house ant, take 7, new diffuser set up... comments welcomed!

[ctron]

Ctron,

With a single flash an a polystyrene cup, you have one of the best setups for studio lighting insect that you could ask for. However, you want to do essentially the opposite of what you're doing here: Instead of shining the light through the cup, shine the light into the cup and let it bounce around inside. So use no diffusion at all--just let the light bounce from a wide range of directions--and you should obtain nice, even light. Bounced light can be much easier than diffused light, and can produce better results.

• To do this:
  
  1) Cut a lengthwise portion out of the edge wall of the cup, and shine your flash into it.

I assume you mean like the attached image.



....and when the cup is lifted, we see the specimen held by toothpick and clay

2) Importantly, block any portion of the flash from shining on the ant directly. Only light bouncing off the inner surface of the cup should hit the ant. You can use part of the polystyrene you removed to make the light blocking "flag."

With the angle of the flash, I don't think any of it was hitting the specimen directly, but I'm not sure. Do you see anywhere in the image where a flag may need to be placed?

3) Since you'll no longer be shining your light through the polystyrene, you'll lose far less of it, so you can probably set your flash for at a lower power, which will improve recharge time and save batteries.[/list]

Yes, I was able to reduce by a couple of fractions/ stops.

I would normally suggest shining the light from underneath the subject, but your subject support is already taking up that space. So you can put the open portion of the cup, and your flash, above the ant instead. But if you do this, photograph the ant upside down and rotate the final image 180 degrees. As you likely know, our eyes/brains treat the direction the light is coming from as "visual up," since we evolved in a world where the sun was above us. In this lighting arrangement, the side of the ant nearest the flash is actually the least lit, as the flash is blocked in that direction--so visual up is opposite the flash.

With careful placement today, I was able to turn the ant upside down and keep it in position.

One problem I keep running into is when to stop taking images for the stack. I keep stopping a bit too soon and this makes the extreme end portion of the image blurry. I suppose I might be able to mark start/ stop areas on the course focus knob on the Nikon block(?).

I have a slice of masking tape on the fine focus wheel of my focus block, which helps me count revolutions.

Also, I try to shoot "through" the subject, including some extra shots to front and rear. Most of the time, I throw these away. But sometimes they save me from having to reshoot when I miss on one end or the other.

Further, some members of our community mount an iris as close as possible to the rear of the objective. Then at the ends of the stack, they stop down for a few pictures (which may be more widely spaced). The stopped down images have greater depth of field, though less resolution, and can provide a more pleasing transition from in-focus to out-of-focus elements.

I can't really go by the back of the camera histogram for stacking. If I try to keep highlights where they should be, the final stack is too dark. Seems like I have to almost clip the highlights, but the stack doesn't turn out overexposed. Not sure what's going on there.

If you're using the live-view histogram, don't trust it overmuch, as in my experience they aren't all that accurate. If you're shooting an image and then checking the histogram, this can be much more accurate. Make sure you are using an RCG histogram, and not single-channel or luminosity histogram. Also, at magnifications like these, small highlights go in and out of focus shot by shot. When such a highlight is out of focus, the bright bits are averaged with less-bright bits, so OOF highlights appear darker than they will be when in focus. I make it a point to set my exposure with an in-focus frame, and test it on a few frames with different points of focus.

Not sure if any of the above answered your questions?

Cheers,

--Chris

I think it did help, yes, but I'm still running into difficulties. I did replace the cup I was using with the one above and inverted the ant, but I then was ending up with a very bright ant bottom and too dark of a topside. I also had to use Lightroom to reduce exposure by 2 stops as especially the bottom of the ant was way overexposed. If I try and adjust flash so that nothing is blown out, I notice that Zerene has the tendency to produce a final stack darker than the individual frames, which may have looked quite good during acquisition. I then have to try and increase the brightness of the darker result.

The flash lighting difficulties are still the main thing holding me back. I just can't seem to get uniformity, either too bright or too dim. If I get one part of a specimen just right, then the other part is blown out. I'm not using the camera instant histogram either.... I go back after the image is taken and view it. If it's where it should be the final stack ends up too dark.

I notice that some people make sub-stacks to be later combined to produce a composite. I'm not exactly sure of the reason. One could be that I notice Zerene has the tendency to change areas already stacked sometimes where maybe there would have been a better stack of just a particular area earlier on. It could also be for exposure reasons too. As I said, I reduced by 2 stops all frames in Lightroom and then stacked. I have another stack where nothing was changed, and I could probably reduce by 1 stop for another stack. Sort of like making an HDR except software changing exposure and not actually taking new images. I just hoped I could avoid the extra work of these methods.

I'm also not sure if an objective is suited to specimens filling only half frame. I don't see many ant whole bodies taken with objectives, usually only the heads. Rik kindly pointed out the odd positions dead ant bodies would be in and I agree, but maybe there's some flaring going on too with objects not filling the frame. I know I've pretty much tried everything and still ending up with speculars and or washed out detail areas.

Just some thoughts....

[dack9]

To do this:

1) Cut a lengthwise portion out of the edge wall of the cup, and shine your flash into it.

2) Importantly, block any portion of the flash from shining on the ant directly. Only light bouncing off the inner surface of the cup should hit the ant. You can use part of the polystyrene you removed to make the light blocking "flag."

3) Since you'll no longer be shining your light through the polystyrene, you'll lose far less of it, so you can probably set your flash for at a lower power, which will improve recharge time and save batteries.[/list]I would normally suggest shining the light from underneath the subject, but your subject support is already taking up that space. So you can put the open portion of the cup, and your flash, above the ant instead. But if you do this, photograph the ant upside down and rotate the final image 180 degrees. As you likely know, our eyes/brains treat the direction the light is coming from as "visual up," since we evolved in a world where the sun was above us. In this lighting arrangement, the side of the ant nearest the flash is actually the least lit, as the flash is blocked in that direction--so visual up is opposite the flash.

--Chris

Dear Chris,

If it is possible, could you add photo or drawing of this light scheme? Because for me, using only you description it is hard to repeate the scheme... )

[ChrisR]

I notice that Zerene has the tendency to produce a final stack darker than the individual frames, which may have looked quite good during acquisition.

Alarm bells here.
This is only a behaviour I recognise where you have one dark frame in the stack, sometimes the first, which is much darker than the rest. This upsets the auto-light-balancing in the stacker. Check carefully that you don't have an odd rogue frame.
It can make a real mess!
You could try rerunning the stack with the (Preferences) Brightness box unchecked.

[ctron]

I notice that Zerene has the tendency to produce a final stack darker than the individual frames, which may have looked quite good during acquisition.

Alarm bells here.
This is only a behaviour I recognise where you have one dark frame in the stack, sometimes the first, which is much darker than the rest. This upsets the auto-light-balancing in the stacker. Check carefully that you don't have an odd rogue frame.
It can make a real mess!
You could try rerunning the stack with the (Preferences) Brightness box unchecked.

Well, as my flash warmed up at the higher powers, sometimes it would miss and I did go back and delete those frames. Now I think the result is darker because the light from the flash is varying as the specimen is moved. One possible solution I can think of it applying a shadow/highlight function like Photoshop has to equalize brightness which might greatly help with a uniform result. I haven't tried this yet as it will take even more time than Lightroom. I will try unchecking brightness first,might save a lot of work. Thanks for that tip.

[ChrisR]

Another low grunt trial could be a stack of just a few nice frames, to see if it's different.

I have that objective, and a cup - no ants here though.

[ctron]

Another low grunt trial could be a stack of just a few nice frames, to see if it's different.

I have that objective, and a cup - no ants here though.

Yes, I definitely see differences although I don't know if I'm going to go with substacking. Just comparing stacks from native exposure and 2 stops less results in a different sized subject which is next to impossible to combine unless very carefully resizing is done. I'm not sure what would happen if I stacked smaller numbers from the same session though.

My latest result, based on all the techniques discussed so far in this thread is here.

One thing I haven't asked about at all are camera settings. I tend to stick with 100 ISO in order to avoid noise, but I'm sure some folks are going higher. I'm not sure of the benefit though unless it is to reduce flash output, but there may be other unknown (to me) reasons. Also, any other camera settings to be aware of.

[naturepics43]

An observation from a novice. If I understand your set-up, you move the specimen toward the objective & your flash is stationary. I think the blob of clay is blocking a lot of light causing shadows to fall in different places as you advance through the stack since the flash doesn't move with it. Just a thought.

[ctron]

An observation from a novice. If I understand your set-up, you move the specimen toward the objective & your flash is stationary. I think the blob of clay is blocking a lot of light causing shadows to fall in different places as you advance through the stack since the flash doesn't move with it. Just a thought.

That's a very good observation, something I haven't thought about, and could very well be causing light variance. My flash set up, so far, has been on a separate, stationary tripod and does not move as the subject does.

As far as the clay goes, I found it convenient but it could be detrimental.

I welcome suggestions for options other than clay as well as what I could attach to the side of the Nikon scope to allow flash use. Willing to try changing options for both of these factors. I'm been looking for some sort of flash bracket anyway, especially something DIY. However, attaching this to just the moving part of the scope could be a challenge.

[naturepics43]

The clay is OK, just try to flatten it out so it is below the height of the ant. Could you replace the switch plate platform for a wider platform? Then you could set the flash unit on the platform. The mis-firing flash you mentioned may be due to the cameras built in flash circuit getting hot & shutting off until it cools. I had that problem with my Nikon D 7000. I now use a cord from the camera hot shoe to one of the flash units. No more mis-firing.

[rjlittlefield]

This section of the Zerene Stacker FAQs may be helpful:

My colors changed a little. Why is that?

There are three reasons that output images can have different colors from the input: 1) brightness adjustment, 2) PMax, and 3) “Retain extended dynamic range” when saving. “Brightness adjustment” refers to Zerene Stacker's attempt to correct for uneven exposure between various input images. That feature is turned on by default, but you can turn it off by un-checking Brightness at Options > Preferences > Alignment. “PMax” refers to the PMax stacking method, which often makes slight changes in brightness, contrast, and saturation as a side effect of doing its focus stacking. This behavior is an unavoidable side effect of PMax and should be considered as one of the tradeoffs of PMax versus DMap. “Retain extended dynamic range” when saving causes the range of internal pixel values to be compressed if necessary to fit within the 0-255 range of image files. Internally the range can exceed 0-255 as a result of PMax, brightness adjustment, or even just pixel interpolation during alignment.

Color/brightness/contrast changes can be completely avoided by using the DMap stacking method, with Brightness adjustment turned off at Options > Preferences > Alignment, and “Retain extended dynamic range” turned off at Options > Preferences > Image Saving or in the file save dialog.

More information is provided by this one:

What does "Retain extended dynamic range" mean?

As background, you need to know that the PMax stacking method often causes contrast to increase, pushing darks darker and brights brighter. If your source images are already high contrast, then the increase can internally push pixel values to “darker than black” or “brighter than white”. Such values cannot be saved in ordinary image files. By default, Zerene Stacker clips these pixels to exactly black or white when the file is saved, thus throwing away some information you might like to keep. Placing a checkmark on “Retain extended dynamic range” essentially does a “levels adjustment” that reduces contrast and possibly brightens the image, exactly enough to occupy the full range of allowed pixel values, 0-255 in an 8-bit image. This preserves all the computed pixel values so that you can apply your own levels or curves adjustment in Photoshop or any similar tool to get whatever appearance you like best. When using “Retain extended dynamic range”, it's also a good idea to use 16-bit TIFF output, so as to preserve good gradation that might be lost if the extra dynamic range were compressed into 8 bits.

Note that in previous versions of Zerene Stacker, this option was named “Retain full dynamic range”. It has been renamed to more accurately reflect its function.

One way to help evaluate your stack would be to turn off Options > Preferences > Alignment > Brightness, stack using DMap, and look at the result. If you see banding or blotches in the OOF background, then you're having trouble with flash variation from frame to frame. If the lighting of the subject itself looks OK with DMap but not with PMax, then apparently your difficulties are introduced by PMax. In that case you might want to make friends with DMap and with the hybrid DMap+PMax approach using retouching. This is covered by the first few tutorials at http://zerenesystems.com/cms/stacker/do ... rialsindex.

--Rik

[ctron]

One way to help evaluate your stack would be to turn off Options > Preferences > Alignment > Brightness, stack using DMap, and look at the result. If you see banding or blotches in the OOF background, then you're having trouble with flash variation from frame to frame. If the lighting of the subject itself looks OK with DMap but not with PMax, then apparently your difficulties are introduced by PMax. In that case you might want to make friends with DMap and with the hybrid DMap+PMax approach using retouching. This is covered by the first few tutorials at http://zerenesystems.com/cms/stacker/do ... rialsindex.

--Rik

Attached is the DMap result, brightness off, no editing other than cropping for the forum.



This tells me that I'm still having trouble with flash illuminating the darker areas. Anything beyond that, not sure.

Honestly, I'm about out of options with flash, not sure what else I can do. As a last resort, I may try a polystyrene chip cone diffuser with two flashes on opposite sides, but to do so will mean major changes in my set up. Not sure it's worth the trouble. I've tried to keep the set up as simple and as cheap as possible before settling on a permanent set up that works, which, to date, I have not yet found. Not even close.

I have to ask, is anyone using a lens hood on their objective? Doesn't look like it from archive research. I think this would be difficult to implement since we're already very close to the subject.

Two other options I'm looking at are going back to constant lighting, which I seemed to have more luck with, and using one of the recommended copy/ enlarger lenses on bellows. I have 10 W constant LED lights on the way, but won't shell out for the latter unless there's no way I can get the lighting right with the objective.

[ChrisR]

No time now, but move the DMap sliders to minimize the "cloudy" edges, then retouch them from PMax.

You're getting a lot of light from the backround, reflecting off the ant. I think it would work better with a duller background.
For such deep stacks, substacks do help too.

[Lou Jost]

I do use a lens shade made of Protostar with my microscope objective.

As people have pointed out, it is essential to move the light with the subject (or move the camera and leave light and subject stationary).

I've never had the problem you describe regarding exposure differences between individual frames and Zerene stacked results. There is an increase in contrast in PMax output relative to individual frames, but not an overall shift in exposure.

[rjlittlefield]

Those are quite some impressive loss-of-detail halos. To improve the DMap result, I suggest pushing the DMap contrast threshold slider a bit lower, so that essentially the whole ant retains natural colors while the background is left to go "black in preview". See the "How To Use DMap" tutorial at http://zerenesystems.com/cms/stacker/do ... utorial003 for some explanation.

Given your problems adequately illuminating the dark ant, I suggest using a darker background to reduce the subject/background contrast. When setting up the background, take note that your NA 0.25 objective is accepting a cone of light that's about 30 degrees wide. You'll need to cover an area at least that wide with the darker background, or the background will end up lighter than you think it ought to be.

I would be interested to see your entire frame, not cropped. I gather that you have a dark subject floating in a sea of brightness, but I don't know how big the sea is.

There is an increase in contrast in PMax output relative to individual frames, but not an overall shift in exposure.

That's true in general, but when the subject is small and dark, it will get a lot darker while the background gets only a little lighter.

About the lens hood, I personally would not spend the effort to try using one with this sort of objective. (Truth be told, I hardly ever use a lens hood with any objective, even LWD models.)

--Rik

[ctron]

Alright, latest stack. This time based on Chris R's suggestion to try using sub-stacks. There were 5 sub-stacks used here.