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Duckweed in UV

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#1 nfoto

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Posted 08 September 2019 - 16:21

Currently I'm working with various duckweed (Lemna) species for my ongoing aquatic macrophyte documentation project. This necessitated working at magnifications >> 1 as these plants are tiny. A "big" Lemna might be 4-5mm on the major axis and most specimens are much smaller. I run most photos at the 3-5X range provided by my Laowa 25mm f/2.8 Ultra-macro lens, or 5-10x with the various Mitutoyo objectives at my disposal. Alas none of these optics can be used for UV.

During the study I observed flowering duckweeds so only natural to want to shoot these über-small flowers in UV too. We are talking about flowers reduced to the very basic, meaning 1-2 stamens and 1 style maximum per flower, all wrapped in a gossamer-like short spatha. They are often hard to detect by the naked eye as the entire package might only be a fraction of a mm in size. I examine my duckweed collections under a stereo microscope thus any occurrence of flowers is "easy" to detect, yet very far from easy to photograph thanks to the minute size and the fact most parts are hyaline.

To get adequate magnification, I cobbled together a makeshift solution comprising the UV-Nikkor as master lens with various optics reverse-mounted to its front. The Noflexar 35/3.5 proved to be a nice combination, as did the EL-Nikkor 50/2.8 and 63/3.5. The Noflexar combo reached 4.4X so was the preferred approach.

For visible-light photographs I used a Z7 coupled to the Laowa lens, with cold-light fibre optical light (FI Heim, Switzerland). For UV I pressed one of my Broncolor studio flashes (1600 Ws) with uncoated Xenon tube into service in conjunction with the Nikon D3200 (internal Baader U filter).

Here is a snapshot of the "duckweed corner" of my studio.

Attached Image: U201909072899_birnas_UV_lab_duckweed.jpg

A duckweed flower in visible light (shot at 5X, cropped). The entire frond is approx. 1.7mm long.

Attached Image: X201909073810-duckweed_flower_VIS.jpg

Here is the very same specimen, now shot in UV and cropped slightly.

Attached Image: X201909073875-duckweed_flower_UV_4,4X_crop.jpg

(the spooky shadow behind the flower in UV is from a small mosquito larva that suddenly became active in the UV session!)

#2 Andy Perrin

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Posted 08 September 2019 - 16:59

Neat! Are these images going to be focal stacked next?

#3 nfoto

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Posted 08 September 2019 - 18:06

They are already stacked in Zerene Stacker. Twenty to forty frames each. One has to limit the number of exposures whilst shooting UV macro as the beefy output from the studio flash up close can induce wilting of the specimen, even when they are "water-cooled" as in this instance. On the other hand, exposure is no problem. I ran the UV @4.4X with nominal f-number 8 and ISO 100. The effective f-number is much smaller, probably around f/40.

#4 dabateman

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Posted 09 September 2019 - 03:38

Excellent photos. I also like the setup image. That tilt slidding macro arm is amazing.
I still need to test my objectives for uv transmission. I have some cheap Chinese 1x, 2x and 4x which I think were ok for UV and working distance. I can't remember if my zeiss 16x was good. I also can't remember if either my 40x or 63x were good and I know I didn't test my 25x or 80x.
The 80x is really hard to focus as its zero corrected. So you just see the surface of objects. Which makes sense as its a metallurgical objective.
I don't have a fancy stage though, I just recently got 2 iShoot macro focus rails. So can now adjust in fine increments. I still need a lifter to get the third dimension. I will most likely get one in October with ebucks.
As for holding the sample I cut a square 75x75 mm out of a old TV table and etched in a groove to hold hold a UV transmitting PMMA sheet. Then a shine UV light up through my sample. I can also direct a uv light on to the surface.
This is how I shot a uv trans image through a flower, way back. Still need to follow up on the setup. I have some odd mold and cells I want to image.

#5 JMC

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Posted 09 September 2019 - 08:25

Amazing pictures, and that is quite a setup.
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#6 GaryR

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Posted 09 September 2019 - 13:24

Monstrous setup, amazing images and backstory.
Had to check out Zerene Stacker...thanks for the info.

#7 nfoto

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Posted 10 September 2019 - 20:33

I'm a little surprised the support stage wasn't mentioned.... this is the "Lego" system of science labs in their time, i.e. the versatile Nikon Multiphoto photomacrographic system. It catered for formats up to 4x5" film via a multitude of extra parts and bolt-ons. The support structure is extremely rigid and very heavy, and is equipped with rubber-dampened adjustable feet. I have several units of them and most of the components so can set up easily for a specific task.

Here is snapshot of a Multiphot set up for 10X photomacrography with the Z7 and the Mitutoyo 10x/0.28 objective.

Attached Image: U201909092907.jpg

Stacking is done by a Stackshot rail mounted on the substage thus not lifting the camera rig; the camera stays put. Illumination is provided by a Novoflex cold-light device, or can be done by studio flash (for UV). I'll shortly experiment with various short focal length lenses to learn what they can achieve in UV terms. I recall doing some UV work using an impromptu setup with a Canon 20mm f/3.5 macro lens and wish to extend the experiments using the Multiphot instead.

Multiphot systems do occasionally occur on eBay. Beware of the exorbitant asking prices for any of the original four Macro-Nikkor lenses delivered with the system. The optics *are* good, but modern lenses are better. And not the least, tend to be much less expensive.

#8 Andy Perrin

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Posted 11 September 2019 - 04:50

Wow, it looks like the distant future of my TriWave setup:
Attached Image: IMG_7943b.jpg

#9 UlfW

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Posted 11 September 2019 - 07:09

That huge old rig looks very impressive and stable.

An alternative when building from scratch could be to build with some kind of aluminium slot-profile system.
Some companies offer custom cut parts and suitable accessory components.
These kind of parts are not only used for hobby DIY projects but also advanced professional machinery.
With well chosen parts they can be very stable and still easy to reconfigure.
I built some of the structures for my extreme macro photography with such profile systems.
https://www.ultravio...dpost__p__22009

I do not know any american company that are selling this types of profiles and components.
One german company I have good experiences with is Motedis.de
Here is their UK-webshop https://www.motedis....s:::999991.html
Ulf Wilhelmson
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#10 JMC

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Posted 11 September 2019 - 08:12

View PostUlfW, on 11 September 2019 - 07:09, said:

That huge old rig looks very impressive and stable.

An alternative when building from scratch could be to build with some kind of aluminium slot-profile system.
Some companies offer custom cut parts and suitable accessory components.
These kind of parts are not only used for hobby DIY projects but also advanced professional machinery.
With well chosen parts they can be very stable and still easy to reconfigure.
I built some of the structures for my extreme macro photography with such profile systems.
https://www.ultravio...dpost__p__22009

I do not know any american company that are selling this types of profiles and components.
One german company I have good experiences with is Motedis.de
Here is their UK-webshop https://www.motedis....s:::999991.html

I've used that type of aluminium t-slot framing before. Very stable. Ulf, I've had imaging systems come over from the US with that type of framing, however they spell Aluminium weirdly over there ;) , which may be why you haven't found a supplier over there.

Edited by JMC, 10 June 2020 - 13:22.

Jonathan M. Crowther

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#11 Andrea B.

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Posted 11 September 2019 - 17:02

omg, Birna, where were you hiding that stuff when I visited Norway?
What an amazing setup!!! That thing must weigh a ton (metric or English whatever)!!

Is that the Stackshot you got from me? I'm very pleased to see it in use!

Who makes those bendy-arm little lights? Are they LED? Can you get UV versions?

....flowers reduced to the very basic, meaning 1-2 stamens and 1 style maximum per flower, all wrapped in a gossamer-like short spatha.
This is soooooo cool to see! And how incredibly interesting that there is a small UV-absorbing tip. This has to be for protection, don't you think? Rather than some kind of insect-visible UV-signature?

I've wanted to see a sample of your aquatic work for ever so long. Thank you for posting this!
Andrea G. Blum
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#12 nfoto

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Posted 12 September 2019 - 12:48

Glad you enjoyed this small glimpse into the world of aquatics, Andrea. When you visited Norway the gear still wasn't unpacked after my last move, but the last years project on documenting aquatic species necessitated digging out the Multiphots and their various components once again. Still haven't located all the pieces though, but the systems now are functional. And yes, one of the Stackshot rails is the one I got from you many years ago :) Cognisys, the maker of Stackshot etc., does make some pretty useful equipment for specialised photography. There are improvements one can add by oneself too, as Cognisys strangely enough hasn't fully acknowledged the superiority of the Arca-Swiss clamp system. I might get a third rail system, but not decided on this purchase yet.

The mirrorless cameras (mainly Z6/7 in my case) are faster and simpler to operate on these high-magnification systems than the traditional DSLRs, but provided one uses mirror lock-up and shutter pre-release (easily accomplished by configuring the Stackshot controller), the latter camera systems do still have a slight edge for many situations. Plus they are less prone to getting a dirty sensor during the long and tedious stacking sessions.

The illumination shown here is cold-light fibre optics, which can be augmented by UV flash if necessary. Many companies make such light sources and the goose-neck flexible rods to transmit light from the source to the object under study. Very convenient as one can have high wattage output delivered without any heat .... I have several such illumination systems; those made by FI Heim (Switzerland) only allow using projection halogen lights (which can be extremely bright), whilst a system by Novoflex combines cold-light fibre and microflash. The latter supports using the Broncolor studio flash as light source.

As to the "basic" flowers of the duckweed, they are highly reduced yet functional and one observes fruits once in a while. The fruit is so big it almost makes the little duckweed frond to capsize!!

Attached Image: X201909063764_duckweed_fruit_5X.jpg

#13 dabateman

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Posted 13 September 2019 - 02:34

That kind of reminds me of a budding yeast. Interesting how small that thing is.
I will have to show a photo of my setup. Much simpler than Andy's. Just tripod, macro rail, camera. I recently got a touch monitor to add in for hands free camera adjustments. But still haven't connected it yet.


#14 Andrea B.

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Posted 13 September 2019 - 16:44

Such an incredible photo !! All that amazing detail in one tiny little duckweed leaf.

I don't know if I would have the patience to create these detailed macro photos. :rolleyes:
Andrea G. Blum
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#15 Andy Perrin

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Posted 13 September 2019 - 17:31

It looks kind of like a rubber ducky.

#16 nfoto

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Posted 19 October 2019 - 19:24

Last UV duckweed series has been with the combination of Cassar-S 50mm f/2.8 reverse-mounted on one of my UV-Nikkor lenses. I get up to m ~ 4.2 with this combination and it obviously lets a lot of UV through.

The Cassar is set to the working aperture (f/8 mostly) and the master lens, UV-Nikkor, is used wide open. I cannot detect any vignetting thus the two lenses are optically well matched in this configuration.

The working distance is < 4 cm thus on the short side, yet suffices to allow lighting of the samples. A single step ring allows mounting the Cassar onto the master lens.

This is an overview of the configuration. The Cassar housing is all aluminium but the finish is not very shiny thus flare caused by the lens' exterior is minimal if present at all. The flash is one of my Broncolors with 1600 Ws output and uncoated Xenon tube, giving plenty of UV output for the occasion. Apparently this UV-Nikkor is the oldest one as it is labelled with my old name and address, so must be nearly 30 years old now - -yet still going strong. This stuff was built to last.

Attached Image: U201910194025_Cassar50f2,8_UV-Nikkor.jpg

Overall results were very good in terms of sharpness and contrast. The image below is a 100% crop from the periphery of the frame, showing excellent detail of surface cell, stomata, and the elongated cells of the root hair of a Lemna turionifera. My impression so far is that this way of stacking lenses to reach 3-4X magnification for UV photomacrography is equal or better than what the various EL-Nikkors achieved. And the orange colour cast of the unprocessed NEFs indicate the stacked lenses reach pretty deep into UV-A as well.

Attached Image: X201910194059_corner_100pct_crop_Cassar50S_UV-nikkor_D3200_BaaderU.jpg

#17 Andy Perrin

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Posted 19 October 2019 - 20:39

Very sharp indeed.

#18 nfoto

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Posted 04 June 2020 - 18:53

My new dietary regimen excludes bananas, but such fruits can find other fields of application besides being eaten Posted Image Some of my Duckweed cultures are grown in enclosures with slices of banana or apple inside; this to generate ethylene gas. I do this to trigger formation of gibbous fronds of some species (such as Fat Duckweed Lemna gibba, which unfortunately can exist in a non-gibbous form since these plants don't read the Flora description ...). Ethylene, being a gaseous growth hormone, is claimed to induce gibbosity if the culture can produce such growth form.

After about 5 weeks sure enough. some fronds tend to be more swollen, so that's fine as I already have identified them as Lemna gibba on other criteria. However, with these morphologically reduced and highly plastic aquatic species, there is always a question about their identity unless true differential criteria are observed. Hence my culture experiments.

What I hadn't thought of beforehand is that ethylene apparently can trigger flowering as well. The putative L. gibba culture started flowering yesterday, many weeks or even months before any flowering would have occurred in the wild. The Duckweed flower is truly minuscule, comprising 1-2 stamens and a style all of which is enclosed in a basal, hyaline spatha.

Here is a UV capture of one such flower, shot at 4X magnification with a reversed Coastal 60mm f/4 on my UV-only modified Nikon D3200.

Attached Image: X202006044367_the_tiniest_flower_of_them_all_UV_D3200.jpg

The small object on the left, looking like a minute apple, is in fact a pollen grain of a Scots Pine Pinus sylvestris. That pollen grain is 65 µm (0.065mm) so sets the overall impression of scale here. The complete stamen with filament and anther is around 400 µm. A normal human hair is around 75 µm (Europe) to 100-120 µm (Asia). Not much size-wise for this flower, but it does have what is required to ensure a future seed set. Because the Duckweeds have such efficient vegetative multiplication and dispersal, one might wonder why the plant bothers to flower at all. Maybe a fall-back option to ensure a tiny chance of genetic recombination? Nature as usual answers with more questions.

#19 Bill De Jager

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Posted 04 June 2020 - 23:44

Birna, this is very informative and absolutely amazing. We have Lemna in California, but I think I first noticed it in canals in the Netherlands (where it can be very abundant) when visiting family as a child.

At present I have several bellows systems and suitable lenses which I've used for a few macro photos on tabletops. I see the advantage of an upright system so the camera and lens can be held rigidly stable for stacking, and so the subject can rest on a surface and be photographed from above with more lighting flexibility. Do you have any suggestions for a relatively affordable support system suitable for a beginner in macro photography? Someday I may try to get a Multiphot, but there is no point in making that financial investment unless I know can put it to good use.

#20 OlDoinyo

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Posted 10 June 2020 - 02:23

It amazes what can be seen if one looks closely enough. I have regarded Lemna mainly as common pond scum most of my life, never suspecting all these details (though I did hear somewhere that they are among the smallest angiosperms on earth.) One wonders how pollination of such miniscule structures occurs.

Edited by OlDoinyo, 10 June 2020 - 02:26.