about 1x magnification).

The objectives I have so far fall into 2 types - specifically designed for UV transmission down to below 300nm, and potentially useful for UV imaging but only UVA not UVB.

1. Specifically designed for UV even below 300nm. I have a Leitz 16x UV objective which seems to show good transmission even down at 280nm, and I suspect it would still be good down to about 220nm. This is a refractive lens, and I'm guessing has quartz and/or calcium fluoride lens elements. I also have a Beck 52x reflecting objective, which again is suitable for down to and below 300nm. I have a 35x one on the way too, but I'm not sure what condition that is in yet.

2. Useful for UVA imaging, but not UVB. I have a 10x Olympus UV FL objective. This has good UV transmission down to about 320nm so is fine for 365nm imaging which would be the commonly used wavelength for UVA work. I suspect the other UV FL lenses would be good for UVA work too, although I do not know whether they have focus shift between visible and UV.

As for a light source I have a couple of Hamamatsu LC8 200W lamps. I have these in HgXe and Xe variants, and they give out light between about 270nm and 420nm. Obviously the HgXe one has strong mercury lines in it, while the Xe one is smoother in spectral distribution. The plan will be to use a flexible light guide to get light under the stage for transmission work.

As for the condenser, I have hopefully just sourced a Zeiss quartz condenser, although I wont see that for a few months (it is in the US and is going to a family member out there who will keep it until I next see them).

I have a couple of monochrome converted DSLR cameras which I'l be using for imaging. With these I can still get an image at 300nm, but not really below that. I may go down the route of a dedicated scientific UV camera at some point, but I'd like to stick with SLRs for now.

Hope that helps.
Last edited by jmc on Mon Dec 28, 2020 9:32 am, edited 1 time in total.
Jonathan Crowther

Lou Jost
Posts: 5944
Joined: Fri Sep 04, 2015 7:03 am
Location: Ecuador
Contact:

Post by Lou Jost »

Jonathan, thanks very much for this helpful information. I am also just exploring UV now, so this is very useful.

kds315*
Posts: 224
Joined: Mon Feb 02, 2009 1:06 am
Contact:

Post by kds315* »

Don't forget the really excellent full color corrected ZEISS ULTRAFLUAR objectives,
which I just happen to have a specific 6x type (a one-off one), but there are others too..
https://www.micro-shop.zeiss.com/en/se/ ... r-10x-0.2#

Zeiss Jena made an UV MIKROTAR lens:
http://www.macrolenses.de/ml_detail_sl. ... tiveNr=366

Bausch & Lomb also made an UV Objective:
http://www.macrolenses.de/ml_detail_sl. ... tiveNr=324

Edmund Optics offers some low mag objectives:
https://www.edmundoptics.com/p/finite-c ... ive/32167/

Here a shot done with a UV-Mikrotar (a Nichia high power UV LED 4x dice):

Image

You found a good place to get answers!

Cheers, Klaus
Last edited by kds315* on Thu May 28, 2020 5:56 pm, edited 3 times in total.
Klaus

http://www.macrolenses.de for macro and special lens info
http://www.pbase.com/kds315/uv_photos for UV Images and lens/filter info
http://photographyoftheinvisibleworld.blogspot.com/ my UV diary

Lou Jost
Posts: 5944
Joined: Fri Sep 04, 2015 7:03 am
Location: Ecuador
Contact:

Post by Lou Jost »

Klaus, thanks, I know the Ultrafluar is good, but very expensive. Are the other ones you mentioned really good? For both visible and UV light, or just UV? I would like to use through-the-objective UV lighting to excite fluorescence.

And why do so many UV objectives require a coverslip? Doesn't a coverslip reduce transmission? Or is it so thin that it doesn't matter?

Post Reply Previous topicNext topic