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Varroa destructor

 
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GrahamR



Joined: 14 Feb 2018
Posts: 3
Location: Cheshire, UK

PostPosted: Wed Feb 14, 2018 3:24 pm    Post subject: Varroa destructor Reply with quote

Hi All, this is my first post. I have been following this forum for several months now and have learnt so much. I'd like to start with a big THANK YOU to everyone who has shared their knowledge and made this such a valuable resource. Two months ago I started to build my own macro rig, using as far as possible, equipment I already had. The result is lots of compromises but I can always upgrade over time.

Basic setup is:
EOS500D with a 50mm f1.8 FD lens reversed onto a 100-200mm f5.6 FD lens.
Homebuilt stackrail with PIC 16f microcontroller
Two 1970's Vivitar flashguns triggered from the stackrail controller
CombineZ software

Here are some of my recent pictures:

Varroa destructor mite taken from one of my beehives. This mite is causing lots of colony losses throughout the world.
35 images, 32.5um







51 images 32.5um

These are all crops from the full frame. Magnification is around 4x which is not enough to fill the frame. ( In the first photo the visible width of the mite is 1.6mm, which comes out at 6+mm on the sensor). The mess on the right of the mite in the last photo is the pin that it is superglued to. I painted the pin matt black but that hasn't totally hidden it.

Whilst I'm happy with these as first attempts, I know there is a long way to go before I get anywhere near most of the superb images posted here. There is a lot of haloing round the body hairs which I think Zerene would deal with better and I need more magnification for this subject so that the image fills more of the frame.

What should I change first to improve on these? Different lenses, different lighting, Zerene rather than CombineZ. Any and all suggestions will be most welcome!
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rjlittlefield
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Joined: 01 Aug 2006
Posts: 18501
Location: Richland, Washington State, USA

PostPosted: Wed Feb 14, 2018 5:02 pm    Post subject: Reply with quote

Graham, welcome aboard!

I'm thinking that your two biggest improvements would be 1) more diffuse illumination, and 2) replace that reversed 50 mm lens with a 10X NA 0.25 infinite type microscope objective. Those two things, especially combined, will get you a lot more resolution.

The downside is that it will also get you a lot less DOF per frame, only about 8 microns, so your stacks will be deeper and demand more precision from the rail.

The big advantage of Zerene Stacker for this sort of work is its retouching capability. CombineZP's "Pyramoid Maximum" method is similar to Zerene Stacker's PMax. Haloing around body hairs is common, almost unavoidable, and not likely to be much different between the two software packages. (Full disclosure: I'm the fellow who wrote Zerene Stacker, after turning CombineZ's author on to pyramid methods.)

--Rik
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GrahamR



Joined: 14 Feb 2018
Posts: 3
Location: Cheshire, UK

PostPosted: Thu Feb 15, 2018 3:23 am    Post subject: Reply with quote

Rik,

Thank you, that gives me a few things to try out. There are so many variables it is difficult to know what to try first.

I’m currently using a polystyrene cups as diffusers but there are plenty of alternatives suggested on here for me to experiment with.

I have several microscopes so have a few objectives I can try. They are not infinite, and probably not up to the quality of those mentioned in other topics here, but I do have bellows and extension tubes I can use in place of the tube lens. I’m waiting on an RMS adapter which has been on it’s way from China for the past couple of weeks. My stackrail is capable of 2.5um steps in theory but that doesn’t account for stiction etc.
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rjlittlefield
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Joined: 01 Aug 2006
Posts: 18501
Location: Richland, Washington State, USA

PostPosted: Thu Feb 15, 2018 9:52 am    Post subject: Reply with quote

GrahamR wrote:
I’m currently using a polystyrene cups as diffusers but there are plenty of alternatives suggested on here for me to experiment with.

Polystyrene is usually an excellent diffusing material.

What prompts me to say you need more diffuse illumination is for example that very bright blown out highlignt on the left side of the second image. That indicates unambiguously that you have a lot more light coming from the narrow range of angles whose reflections form that highlight, than you do from all the other angles that reflect off the other surfaces of the mite.

Given that you're already using what is probably an excellent diffusing material, maybe what you need to do is just reposition your lights so that the outside of the cup is more evenly illuminated.

Be cautious also about the possibility of undiffused light getting reflected onto the subject in unexpected ways. In your last image, those extremely bright glowing front appendages suggest that maybe some stray light is getting in and lighting up just those exposed parts.

Quote:
They are not infinite, and probably not up to the quality of those mentioned in other topics here, but I do have bellows and extension tubes I can use in place of the tube lens.

That sounds great. With bellows and tubes, and a low power objective like 10X NA 0.25, you'll have some freedom to adjust magnification by changing the tube length.

--Rik
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ChrisR
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Joined: 14 Mar 2009
Posts: 7502
Location: Near London, UK

PostPosted: Thu Feb 15, 2018 11:18 am    Post subject: Reply with quote

You can get a hotspot if you purt a flash hard up against a poly cup - moving them away ~3 inches should be fine.

As you may know, most objectives were designed for colour corrections in an eyepiece, which you won't be using. Which do you have?

When you get your adapters remember to check for and deal with shiny surfaces, eg at the back of the objective. Black paper is usually good enough to kill them.
My first adapter was card with a hole bored through it Wink
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GrahamR



Joined: 14 Feb 2018
Posts: 3
Location: Cheshire, UK

PostPosted: Thu Feb 15, 2018 3:26 pm    Post subject: Reply with quote

Rik, Chris,

You both got it right. The left hand flash was too close to the poly cup. Partly to give more (too much) illumination, and partly to avoid it falling off the edge of the table. I definitely need more space!

Here is the mite with the left flash moved further away.



Chris, the objectives I have are probably Chinese. I bought the microscopes in the UK some years ago from a reputable supplier, but they are not named and I expect they are of Chinese origin. I have a couple to choose from with the usual x4, x10, x40 objectives. It will be interesting to see how they perform once the adapters arrive.

Last week I bought a microfiche lens from ebay. At £6.99 I thought it was worth a punt.

https://www.ebay.com/itm/192447492170

The vendor had several similar lenses for sale.

Today I have made an adapter so that I can mount it onto the 200m tube lens. Here is the first stack result:




140 images, 10u step, Combine Z. Unlike my previous uploads, this one is not cropped. I have removed a few 'stack worms' using Photoshop.

Magnification when mounted on the tube lens is approx. x10

What a difference a day makes!
Very Happy
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ChrisR
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Joined: 14 Mar 2009
Posts: 7502
Location: Near London, UK

PostPosted: Thu Feb 15, 2018 4:45 pm    Post subject: Reply with quote

Progress!! Nice to see.Very Happy
If you have a Photoshop version, I might suggest Image>Adjustments>Curves>Options>Enhance per Channel Contrast

You can be lucky or not, with random objectives.
You may find the image circle yours cover, isn't as large as you'd like.

There are some good ones mentioned in this thread : http://www.photomacrography.net/forum/viewtopic.php?t=36036

and some new infinites which therefore may not be suitable for you in this thread in the FAQs http://www.photomacrography.net/forum/viewtopic.php?t=27205
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