First attempt with Nikon 10x objective

[lauriek]

Mounted on OM auto bellows with PM-MTOB OM-RMS mount, bellows at maximum extension. DOF sure is thin with this setup!!

Stack of 23 images, obviously should have shot more images but I'm just getting to grips with the new lens!

Subject is a forget-me-not flower, entire flower approx 3-4mm across, I suspect we're looking at a horizontal field of view of approx 1mm here...



and a 100% pixel crop of an in focus section:



Overall I know the stack is not deep enough but I'm quite pleased with the results. This is straight out of combine ZM, I've been playing with the settings a bit in this latest version and found the best results for this particular stack with a modified "do stack" macro with "find detail" settings modified to either 1,1,0 or 3,1,0 (I wish CZM used some of the settings used on the stack in the output filename in the same way as HF!!)

Does anyone know exactly does this first number means, as I've struggled to understand the help on this - I actually thought a larger number might work better on this but it did not, I kept playing with the numbers and this was by far the best result - although it did miss some detail on the bottom right of the image which is there in some of the originals.. (Also any CZM experts, what does the number [default 5] mean in the "remove islands" function?)

As with any of my image posts, comments are appreciated...

Cheers!

ETA Thanks to Rik for bringing this lens to my attention, and Charlie for pointing out someone with some for sale at a reasonable price!!

[rjlittlefield]

Laurie,

Welcome to the wonderful world of high-mag stacking! I hope you are encouraged by these results.

Regarding the "remove islands" function, John Hollenberg's review article says that "Islands made up of less pixels than specified in the parameter are removed. Thus, for Remove Islands (5), islands with 4 pixels or less will be removed."

About the three-argument "find detail" command, I can't offer any insight and I'm a bit skeptical that the thing is working correctly. I know there's a bug prowling around there someplace but I'm not sure what triggers it. What I do know is that someplace between the Feb 3rd and Mar 15th versions, Do Stack stopped correctly handling my hobo-face stack. That corresponds to the time when "find detail" switched from one argument to three. I just discovered this problem a couple of days ago and it's still in Alan's queue of stuff to look at.

As a general comment about your image, the actual-pixels shot suggests that you're giving away some field size in exchange for only empty magnification. I suggest shooting the same highly detailed subject at a range of magnifications, with the goal of finding the smallest magnification that captures all the detail the lens provides. I'm guessing that'll happen at a lot less than full bellows extension.

--Rik

[Charles Krebs]

Laurie,

I don't have access to the specs where I am right now, but if you "maxed-out" an Olympus bellows (about 190mm or so) and then add in the camera body "flange-to sensor" distance (probably around 40mm or so) you were working at a magnification a little greater than 15X!

We warned you about the DOF

After awhile you begin to realize what will, and will not, work well (stacking) at these magnifications. In a "normal", single macro shot we are accustomed to seeing the DOF fall off gradually behind, and in front of the part of the subject focused upon. With stacking at 10X and higher, there is no gradual DOF fall off... it goes completely out of focus as soon as you stop taking the stack. Often you'll get the section you are most interested in, but if there are "deeper" parts in the frame the rapid drop to "completely out-of-focus" gives a look we are not used to seeing, and can look a bit odd at times. This is not always an issue, but if you want to avoid it on certain shots it will cause you to carefully choose your compositions. It's not uncommon that you will get about 90% of what you want in, say, 30 or 40 images... but then to "pull in" that last 10% you need to double or triple the stack. Careful choice of subject and composition makes life much easier.

[acerola]

Is there any way to introduce an "aperture" to the optical path for this type of setup? In this way you could achieve an image with more DOF. I know it would be heavily affected by diffraction. But this picture could have been used to masking out the out of focus ranges of the stack, where people did not expect good resolution.
In this way a high percentage of the frames (Charlie mentioned above) could be spared.

[rjlittlefield]

Is there any way to introduce an "aperture" to the optical path for this type of setup? In this way you could achieve an image with more DOF. I know it would be heavily affected by diffraction. But this picture could have been used to masking out the out of focus ranges of the stack, where people did not expect good resolution.
In this way a high percentage of the frames (Charlie mentioned above) could be spared.

In theory, yes. In practice, I've tried it only a couple of times and ran into enough problems that it was not worth the trouble.

The theory applies to ordinary lenses with diaphragms also, and it's easier to talk about those.

Suppose you have a short f/2.0 lens that you are running at high magnification. Choose a focus step that is just right for f/2.0, call it X. Cover the area that you want to be sharp with numerous frames at f/2.0, step X. At the end(s) of the sequence, shoot several more frames like this: f/4, step 2X; f/8, step 4X, f/16, step 8X, in each case adjusting the exposure time to compensate for the stopping down. Then just stack all the images.

Of course you can use intermediate f-stops also, and/or shoot multiple frames at the smaller stops.

When I tried this, the major difficulty I ran into was matching the exposures at various f/stops. Again in theory, the exposure at f/16 should be exactly 64 times the exposure at f/2. (64 = 16/2, squared.) But in practice, I was having to shoot the f/2 frames at 1 second for vibration reasons. So the f/16 frames would have been at 64 seconds, where sensor noise and hot pixels became serious problems. Worse, the proper f/16 exposure was not exactly 64 seconds but some different time that was tedious to determine. This was with continuous illumination. I think similar problems would have cropped up with flash.

With a microscope objective, it is possible but difficult to introduce an adjustable aperture, leading to another impediment. As a heroic effort, one might shoot the high res part of the stack with a microscope objective, then switch to a macro lens with a smaller and adjustable aperture to shoot the rest of the stack. Of course this would introduce magnification differences in the camera images. Those differences could be handled with state-of-the-art image registration tools such as PTGui, but again it's a lot of trouble.

I don't mean to sound too negative here. The idea has merit, and quite possibly a workflow could be developed that would be reliable and fast enough to make it worth the trouble for some images. It's just not what you'd call "easy".

--Rik

[lauriek]

Rik wrote:
I hope you are encouraged by these results.

Considering this was just a first play I'm very encouraged. Way to go yet to work out optimal setup for this lens but very pleased so far - it is hard work but for some reason I don't mind hard work like this!!

Thanks for the help on the numbers in CZM, encouraging to know they don't work as expected and it wasn't just me!!

Rik wrote:
As a general comment about your image, the actual-pixels shot suggests that you're giving away some field size in exchange for only empty magnification. I suggest shooting the same highly detailed subject at a range of magnifications, with the goal of finding the smallest magnification that captures all the detail the lens provides. I'm guessing that'll happen at a lot less than full bellows extension.

I presume the empty magnification you're talking about is because I'm running /this/ lens at higher magnification than it's specified for - ie according to Charlie I'm at a little over 15x which ties up well with my estimate of field size - and it's a 10x objective (So it's resolution is likely to be designed for 10x?) . You're not saying that I can't use a higher magnification objective on this sensor?

Charlie wrote:
We warned you about the DOF

You did Charlie!

As I mentioned above I don't mind this kind of hard work for some reason, I was just surprised at the thinness of the slice of 'in focus' through the viewfinder (well on my live-view screen) - it's miniscule!!

Thanks for the tips!

[lauriek]

Is there any way to introduce an "aperture" to the optical path for this type of setup? In this way you could achieve an image with more DOF. I know it would be heavily affected by diffraction. But this picture could have been used to masking out the out of focus ranges of the stack, where people did not expect good resolution.
In this way a high percentage of the frames (Charlie mentioned above) could be spared.

I was wondering about this myself last night - with a 3d subject in front of this lens, I noticed that you can see objects in focus, which are behind other objects, which if the latter are in focus block the former - this leads me to believe you could even mount an aperture /in front/ of the lens itself.

However I think a part of the reason this lens is so sharp is its fast aperture - compared to most macro lenses approaching this magnification anyway - stop it down in some fashion and I think as you say diffraction would soon become bad!

Rik wrote:
When I tried this, the major difficulty I ran into was matching the exposures at various f/stops. Again in theory, the exposure at f/16 should be exactly 64 times the exposure at f/2. (64 = 16/2, squared.) But in practice, I was having to shoot the f/2 frames at 1 second for vibration reasons. So the f/16 frames would have been at 64 seconds, where sensor noise and hot pixels became serious problems. Worse, the proper f/16 exposure was not exactly 64 seconds but some different time that was tedious to determine. This was with continuous illumination. I think similar problems would have cropped up with flash.

Not relevent to Acerola's question but - A while back I worked out this idea - at the beginning and end of each stack I shoot 1 shot at something like f16, then shoot the main stack at a better f-stop for sharpness - ie say f5.6 with a reversed 50mm. Then at the other end of the stack I shoot another shot at f16. I didn't have too much trouble matching the exposures - but I use flash - I just upped the flash power by 1 stop for each stop I change on the lens and this seemed to work fine. I think the problem you were suffering was something akin to reciprocity failure, which iirc comes from long exposures, not aperture.

[rjlittlefield]

Rik wrote:
As a general comment about your image, the actual-pixels shot suggests that you're giving away some field size in exchange for only empty magnification. I suggest shooting the same highly detailed subject at a range of magnifications, with the goal of finding the smallest magnification that captures all the detail the lens provides. I'm guessing that'll happen at a lot less than full bellows extension.

I presume the empty magnification you're talking about is because I'm running /this/ lens at higher magnification than it's specified for - ie according to Charlie I'm at a little over 15x which ties up well with my estimate of field size - and it's a 10x objective (So it's resolution is likely to be designed for 10x?) . You're not saying that I can't use a higher magnification objective on this sensor?

"Empty magnification" describes any time the sensor could capture significantly finer detail than is present in the image. Suppose the lens doesn't care what magnification it's used at. Even so, if you make the extension 5 times longer, so that you're getting 50X instead of 10X, you won't resolve 5 times finer detail because the aperture of the lens won't allow it. Depending on how small the pixels in your sensor are, you may capture more detail by increasing magnification above 10X, but at some point all you'll do is spread the same detail across more pixels -- that's empty magnification.

The literature suggests that with that lens (NA 0.3), it's marginally OK to add an additional 80 mm extension, giving 15X magnification. (See http://www.science-info.net/docs/etc/Tube-Length-na.gif) I haven't run any tests personally.

You can use a higher magnification objective with the same sensor, but it will come with a larger aperture as part of the design.

this leads me to believe you could even mount an aperture /in front/ of the lens itself

That might work. It depends on the lens design. I haven't tested with the Nikon CF.

I think the problem you were suffering was something akin to reciprocity failure, which iirc comes from long exposures, not aperture.

The usual story is that digital cameras don't have reciprocity failure, although they do accumulate noise. I'd guess that's not exactly correct (electrons do leak), but it's probably close enough to not matter. I did check the f/2 versus f/16 exposures at shorter times as well, and for that lens, they just weren't exactly 1:64. That's no surprise. If the actual diameter of the iris happens to be off by 10%, the transmission goes off by 20%, and it's easy for a 2 mm aperture to be off by 0.2 mm (10%). Another complication could have been that I was trying to cover a 6-stop range starting at wide open, versus a 3-stop range starting from stopped down. Hard to know, and I didn't investigate any deeper.

It's good to hear that somebody else had more success with the technique.

--Rik

[acerola]

My original idea is just making one frame at the end of stack with bigger DOF. If I make a short stack sequence, then I get this milky appearance for the out of focus areas. If I make this bigger DOF image. I did not care much about exposure, diffraction and noise. Because if the exposure is just half right I can make a suitable picture from RAW. All I need is a more suitable (for the sharp stack) out of focus area. Its bound to blurred so the diffraction is not a big problem and you can heavily noise filter the noisy image too. You can use Photoshop to mask the few critical areas from this picture to the stacked image by hand.

My only problem for trying this is to where to put the aperture. So one of the solution could be in front of the objective.

[rjlittlefield]

Yes, placing a new aperture in front of the objective looks promising.

I checked by using a piece of aluminum foil with a pinhole in it, and visually observing through a microscope.

The effect of the pinhole was to grossly degrade the image resolution while greatly increasing the DOF, with no trace of vignetting.

The added aperture also changed the center of perspective of the lens, an effect that is explained here. I doubt that this would be important for stacking applications, but it's a good thing to be aware of just in case you see bizarre effects. The most obvious effect in my tests was that when the pinhole was off-center, then focusing straight up and down caused the image to shift left and right, as if I were looking through one eyepiece of a stereo microscope. If you're careful to get the added aperture centered, this won't happen.

--Rik

[Charles Krebs]

Not a bad idea really... take a really "stopped-down" shot to incorporate somehow for the foreground or background "out-of-focus" areas in oder to get a more natural DOF "fall-off". It would not matter if it were mushy from diffraction. It would be interesting to try with a lens that has an adjustable aperture. I suspect it might work better with magnifications lower than we're talking about here.

For reasons I won't go into here, a reduced aperture can be helpful with some objectives in microscope darkfield illumination. Some objectives were provided with "funnel stops" that inserted into the rear of the objective, to effectively reduce the aperture. These seem to have been more common in the past. I've never seen one, and rarely see them mentioned.

When you look at the "numbers" it seems clear that the NA of these objectives was determined with resolution in mind. If a method were arrived at to "stop them down", the resolution "hit" would be very noticeable. I do have several objectives with diaphragms, and if they are accidentally moved off of full aperture, the resolution drop is clearly seen.

[acerola]

I have used this method for my ant stacking picture. I made a frame stopped down to check the composition. And the stack background has very uneven lightning from frame to frame. I had to use ETTL flash settings because my bigger flash (only it has manual settings) was at repair. I needed an easy solution for the background and it was the stopped down picture. I was glad later on as it solved the DOF transition as well.
I will try the added aperture when I have time. I also heard about an add on (rear) diaphragm on a Hungarian microscopy forum. (I'm not in microscopy yet, i just checked in before I bought the objective ). It would be nice to try some out.