UV (365nm) and visible (546nm) microscopy of a diatom

[Guest]

Let's say I want to buy used and have a total starting budget of $1400. I want a field of view from 2mm to 0.2mm. Working with UV reflected and fluorescence. Ability to attach a Sony a6000. Working in the 365nm UVA range and visible as well. Upgradeable. Nikon maybe or something else. I can handle a beater if the parts are available. For example Nikon has a competition ever year featuring microscopic photography. I find that extremely fascinating. For example, I would like to put it together in phases $600 plus $400 plus $400 etc then accessories and replacement parts. The problem listed above is buying the wrong stuff and over spending. Without lab knowledge I would have no idea what to buy, who to ask or where to start. 

 

I have a canon macrophoto 20mm f3.5 rated up to 20x. I also have a Tominon 17mm and a special microfiche lens. None of them are easy to use on a tripod and I don't think they are any good for UV. Would I be better off getting a solid macro stand or just diving into the microscope realm?

[dabateman]

BlazerOne, 

Birna would be the expert to address that from an imaging point of view. Especially with her duckweed images:

https://www.ultravioletphotography.com/content/index.php?/topic/3489-duckweed-in-uv/

 

I will have to think about it as well.  The Sony A6000 sensor is 23.5mm wide, so you are asking for 11.75x to 117.5x magnification range if you want 2mm to 0.2mm to fit the sensor width. 

So you are just outside the range of a disection microscope but not by much. 10x objective x 10x optical path = 100x magnification. 

For reflectance, an inverted scope or metallurgical scope and objectives might work for you. Like this (see figure 4):

https://www.olympus-lifescience.com/en/microscope-resource/primer/anatomy/reflected/

 

You would want a good 10x or 16x objective.  They aren't too hard to get.

 

[Guest]

Ok wow! So, to match Birna's setup all I need is a pile of dirty laundry, an automobile lift, the arms from Dr. Octopus, a neutron star, and two lenses kissing each other. 

[dabateman]

Maybe 

Now you have me searching again. 

This looks really cool, but can't do reflectance as no EPI head:

https://www.ebay.com/itm/Olympus-IM-Inverted-Phase-Contrast-Microscope-/255022994029?mkcid=16&mkevt=1&_trksid=p2349624.m46890.l49286&mkrid=711-127632-2357-0

 

I don't know if the cheap metallurgical microscopes can see UV:

https://www.ebay.com/itm/AmScope-50X-500X-Inverted-Trinocular-Metallurgical-Microscope-/383430987818?mkcid=16&mkevt=1&_trksid=p2349624.m46890.l49286&mkrid=711-127632-2357-0

 

Or from India:

https://www.ebay.com/itm/Radical-600x-Inverted-Metallurgical-lab-Reflected-Light-Microscope-/113665254073?mkcid=16&mkevt=1&_trksid=p2349624.m46890.l49286&mkrid=711-127632-2357-0

[dabateman]

Actually thinking about it more.

I don't think you would be happy with such extreme magnification. 

You might be better off with a Milar lens like these:

https://www.ultravioletphotography.com/content/index.php?/topic/4316-leitz-30mm-f45-milar-macro/

 

These cheap stands have had some great reviews: 

https://www.wemacro.com/?product=vertical-stand

 

Or buying a disection trinoccular microscope.  You have to think about what you want to see, cells or structures. 

[colinbm]

20 minutes ago, dabateman said:

Actually thinking about it more.

I don't think you would be happy with such extreme magnification. 

You might be better off with a Milar lens like these:

https://www.ultravioletphotography.com/content/index.php?/topic/4316-leitz-30mm-f45-milar-macro/

 

These cheap stands have had some great reviews: 

https://www.wemacro.com/?product=vertical-stand

 

Or buying a disection trinoccular microscope.  You have to think about what you want to see, cells or structures. 

This is great stuff Dave 

[Guest]

Thanks for the feedback. The JMC topics are like easter eggs full of goodies. I just have to find them all. Going to keep a look out for the 30mm Milar or 25mm Luminar.

[JMC]

Some new images, and I thought I'd add to this thread rather than start another new one.

 

This is a diatom slide which has had a thin layer of aluminium applied to the samples with the goal of improving contrast for visible light microscopy. The slide was made by John Dale in the 1990s, and I received it as part of a box of slides from the Postal Microscopical Society of which I am a member. Here's the slide.

 

Imaging with done with my UV microscope, and are simple brightfield transmission images with the light below the sample. This was done with my monochrome d800 Nikon.  I was limited to visible and 365nm with this as the sample was made with a normal glass slide and coverslip. Resolution has been dropped massively for sharing, as these were full frame images originally.

 

First though before the UV images, this was a visible light one at 546nm with a relatively low power objective to give more of an overview of the sample.

 

With the low power objective above, you can see the cracks that have formed in the Al layer over time.

 

And now at 365nm with a Nikon UV-F 100x glycerin immersion objective.

 

 

 

 

 

 

Apart from the first image of the UV images (which was stacked) these are all single photos. The camera produces a slight colour cast in the UV, which I normally remove, however I have left it here as I quite like it.

 

Also, a video shot with the same setup at 365nm;

 

The video shows me moving the sample vertically, and reveals some of the structures in different parts of the diatom.

 

[colinbm]

This is exciting Jonathan.

[photoni]

@JMC as always ... amazing photos

[Andy Perrin]

Very very cool!

[JMC]

Thanks guys.

[Mambo]

Amazing stuff, phenomenal!

 

-May I ask what your exposure settings are in the yt video clip?
Curious if we can help get rid of that banding (or not).
Looks like your shutter is out of sync with the lights causing flicker/banding.

[JMC]

10 minutes ago, Mambo said:

Amazing stuff, phenomenal!

 

-May I ask what your exposure settings are in the yt video clip?
Curious if we can help get rid of that banding (or not).
Looks like your shutter is out of sync with the lights causing flicker/banding.

Thanks, I have gone back and checked and it was 23 frames per sec. Can't remember the ISO setting the camera was on but it was high.

[Mambo]

Is that the video frames per sec? -I was thinking more along the lines of the exposure settings. 100th of a sec? faster/slower?
 

Also it can be tricky going back and forth between stills and video!

[JMC]

1 hour ago, Mambo said:

Is that the video frames per sec? -I was thinking more along the lines of the exposure settings. 100th of a sec? faster/slower?
 

Also it can be tricky going back and forth between stills and video!

No idea, sorry, can't remember the original settings. The mercury xenon lamp has a flicker to it at something like 500Hz from memory, so this'll probably be the problem.

[JMC]

Update from a diatom slide imaged yesterday - Cymatopleura elliptica, made by Klaus Kemp. The image was done using 365nm light on my modified Olympus BHB microscope and is a stack of 18 photos. Objective was a Leitz 100x Pl Apo NA 1.32-0.60 (oil immersion). Condenser a Watson Holoscopic one (again oiled). The objective iris was closed down slightly with the aim of producing circular oblique lighting. A Nikon 5x photoeyepiece was used along with a monochrome converted Nikon d800. Light source - Zeiss 50w HgXe. Filters - 2x 365nm, 10nm bandpass Edmund Optics. Reduced in resolution for sharing, although the crop is at original resolution.

 

 

Cropped image at original resolution.

 

[colinbm]

Pretty good Jonathan. Is this visible to the naked eye, or only because it is on a slide ?

 

[JMC]

1 minute ago, colinbm said:

Pretty good Jonathan. Is this visible to the naked eye, or only because it is on a slide ?

 

Not sure I follow you Colin. The diatom is about 100 microns long so would be visible to the naked eye.

[colinbm]

Thanks Jonathan
I have some small Micrometeorites this small & if you didn't know where to look you wouldn't see them.

 

[JMC]

No problem Colin. Yes dealing with small subjects can be challenging. All of my samples are on microscope slides which makes things easier, but sometimes even finding them on the slide can be interesting.

[JMC]

Another diatom, this time Amphipleura pellucida, imaged on my Olympus BHB microscope (a 63x Leitz Pl Apo NA 1.4 objective) using 365nm UV light and with a monochrome converted Nikon d850 camera. Image captured as a 10 shot average NEF file in the camera, and the processed in Monochrome to DNG, Darktable and Photoshop. Cropped from the original image.

 

 

[colinbm]

Good Jonathan. No stacking ?

[JMC]

31 minutes ago, colinbm said:

Good Jonathan. No stacking ?

No, no stacking with this one Colin as the diatom is so thin and the interesting bits were in focus with the stage in one position.

[colinbm]

Ok, it must be thin. At that magnification I am using 0.6um stacks & they seem too big ?