I am not able to get a very sharp image even with post adjustments.
can any one give me some advice? thanks in advance. Sam
Diatom in a CUP
Moderators: rjlittlefield, ChrisR, Chris S., Pau
Diatom in a CUP
DIC splanapo 40X, Canon 70d. 40 stacks.
I am not able to get a very sharp image even with post adjustments.
can any one give me some advice? thanks in advance. Sam
I am not able to get a very sharp image even with post adjustments.
can any one give me some advice? thanks in advance. Sam
-
Charles Krebs
- Posts: 5865
- Joined: Tue Aug 01, 2006 8:02 pm
- Location: Issaquah, WA USA
- Contact:
Sam,
A few things to consider....
What arrangement/optics do you have to "send" the image into the camera? Are you using an NFK photoeyepipece... and if so, what magnification?
Check your condenser aperture setting. If you close down the condenser aperture too much the image will quickly get "softer", even though contrast and DOF will increase. With that 40X in DIC you will likely find the "sharpest" image very close to (if not at) full aperture.
I have never used a 70D body. It does have EFSC, but I have not seen any information as to whether or not it has also has the small vibration that unfortunately showed up in the 60D. Were you shooting from live-view with one of the two silent modes turned on?
The 40/0.95 S Plan Apo (like all similar high/dry objectives) is very sensitive to cover glass thickness and subjects that are not positioned directly against the cover-glass (i.e. there is some distance filled with mounting medium between the subject and cover-glass). That's the reason for the adjustment collar, and it is very critical to have it adjusted correctly. And even with the adjustment collar you will encounter slides where the subject is too "deep" and you can't correct as well as you would like.
Lastly...
Even if everything is set up well, keep in mind that under the best conditions you will working at an "effective aperture" that is likely around f30 or smaller. So you are very much into diffraction territory, and resolution will suffer accordingly. If your frame of reference is a conventional (non-microscope) image, or even a microscope image from a lower power, high NA objective (like the 4, 10 or 20 S Plan Apos) then you will likely be somewhat disappointed.
But first consider the points mentioned at the beginning of this reply.
A few things to consider....
What arrangement/optics do you have to "send" the image into the camera? Are you using an NFK photoeyepipece... and if so, what magnification?
Check your condenser aperture setting. If you close down the condenser aperture too much the image will quickly get "softer", even though contrast and DOF will increase. With that 40X in DIC you will likely find the "sharpest" image very close to (if not at) full aperture.
I have never used a 70D body. It does have EFSC, but I have not seen any information as to whether or not it has also has the small vibration that unfortunately showed up in the 60D. Were you shooting from live-view with one of the two silent modes turned on?
The 40/0.95 S Plan Apo (like all similar high/dry objectives) is very sensitive to cover glass thickness and subjects that are not positioned directly against the cover-glass (i.e. there is some distance filled with mounting medium between the subject and cover-glass). That's the reason for the adjustment collar, and it is very critical to have it adjusted correctly. And even with the adjustment collar you will encounter slides where the subject is too "deep" and you can't correct as well as you would like.
Lastly...
Even if everything is set up well, keep in mind that under the best conditions you will working at an "effective aperture" that is likely around f30 or smaller. So you are very much into diffraction territory, and resolution will suffer accordingly. If your frame of reference is a conventional (non-microscope) image, or even a microscope image from a lower power, high NA objective (like the 4, 10 or 20 S Plan Apos) then you will likely be somewhat disappointed.
But first consider the points mentioned at the beginning of this reply.
optics used
hi Charles,
thanks for the reply, I used NFK 2.5 and the DIC intermediate has 1.25X magnification. I have a superwide trinocular head, but the view taken by the 70d is limited by the combination of 1.25X in the intermediate and the 2.5x of the NFK. I also have the mag changer attached (two attachments below the trinocular head) but it can set to 1X, so not contributing to change the size of the image. although it does takes away a little light when attached.
the condenser was set to 0.7-0.75, since splanapo was not designed for DIC usage, so I closed the aperture a little to reduce the glare. (this might be the problem? I should made it wide open at 0.9, that's the maximum on the condenser.)
for the 70d, I have the live view enabled and the silent mode set to 1. the 40 photos were taken raw then converted to TIFF, each was taken with shutter 1/25. (the vibration and the high power used may also be the problem)
the cover glass might not be the problem because I am seeing a crisp image via the eyepieces.
"Even if everything is set up well, keep in mind that under the best conditions you will working at an "effective aperture" that is likely around f30 or smaller. So you are very much into diffraction territory, and resolution will suffer accordingly. If your frame of reference is a conventional (non-microscope) image, or even a microscope image from a lower power, high NA objective (like the 4, 10 or 20 S Plan Apos) then you will likely be somewhat disappointed. "
I am a little confused by this,
does this formula work:
Effective Aperture = Lens Aperture x (1 + Magnification)
any adjustment I can do to reduce or prevent the diffraction for the high NA Objectives?
Charles, thanks again for replying to my questions. Sam
thanks for the reply, I used NFK 2.5 and the DIC intermediate has 1.25X magnification. I have a superwide trinocular head, but the view taken by the 70d is limited by the combination of 1.25X in the intermediate and the 2.5x of the NFK. I also have the mag changer attached (two attachments below the trinocular head) but it can set to 1X, so not contributing to change the size of the image. although it does takes away a little light when attached.
the condenser was set to 0.7-0.75, since splanapo was not designed for DIC usage, so I closed the aperture a little to reduce the glare. (this might be the problem? I should made it wide open at 0.9, that's the maximum on the condenser.)
for the 70d, I have the live view enabled and the silent mode set to 1. the 40 photos were taken raw then converted to TIFF, each was taken with shutter 1/25. (the vibration and the high power used may also be the problem)
the cover glass might not be the problem because I am seeing a crisp image via the eyepieces.
"Even if everything is set up well, keep in mind that under the best conditions you will working at an "effective aperture" that is likely around f30 or smaller. So you are very much into diffraction territory, and resolution will suffer accordingly. If your frame of reference is a conventional (non-microscope) image, or even a microscope image from a lower power, high NA objective (like the 4, 10 or 20 S Plan Apos) then you will likely be somewhat disappointed. "
I am a little confused by this,
does this formula work:
Effective Aperture = Lens Aperture x (1 + Magnification)
any adjustment I can do to reduce or prevent the diffraction for the high NA Objectives?
Charles, thanks again for replying to my questions. Sam
The 70D is not fully vibration free according with this test:
http://www.photomacrography.net/forum/v ... 406#145406
So to avoid any possible vibration blur chance, test it with expossure times about 2s.
1/25s is very prone to show shutter induced vibration if present
http://www.photomacrography.net/forum/v ... 406#145406
So to avoid any possible vibration blur chance, test it with expossure times about 2s.
1/25s is very prone to show shutter induced vibration if present
Pau
Vibration
Hi Paul.
I did read that post. But some other members also have confirmed 70d is indeed vibration free. I use the canon utility for tethering. I usually zoom in to check the pixels before each shot. And at pixel level the image is already blur. I will rearrange my setup again. And also do some test to check the vibration on 70d.
Thanks for the advice. Sam
I did read that post. But some other members also have confirmed 70d is indeed vibration free. I use the canon utility for tethering. I usually zoom in to check the pixels before each shot. And at pixel level the image is already blur. I will rearrange my setup again. And also do some test to check the vibration on 70d.
Thanks for the advice. Sam
-
Charles Krebs
- Posts: 5865
- Joined: Tue Aug 01, 2006 8:02 pm
- Location: Issaquah, WA USA
- Contact:
the condenser was set to 0.7-0.75
No not that one. For a microscope use m/2NA. You have an intermediate image (the actual image formed by the objective) of 50X, and the NA used is a little less than the full stated aperture on the objective (let's say about 0.825). So your effective aperture is roughly efff/30. But the primary point is not to come up with a hard "value" but to simply realize that you are unavoidably into "diffraction territory", so images taken at significantly larger effective apertures (if used for comparison) will almost certainly appear "sharper".does this formula work:
Effective Aperture = Lens Aperture x (1 + Magnification)
any adjustment I can do to reduce or prevent the diffraction for the high NA Objectives?
All the more reason to do a comparison with shutter speeds of 2-3 seconds or longer. This would at least determine whether vibration is a problem.the cover glass might not be the problem because I am seeing a crisp image via the eyepieces.
Also keep in mind that if you are viewing with super wide-field eyepieces you are seeing a field that is 26mm in diameter, but you are only recording a FN of 11mm with the 2.5X NFK into a Canon APS-C body. This will often tend to give an impression of a sharper image through the eyepieces.
If the longer shutter speeds don't improve things why not try a test comparison where you remove any unneeded optics . You need the DIC intermediate piece and the NFK, but try a shot without the mag changer. I would be surprised if that is causing you any problems... but you never know!
thanks for the tip
hi Charles,
thanks again for the advice, I will run a test shot tomorrow, with mag changer and LBD filter removed. and lower the shutter speed. I will post a revised photo when It is done.
thanks again for all the help. sam
thanks again for the advice, I will run a test shot tomorrow, with mag changer and LBD filter removed. and lower the shutter speed. I will post a revised photo when It is done.
thanks again for all the help. sam
finally I have some time to run a test this weekend. the Mag changer and any additional filters were taken out, using canon utility and zoom to pixel level does seem to improve. here is one by 70D, 28 Stacks. using Splanapo 20x. 1/25.
One thing is certain, the additional optics in the light path does make the image a little blurry. for now, the Mag changer and all the additional filters will sit in the storage box. Charles and Pau, thanks for the advice.
One thing is certain, the additional optics in the light path does make the image a little blurry. for now, the Mag changer and all the additional filters will sit in the storage box. Charles and Pau, thanks for the advice.