Proof of concept: UV-reflectance stacking with MP-E 65

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
This right up front, this is not a great picture, it is a proof of concept, to be refined.

I am interested in UV reflectance photography and to see whether there are insect patterns not visible in "normal" light on some minute orchid flowers. The flower is 1.76 mm (no, not a typo) wide. Shot with full-spectrum modified 5D2, MP-E 65 at f/2.8, manually stacked 10 frames, Baader UV-pass visible opaque filter, ISO 400, about 4 second exposure, illuminated with single UV 365 nm LED flashlight.

First off, the MP-E 65 DOES transmit some UV. That's already great. I also have a Nikon EL 80/5.6 but still waiting for some parts to arrive to put it all together. Should have much better UV transmission, so will be interested to see the differences.

Further refinements are additional UV flashlights (can't diffuse them because of absorption and fluorescence issues), and stacking with StackShot (have that).

Some single shots on some other orchid flowers do show some different patterns, so that works as well. The dotted highlights, those are the individual cells, with their rounded surfaces (verified by scanning electron microscopy). In visible light, the flower is uniformly green.
 

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Reactions: Berowne

Maximilian

The dark side - I've been there
Nov 7, 2013
2,408
219
Germany
Hey that's great!

Although I am personally not interested in doing something like that myself, I know about this UV insect patterns and I'd like to see more if you continue working on that.

Thanks for sharing and good luck!
 

lion rock

EOR R
Jan 1, 2013
1,920
37
It is amazing to try unusual techniques on everything. This opens up another avenue of opportunities.
Glad you've started this method. Please keep perfecting the lighting and send us samples of the various "targets."
We do have to innovate!
-r
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
Glad this generates some interest. I will post some more as they come along. Loads of fun. And may even get a paper out of it :)
 

LordofTackle

EOS RP
Nov 25, 2014
291
0
Zeidora said:
Glad this generates some interest. I will post some more as they come along. Loads of fun. And may even get a paper out of it :)
May I ask what you do for work? :)
Sounds like biologist/scientist.

And keep the good work coming :)

-Sebastian
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
LordofTackle said:
Zeidora said:
Glad this generates some interest. I will post some more as they come along. Loads of fun. And may even get a paper out of it :)
May I ask what you do for work? :)
Sounds like biologist/scientist.

And keep the good work coming :)

-Sebastian
I am a natural history museum curator, officially for marine invertebrates, but lately I've broadened a bit into orchid systematics. I also run the museums scanning electron microscopy facility, and am the go-to person for any odd scientific imaging needs. Apart of the SEM, all the photo stuff is personal. It would be foolish to ask the museum to buy a $400 filter for me. So I just get that myself. I enjoy an imaging challenge, particularly in the macro realm. Like using cross polarization to image harpsichord strings for a multimedia presentation of a baroque orchestra. Or getting z-stacking going on my compound microscope.

So Ph.D. in marine biology, and a bunch of classes in scientific photography in dept phys chem at the University of Basel, Switzerland (Gschwind and Heilbronner were great). Then building on that.
 

LordofTackle

EOS RP
Nov 25, 2014
291
0
Thanks for the answer. Sounds like a really cool job and it must be very nice to be able to combine job and passion that way. :)

And I have to ask: is Heilbronner a person? Cause I am from the (german) city Heilbronn....^^
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
LordofTackle said:
Thanks for the answer. Sounds like a really cool job and it must be very nice to be able to combine job and passion that way. :)

And I have to ask: is Heilbronner a person? Cause I am from the (german) city Heilbronn....^^
Renée Heilbronner was one of the two instructors of the various classes. Gschwind at some point got a position in Zürich, not sure about Heilbronner. It's been a mere 20+ years!

I know Heilbronn, but mainly for the traffic jams on the Autobahn ;-)

And, yes, I'm darn lucky having this job. Curator jobs are pretty impossible to get. Best career advice I got many moons ago was "Mann, vergiss es!" [Dude, forget about it]
 
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LordofTackle

EOS RP
Nov 25, 2014
291
0
Zeidora said:
I know Heilbronn, but mainly for the traffic jams on the Autobahn ;-)

And, yes, I'm darn lucky having this job. Curator jobs are pretty impossible to get. Best career advice I got many moons ago was "Mann, vergiss es!" [Dude, forget about it]
Ad it's gotten waaaay worse since then ;)
Seriously, that highway square between Karlsruhe, Heidelberg, Heilbronn and Stuttgart is a major PITA

I'm currently in Konstanz, were I studied...biology :D
also not that far Basel
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
Here's another installment in my project. Today it's mainly about post-processing. Still not great photographs, but this is about getting technique sorted out.

One problem is that during camera conversion, some of the Bayer filter elements most likely got damaged. This is no big deal for single captures, but if you stack 50 or more, one gets a lot of "creative" color streaks. That leads to a lot of post-processing after stacking. I thought of doing a dust delete data in the 5D2 body, but that does not work, because during conversion, the dust shaker hardware is removed, and without it, the dust delete data cannot be obtained.

But because these color dots are in fixed positions, they can be screened out with a Gaussian blur mask. Do the following in Affinity Photo:
- Open a dozen images in AP.
- Add a Gaussian Blur live filter on top of the first image.
- Add a mask to the Gaussian Blur live filter. Blur of about 2.5 pixels worked well for me, but that depends a lot on your camera and the type of Bayer damage you have.
- Fill mask layer only with black.
- Paint with a small white brush (4 pixels worked well for me) over any color spots in the mask layer. Do that at 150-300%. You'll spend some quality time at this. Don't bother fix those color spots right at the edge of the frame. The Gaussian Blur from the edge of the image is going to mess it up.
- Once done with first image, copy-paste Gaussian blur filter with Mask into next source image.
- Check for pesky color spots in second image and refine mask.
- Copy-refine for the the rest of the images.

Open one of the CR2 source images, develop it. Now start macro recording and paste the final version of the Gaussian Blur live filter with mask on top of the image layer. Stop macro recording and save it as something like "UV prep". The cool thing in AP is, that the live filter with mask is saved in the macro step. So the macro does not paste from computer memory, but the stored filter-mask. That means even if you quit the application, the Blur-Mask is still stored. Super cool.

In Open-batch, select the Macro, select your output format (e.g., tif for Zerene), add files, and press OK [Another cool thing in AP, saving/export is not part of macros, but is in the batch dialog. No more infuriating confusions about save functions as in PS]. I am extremely impressed with AP batch processing. All 12 cores on my MacPro desktop go continuously full throttle (100%) for several minutes. I have not seen that with any other application. They really program the batch processing well.

I also tried APs focus merge function, and am quite impressed. PS is worse than useless for serious macro stacking, so most people use either ZereneStacker or HeliconFocus. AP is quite impressive, but Zerene still seems to have the edge. I have not found a way to touch up the final stack layer by layer, as you can in ZS or HF. If I stack with the color artifacts still in place, AP has trouble with proper alignment, while Zerene nails it. Last but not least, tone mapping is much better in Zerene than in AP. This could be fixed before feeding the files into AP.

One problem with APs tif export is, that Zerene only reads them as B&W file (or single channel?). Feeding the same files into AP gives me a color image. Not sure yet what is going on there, but am reasonably sure that I can figure that one out.

Attached four images.
- Stacktrail Zerene. Notice the color streaks, but clean edges of flower and good tonality straight out of stacking.
- Stacktrails AP. Notice color streaks, and edge of flower is blurry (alignment issue) and image is overexposed.
- AP stack: stack of images after batch processing with Gaussian blur mask in AP. Flower edges are clean, but still a bit over exposed. It seems that AP-alignment is misled by color artifacts.
- B&W Zerene: Cleaned tifs stacked in Zerene, resulting in B&W image (hmmm ....)

Enjoy!

For the botanists, it is Barbosella duesenii
 

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Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
Before I forget, I also thought about reflectors for UV photography. The typical paper reflector does not work, because of UV brighteners in most paper that fluoresce the UV back in as blue light. Plastic does not work, because plastic absorbs UV readily. However aluminum foil works very well.

just 2 more c.
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
One more thing, these latest images were taken with a Nikon EL 80 mm f/5.6. It has about 10x/3-stop better UV transmission, and the MP-E 65 seems to cut off at a longer wavelength. I would have expected worse transmission by the MP-E 65. With bellows and tubes, I can get to about 4:1 magnification.
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
Here's now one that I am reasonably happy with. Oberonia costeriana. Flowers are about 2 mm long.

First image is a visible light stereomicroscope z-stack.
Second is the UV version. No striking patterns, but that is information as well.
The last small methods adjustments include:
- RAW development in DxO.
- Removing color spots in AffinityPhoto. Note that DxO tifs required a new Gaussian Blur mask. Seems that the different RAW developer engines are putting color spots in slightly different places.
- Run AP cleaned-up tifs through DxO once more, so that Zerene will see them as a color file.
- Run stack through Zerene.

Now that methods have been sorted out, the real fun begins. Off to the greenhouse ...
 

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zim

EOR R
Oct 18, 2011
1,863
53
Hi Zeidora,
Fascinating stuff really enjoying reading thanks for posting. You should post the same on AP forum I'm sure it would be well received and it would help the devs as I guess there won't be too many beta testers around doing this kind of detailed work!

Regards
 

Zeidora

EOS 7D MK II
Feb 15, 2015
668
10
Zim: thanks for the flowers ;) Posted over in AP forum and also on the Photomacrography forum.
 

Maximilian

The dark side - I've been there
Nov 7, 2013
2,408
219
Germany
Zeidora said:
Here's now one that I am reasonably happy with. Oberonia costeriana. Flowers are about 2 mm long ...
Really fascinating, Zeidora.

If I can recall it right, those UV patterns sometimes look even more geometrical.
I wonder if it's just because these flowers are just so small?
Or are geometrical patterns not so common?

Do you know more about that?