Zeiss Standard 14

What equipment do you use? Post pictures and descriptions of your microscope(s) here!
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Glycolyse
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Zeiss Standard 14

#1 Post by Glycolyse » Thu Jan 10, 2019 4:45 pm

Hi everyone,

I got my microscope today. Fortunately for me, the seller was very kind to help me mouting everything together (objectives, head, oculars, halogen bulb, transformer).

Image

I took some samples in the sea, so far no living things.

I have a question tho. Every time I increase magnification (from x6,3 to x16 objectives for instance), the image darkens. At x40, I almost can't see. Yet, the transformer is one the "max" intensity (5.5)

https://image.noelshack.com/fichiers/20 ... 153237.jpg

Where could this come from ?

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Re: Zeiss Standard 14

#2 Post by billbillt » Thu Jan 10, 2019 5:05 pm

Hi and congrats on your purchase.. As magnification increases brighter lighting is required.. Also alignment of condenser in the light train is also important for maximum brightness.. Maybe you can see if you can set it up for Kohler lighting?..I don't know what wattage bulb your unit came with, but there is always room for improvement by going to LED lighting..


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BillT

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75RR
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Re: Zeiss Standard 14

#3 Post by 75RR » Thu Jan 10, 2019 7:05 pm

Congratulations, looking very nice!

Every time you change objective or indeed subject/sample you need to set up Köhler again.

Here is a link with an interactive tutorial that shows you how to do it.

In the tutorial you need to setup the illumination first, on your microscope you only need to do that once, unless you then change the bulb or something similar.

http://zeiss-campus.magnet.fsu.edu/tuto ... flash.html

Note that each higher magnification objective requires more light. This usually entails increasing the voltage to the bulb and opening the condenser iris.

also here is an image with condenser controls:
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Re: Zeiss Standard 14

#4 Post by Hobbyst46 » Thu Jan 10, 2019 7:41 pm

Compliments on your purchase, it will be rewarding I'm sure.

I see a can of WD40 in the background of the photo - I would not use it on any part of the microscope. It can do more harm than good.

About illumination issue with 40X objectives: since the microscope has been in use, it is unlikely that the objectives and/or condenser are SO soiled/dusty that the light beam is blocked. Very probably, the illumination should be optimized (Kohler's, as mentioned by billbillT and 75RR above). Start at brightfield ("J" condenser position) and a fully open condenser iris.

About marine organisms: For a start, I would look (a) where algae grow - collect pieces of algae and the water around them into a jar and inspect, (b) sea bottom sand - collect some "mud" or wet sand from the bottom in shalllow water near the shore (just scrape the top level, no need for a deep digging) into a container and inspect.
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Re: Zeiss Standard 14

#5 Post by Glycolyse » Thu Jan 10, 2019 9:18 pm

Hi everyone, thanks for helping !

I am actually stuck about setting Köhler illumination. "fully open the condenser aperture diaphragm (usually controlled by a lever or knurled knob located on the condenser housing" I can't find the condenser aperture diaphragm. I don't know what that is.

Overall view: https://image.noelshack.com/fichiers/20 ... 221330.jpg
Left side: https://image.noelshack.com/fichiers/20 ... 221351.jpg
Right side: https://image.noelshack.com/fichiers/20 ... 221358.jpg
Under: https://image.noelshack.com/fichiers/20 ... 221413.jpg

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Re: Zeiss Standard 14

#6 Post by 75RR » Thu Jan 10, 2019 9:23 pm

"fully open the condenser aperture diaphragm (usually controlled by a lever or knurled knob located on the condenser housing"
Also known as condenser iris. see red arrows on photo of condenser
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Re: Zeiss Standard 14

#7 Post by Glycolyse » Thu Jan 10, 2019 9:38 pm

75RR wrote:
"fully open the condenser aperture diaphragm (usually controlled by a lever or knurled knob located on the condenser housing"
Also known as condenser iris. see red arrows on photo of condenser
Alright, thanks, got it ! ;)

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Re: Zeiss Standard 14

#8 Post by Glycolyse » Thu Jan 10, 2019 9:59 pm

Quick update: I changed the height of the phase condenser, which was really far from the stage. And to be honest, it was mind-blowing. The quality of the image changed drastically ! I'll pursue with the setting of Köehler Illumination.

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Re: Zeiss Standard 14

#9 Post by Glycolyse » Fri Jan 11, 2019 8:36 am

Hi everyone !

I played a lot with the microscope yesterday evening. I found some living organisms, mainly diatoms !

Somes photos (sorry for the low quality, I have to deal with my phone at the moment):

https://image.noelshack.com/fichiers/20 ... 000542.jpg
https://image.noelshack.com/fichiers/20 ... 235159.jpg
https://image.noelshack.com/fichiers/20 ... 234124.jpg

I almost finished the Köhler illumination. The condenser was almost centered already, but when I tried to move the X/Y axis of it, it didn't change anything.

I didn't go with phase contrast yet. Do I have to set Köhler illumination for each objective of the Phase Contrast ?

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Re: Zeiss Standard 14

#10 Post by 75RR » Fri Jan 11, 2019 9:04 am

First pictures are always exciting. Well done.

Do not confuse the condenser x and y axis with centering the condenser. The x and y axis centers the Phase annulus that lines up with the one inside the objective.

To center the condenser you move the two screws that are placed in the condenser holder. One on each side. See image.

You can tell when the condenser is centered by opening the field diaphragm to the edge of the field of view (what you see through the eyepiece).

It should be the same distance from the edge all the way around.

As to Köhler, yes all the time. With practice it just takes a few seconds.
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Re: Zeiss Standard 14

#11 Post by MicroBob » Fri Jan 11, 2019 11:33 am

The centering mechanism works with two screws that work against one spring loaded plunger.
The screws usually move, but the plunger often is stuck due to hardened grease. So you should check wheter the condenser actually moves when you turn the screws out.

Bob

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Re: Zeiss Standard 14

#12 Post by Glycolyse » Fri Jan 11, 2019 4:32 pm

Thanks for your help guys.
As to Köhler, yes all the time. With practice it just takes a few seconds.
For example, I'd like to see with Phase contrast. I put my Ph2 x16 obj, put on the "2" on my Phase contrast condenser, and then adjust for Köhler illumination ? Because at the moment, when I put my Ph2 x16 obj, or Ph2 x40 obj, and then put "2" on the Phase contrast condenser, nothing appears; it is almost pitch black.

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Re: Zeiss Standard 14

#13 Post by Hobbyst46 » Fri Jan 11, 2019 4:49 pm

Glycolyse wrote:Thanks for your help guys.
As to Köhler, yes all the time. With practice it just takes a few seconds.
For example, I'd like to see with Phase contrast. I put my Ph2 x16 obj, put on the "2" on my Phase contrast condenser, and then adjust for Köhler illumination ? Because at the moment, when I put my Ph2 x16 obj, or Ph2 x40 obj, and then put "2" on the Phase contrast condenser, nothing appears; it is almost pitch black.
It is possible to achieve Kohler when the condenser is at position "2". However, I suggest you try the longer path, as follows:
Start with the 10X (if you have one) or the 16X, and condenser "J". Set Kohler. Rotate the condenser to position "2". The illumination should stay the same as it was. Switch to the 16X Ph2 objective. Now remove one of the eyepieces, insert the phase telescope, focus the telescope and try to align the dark phase ring of the objective with the bright ring of the condenser. This is done with the X- and Y- knobs marked in red by 75RR in the photos above. NOT WITH THE CENTRATION THUMB SCREWS. Once the rings overlap, phase contrast will be achieved. Return the eyepiece. Occasionally you can tweak the condenser up and down - but only so slightly. Same procedure for the 40X. With time and experience you can later shortcut directly to position "2" from the start.
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Re: Zeiss Standard 14

#14 Post by 75RR » Fri Jan 11, 2019 5:06 pm

Agree. In Phase, Köhler is done initially in brightfield using position "J", you then rotate the condenser to the appropriate phase number for your phase objective.

If you do not have a Phase telescope yet it is possible to achieve Phase by eye, by looking in turn down the eye-tube and then through the other eyepiece until it looks phase-like.

Not ideal though. A Phase telescope is really a must.

Not much light is needed to overlap the annuli - you probably had the condenser set too low. When you set up Köhler, it will then be at the right height and the illumination sufficient.

Always start with your lowest magnification objective and work up.
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Re: Zeiss Standard 14

#15 Post by Glycolyse » Fri Jan 11, 2019 10:13 pm

Thanks for the tips ! I'll try that out.

I wanted to try Focus Stacking. I took a series of 16 images, and stacked them using the Helicon Focus (30-days trial). I tried to be as accurate as possible, so I customized an used toilet roll (without paper), which I put on one of the eyepiece (carefully). It allowed me to stay as stable as I can while holding the phone, taking the shots, and focusing for each photo.

Here is the result:

Image

What do you think ? I know it is not that great, but I had fun finding a way to do it.

Also, it seems that I might destroy/squeeze organisms while preparing the slide. I often find cell-like structures that looks destroyed, with the intracellular components out of them. Maybe I should bring a photo of what I am talking about ? That would explain why I don't find so much (very rarely actually) living organisms. It might also be my collecting process. I am taking submerged rocks, and using a toothbrush, cleaning them in a bowl. Then, I wash the rock, and put water on it so I collect as many organisms as I can.

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Re: Zeiss Standard 14

#16 Post by 75RR » Sat Jan 12, 2019 6:24 am

That is very good, especially for a handheld phone.

Was that with the 16x?

Have you considered using a phone holder until you sort out a trinocular head?

I believe it will make things a little easier and they are not too expensive.

Perhaps something similar to this:

https://www.amazon.co.uk/Hoyxel-BlackBe ... microscope
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Re: Zeiss Standard 14

#17 Post by Glycolyse » Sat Jan 12, 2019 9:05 am

Thanks 75RR !

I think I used the x40 one, should've mentioned it earlier. I considered a phone adapter, why not. My first thought was to wait for a trinocular head. But if it not too expensive, I should give it a shot.

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Re: Zeiss Standard 14

#18 Post by 75RR » Sat Jan 12, 2019 9:24 am

I am taking submerged rocks, and using a toothbrush, cleaning them in a bowl. Then, I wash the rock, and put water on it so I collect as many organisms as I can.
That is a good way to collect diatoms if the rocks have a brownish colour to them. Do not use too much water, you are only diluting the sample.

If you want to see other things collecting alga is a good way. Many animalcules take shelter in them.

http://www.microscopy-uk.org.uk/index-n ... k/ponddip/
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Re: Zeiss Standard 14

#19 Post by Glycolyse » Sun Jan 13, 2019 11:22 am

Hi everyone,

So far it is very rare to see living organisms. I collected a lot of moss this morning, put some under the microscope, but not so great. I wonder why. Are the organisms pinned by the cover glass ? How do I prevent that ? How do you prepare this kind of glass ?

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Re: Zeiss Standard 14

#20 Post by 92111 » Sun Jan 13, 2019 11:28 am


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Re: Zeiss Standard 14

#21 Post by Glycolyse » Sun Jan 13, 2019 11:34 am

Very cool video, but we can't see exactly how he prepared the final glass at the end. How much drop of water should I put before putting the cover glass?

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Re: Zeiss Standard 14

#22 Post by Hobbyst46 » Sun Jan 13, 2019 11:42 am

Glycolyse wrote:Hi everyone,

So far it is very rare to see living organisms. I collected a lot of moss this morning, put some under the microscope, but not so great. I wonder why. Are the organisms pinned by the cover glass ? How do I prevent that ? How do you prepare this kind of glass ?
If you are looking for marine protists, worms etc, I would try the near-shore "mud" from the see bottom, just the top layer of wet sand at a distance of a meter or less from the front water line (if accessible and not too rocky).
If you are interested in freshwater protist, just look for any standing water - water hole, the slimy mass under a dripping water stream from a tap, an irrigation overflow pipe; collect the sample as is - do not add water.
Growing an initial batch of protists is super-easy: make a hay infusion, or alternatively place a banana peel in a small jar and leave it aside in a warm place (say, 25-30C). Within a week, more or less, it will be swarming with cilliates. Just for a start.
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Re: Zeiss Standard 14

#23 Post by Glycolyse » Sun Jan 13, 2019 4:38 pm

Hi guys, quick update.

I decided to give another shot to sea life. I took some samples, it was very windy so not the greatest conditions out there. At the end of the session, I noticed there was moss growing just a meter from the sea, in water. I took some samples, and I just observed the moss under the microscope. Holy ! There are so much little guys in this one ! I'll post photos if you want.

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Re: Zeiss Standard 14

#24 Post by MicroBob » Sun Jan 13, 2019 6:02 pm

Glycolyse wrote: I'll post photos if you want.
You bet! :D

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Re: Zeiss Standard 14

#25 Post by Glycolyse » Sun Jan 13, 2019 8:52 pm

Here is what I mainly encountered in the moss: https://image.noelshack.com/fichiers/20 ... 180209.jpg

Rotifers I guess ?

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Re: Zeiss Standard 14

#26 Post by 75RR » Sun Jan 13, 2019 10:20 pm

Looks like it might be Philodina

Forum member's lukem321's rotifer key: https://www.dropbox.com/s/9q5iepgwoed6f ... y.pdf?dl=0

Note: As always there is no need to join dropbox to download anything.
Just scroll to the bottom of the 'invite page' and press on 'No thanks, continue to view'
Then press download on top right of the page.
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Re: Zeiss Standard 14

#27 Post by Glycolyse » Sun Jan 13, 2019 10:25 pm

Thank you 75RR. The drawing and the description match with what I saw. They must be Philodina then !

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Re: Zeiss Standard 14

#28 Post by Glycolyse » Sun Jan 13, 2019 10:58 pm

Update about Phase Contrast:

As I don't have a Phase Telescope, I tried to do it judging with the eye. Wasn't too hard for the x16, a bit more for the x40. I didn't go for the x100 yet. It looked like I achieved it - but it was still dark, so I fully opened the condenser aperture diaphragm, but it was not that bright either. Then, I changed the condenser's height (I know, what ? Köhler was already established for the x16). When going lower, the image got brighter, until it was too messy. I tested something else, something really weird. I wanted to know if a buffed phone's screen (in white color) could do something about the image. So I placed it just after the Halogen beam. I'll post 1 photo of each result, obj x16.

Condenser height's established correctly. The image is dark: https://image.noelshack.com/fichiers/20 ... 234937.jpg

Condenser height's has been lowered. The image is brighter: https://image.noelshack.com/fichiers/20 ... 235154.jpg

Phone's screen setup: https://image.noelshack.com/fichiers/20 ... 235249.jpg
Interesting result: https://image.noelshack.com/fichiers/20 ... 235317.jpg

Note: As it is low quality photos, it partially represents reality. For instance, the first image ("dark") is darker in reality. The Phone's screen result is more bluish in reality.

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Re: Zeiss Standard 14

#29 Post by 75RR » Mon Jan 14, 2019 6:52 am

Don't think you quite got Phase yet.

The field of view should be more widely and evenly illuminated when you have achieved Köhler.

Could you post a photo of a larger subject (an alga for example) with the field diaphragm in view (just visible along the edge of the field of view) and sharply focused.


Also you should test if your filament is focused:

The steps are:

1) Remove any opaque filters that may be in the light path.
2) Close the condenser iris completely
3) Place a mirror or slide under the condenser, angle it so that you can see the closed iris
4) You should be able to see the filament coils reflected on the condenser iris. (This is the Front Focal Plane)
5) If no filament coils are visible or they do not look sharp you need to move the light source either in or out until the filament coil is sharply focused.

Image
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Re: Zeiss Standard 14

#30 Post by 75RR » Mon Jan 14, 2019 6:59 am

If you have't found a Phase telescope yet here is an economical one that should be fine.

https://www.ebay.co.uk/itm/23-2mm-Phase ... :rk:7:pf:0
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