1st ladybird

Just bought that first macro lens? Post here to get helpful feedback and answers to any questions you might have.

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Carmen
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1st ladybird

Post by Carmen »

Below are my first humble photomacrographic attempts at a common ladybird. I suspect it may be harmonia axyridis.

First, at approx' 2x. The optical train:
Nikon CFi 4x Plan Achromat
RAF Camera M25x0.75 to M67x0.75 thread adapter
UV Slim filter (approx' 3.3mm)
Canon 100mm f/2.8L Macro IS
finally imaging to a small ASP-C sensor set to 100 ISO

The result below:
Image
121 focus slices were captured. Focus controlled remotely vía Canon's EOS utility software, in increments of one mouse click on the >> button in the GUI of software. Light was provided by the camera's internal flash: a small mirror was placed at aprox 45º to direct light to specimen, and the interior of a smal styrofoam cup used as a reflector.

Focus stacking process P-max with zerene stacker program. I think spent too much time attempting to ajust the color: I completely forgot to adjust the white balance of the camera. But no sharpening, or focus mask was used.

detail of center
Image

Then a close up of the "head" at approximate 5x. The optical train:
Nikon CFi 10x Plan Achromat
RAF Camera M25x0.75 to M67x0.75 thread adapter
UV Slim filter (approx' 3.3mm)
Canon 100mm f/2.8L Macro IS
finally imaging to a small ASP-C sensor set to 100 ISO

The result below:
Image
208 focus slices were captured. Focus controlled remotely vía Canon's EOS utility software, in increments of one mouse click on the >> button in the GUI of software. Again, light was provided by the camera's internal electronic flash.

Again, focus stacking process P-max with zerene stacker program. Color ajustments, but no shapening, or focus mask was used.

detail of "face"
Image

improvised arrangement of mirror and styrofoam cup reflector
Image

I noticed that ilumination by electronic flash, as opposed to long exposures by continuos light, correlates to a slightly sharper image, and also less vignetting.
But given remote control of focus vía EOS Utility, the live view remains activated, which seems to gradually heat the sensor. A hot sensor seems to correlate to more noise.

QUESTION 1
What causes the star like artifacts? I have noticed a negative correlation of focus close to infinity. Also, they appear to be more prominent toward periphery of image. IIf these are the causes of the star like artifacts, then it suggests that the remote control method of focus vía EOS Utility software is more adequate for relatively flat objects.

QUESTION 2
What is your prefered method to inmovilize this type of specimen, yet maintain a life like appearance? that is given a preference for the specimen to appear more alive, and given the necessity of an inmobilized specimen for focus stacking process.

QUESTION 3
If you can you positively identify this specimen, what is it? And more importantantly, how do you know?

Comments, questions, and considered opinions are greatly appreciated!
Last edited by Carmen on Wed Jun 03, 2015 10:56 am, edited 1 time in total.

ChrisR
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Post by ChrisR »

Q1 Look at the individual frames, and whether they appear on a Dmap stack. If you have sharp-edged artefacts in the individual frames, Pmax will collet them up and show you something like a star. If you have Coma, then focusing through the detail will produce a cross. This may be more likely towards the edge of the frame particularly when usuing in-lens focusing.
Try retouching from Dmap, which will only use one or two frames, or from an aligned source image.

Q2: My knowledge is limited to using the freezer, or ethyl acetate. Either way the legs and antennae will curl, and need coaxing into position, possibly after "relaxing" in a damp environment, later.

Q3: No knowledge! Good Luck!. In the UK we have limited native species but more recently some new arrivals. Many of them seem to be "variable" enough to confuse me completely.

NB I would try removing the filter. You're introducing more surfaces to give problems or reduced contrast from reflections. You shouldn't have a UV issue :).

You're getting quite a noticeable "halo" around the beetle.
You might like to try an adjustable iris, placed immediately behind the objective, for exposing the background. At a smaller aperture, you would lose definition, but that wouldn't matter for the background. The cone of light between the subject and the lens though, would be much narrower, so the beetle wouldn't be in the way of it as much. That would give you something to retouch from, correspondingly closer to the animal.
You can, interestingly, see how the halo width changes in your result, according to the angle of the specimen and hence the distance between it and the background. Where the distance is greater (head end) the halo is much more obvious, because more of the cone of light is obscured.

rjlittlefield
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Re: 1st ladybird

Post by rjlittlefield »

Carmen wrote:Color ajustments, but no shapening, or focus mask was used.
This is a good way of showing how well the optics are doing.

But to make the best picture, some sharpening will be required. This is because the image has been softened by diffraction, so that fine detail has lost contrast compared to coarser detail. Digital sharpening can largely correct that difference, making the image be more like what it would be in the imaginary case without diffraction.
improvised arrangement of mirror and styrofoam cup reflector
This is very clever, especially the use of the paper tab to shadow the subject from direct illumination by the flash. Thank you for showing this setup.
I noticed that ilumination by electronic flash, as opposed to long exposures by continuos light, correlates to a slightly sharper image, and also less vignetting.
The part about sharper is easy to understand. That's a matter of the short flash exposure freezing out any vibration.

The part about less vignetting is strange. I cannot think of any reason why you would get less vignetting with flash, assuming that the same aperture was used in both cases.
A hot sensor seems to correlate to more noise.
Definitely. This is one of the tradeoffs in using EFSC. To get the least pixel noise, you would have to shoot in normal mode, not live view, and then flash illumination at your camera's lowest ISO setting would be the best.
What causes the star like artifacts? I have noticed a negative correlation of focus close to infinity. Also, they appear to be more prominent toward periphery of image. IIf these are the causes of the star like artifacts, then it suggests that the remote control method of focus vía EOS Utility software is more adequate for relatively flat objects.
I agree with ChrisR. The fact that these are more prominent away from center strongly suggests that they are due to some sort of aberration. That aberration probably arises in the objective, and it will be worse away from the objective's design point for focus. It will also be worse at higher magnifications, because of the higher NA. Focus-stepping in the tube lens definitely does impose a limit on stack depth. In my one test with a 50X NA 0.55 objective using AF-motor focusing, I was only able to stack about 40 frames before the image quality degraded too much to be acceptable. See the discussion and images at http://www.photomacrography.net/forum/v ... 2144#92144.
What is your prefered method to inmovilize this type of specimen, yet maintain a life like appearance? that is given a preference for the specimen to appear more alive, and given the necessity of an inmobilized specimen for focus stacking process.
Most beetles and some other hard-bodied subjects retain a life-like appearance even when dry. For these subjects, you can arrange the specimen into a lifelike position with small pins while it is fresh, let it dry, then remove the pins to photograph it. That's the way the tiny carpet beetle at http://www.photomacrography1.net/forum/ ... php?t=4812 was prepared.

Soft-bodied specimens are a different matter entirely. For those, the best strategy I've found is to photograph them while they're still fresh. Flies can be rehydrated to some extent, but they never look quite the same as when they're live or very recently deceased.

--Rik

Carmen
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Post by Carmen »

Thank you ChrisR for you insight and kind advice.
ChrisR wrote:Q1 Look at the individual frames, and whether they appear on a Dmap stack. If you have sharp-edged artefacts in the individual frames, Pmax will collet them up and show you something like a star. If you have Coma, then focusing through the detail will produce a cross. This may be more likely towards the edge of the frame particularly when usuing in-lens focusing.
Try retouching from Dmap, which will only use one or two frames, or from an aligned source image.
I examined the D-map stacked image, and also the individual captures of focus slices. They appear to be reflections of the flash, possibly from the polished surface of the microscope objective. They remind me of reflections of the sun in a tiny droplet of water. I retouched the Pmax output with the slightly improved bits from Dmax output. See below:
Image

If they are from reflections of the flash in the polished surface of the microscope objective, then I suspect that probably a sort of mini black lens Hood may serve to minimize them.
ChrisR wrote:You're getting quite a noticeable "halo" around the beetle.
You might like to try an adjustable iris, placed immediately behind the objective, for exposing the background.
Wouldn't a sort of mechanical iris between the microscope objective and the tube lens mask off the perifery? ¿wouldn't that cause some vignetting? What sort of results would you reasonably expect? Has anyone confected such an iris from an old filter ring, perhaps with black cloth or bits of black paper? It sounds intriguing.

Can anyone think of any other plausible causes of the "halo" around the beetle which to investigate? Dust in optical train? software? etc...

CrisR, to be honest, the reason for the UV filter is partly because I was lazy, and partly because I was lead to think, from initial tests, that the extra 3.3mm space between micro' objective and tube lens slightly improved image quality. The RAF adapter per se positions the micro' objective very close to tube lens Surface. The filter is expensive multi coated type, but will try without it.

Carmen
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Re: 1st ladybird

Post by Carmen »

rjlittlefield wrote:The part about less vignetting is strange. I cannot think of any reason why you would get less vignetting with flash, assuming that the same aperture was used in both cases.
Rik, one possible explanation ocurres to me. When I used film, I often needed to compensate long exposures with an extra aperature or more, due to the poor reaction of the film at these extremes. If the same thing occurs with long exposures on a sensor, and the sensor is on the on the threshold (i.e. increasingly less responsive at such marginal conditions), then a vignette of perhaps 1 aperature may appear to be amplified to 2 aperature because it is on the threshold so to speak. Bear in mind, this is only a conjecture. At any rate, the vignette appears less noticeable at faster shutter speeds than at 5, 10, 20+ second exposures.

Regarding the star like artifacts:
rjlittlefield wrote: I agree with ChrisR. The fact that these are more prominent away from center strongly suggests that they are due to some sort of aberration. That aberration probably arises in the objective, and it will be worse away from the objective's design point for focus. It will also be worse at higher magnifications, because of the higher NA. Focus-stepping in the tube lens definitely does impose a limit on stack depth. In my one test with a 50X NA 0.55 objective using AF-motor focusing, I was only able to stack about 40 frames before the image quality degraded too much to be acceptable. See the discussion and images at http://www.photomacrography.net/forum/v ... 2144#92144.
Rik, Thnx again for the pertinent thread. And your considered opinion and test seem to confirm my suspitions. In consequence, I think it best to restrain this provisional internal tube lens focus method: perhaps 50 focus slices max', rather than 200+ slices, at least with this nikon 10x lens.

Rik, thnx for the considered suggestions on photographing insects. I have some specimens in the freezer from last summer, and fear they've suffered freezer burn. But as they say, (roughly translated) every experience serves to improve future attempts.

One question please!
Have you, or anyone else, tried this same Nikon CFi 10x Plan Achromat microscope objective with the Canon 200mm f2.8 as a tube lens, and projecting to an APS-C (18 or 20mp) or relevantly similar sensor? I am keen to see how much detail it yields at that magnification!

rjlittlefield
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Re: 1st ladybird

Post by rjlittlefield »

Carmen wrote:Have you, or anyone else, tried this same Nikon CFi 10x Plan Achromat microscope objective with the Canon 200mm f2.8 as a tube lens, and projecting to an APS-C (18 or 20mp) or relevantly similar sensor? I am keen to see how much detail it yields at that magnification!
I do not have that exact equipment to test.

However, I can confidently predict that, compared to what you're getting now, the images at 10X (on 200 mm tube lens) will show more detail on subject but will look slightly less sharp in the image. See http://www.photomacrography.net/forum/v ... 082#101082 for discussion. Working from images shown and linked there, here are corresponding areas of a sandpaper target photographed at 10X on sensor and 5X on sensor (using 15 megapixel APS-C).

Image
At any rate, the vignette appears less noticeable at faster shutter speeds than at 5, 10, 20+ second exposures.
Very interesting. I will put this on the list of things to explore at some point.

--Rik

Carmen
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Re: 1st ladybird

Post by Carmen »

rjlittlefield wrote: ...I can confidently predict that, compared to what you're getting now, the images at 10X (on 200 mm tube lens) will show more detail on subject but will look slightly less sharp in the image. See http://www.photomacrography.net/forum/v ... 082#101082 for discussion. --Rik
thnx again Rik for the pertinent link! Your considered prediction aligns with what I expect.

I expect to have a full frame camera toward the end of this year, or beginning of next year. I was considering the 200mm f2.8 as a tube lens. It's a respectable lens, and the full frame sensor resolves fewer lines per mm, so it would seem to be adecuate for the purpose. Perhaps even moreso than the thorlabs tube lenses.

rjlittlefield
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Re: 1st ladybird

Post by rjlittlefield »

Carmen wrote:I expect to have a full frame camera toward the end of this year, or beginning of next year. I was considering the 200mm f2.8 as a tube lens. It's a respectable lens, and the full frame sensor resolves fewer lines per mm, so it would seem to be adecuate for the purpose.
That should work fine. I cannot recall anyone ever reporting problems with a 200 mm f/2.8 fixed-length lens.

--Rik

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