Transmitted Light DIC for Biological Specimen Microscopy

[dolmadis]

Hi All

I would appreciate some input and advice as to the use and benefit that can be derived from Transmitted Light DIC for biological specimens.

And perhaps some views on Reflected DIC versus Transmitted DIC please?

If, as I suspect from what I have read about these contrast techniques, Transmitted DIC is to be preferred then what could be the budget route to enjoying this?

Thanks as always.

John

[Ichthyophthirius]

Hi John, That's a very broad question. Which microscope do you have and what specimens do you want to investigate?

You can find a lot of useful information on this website: http://www.olympusmicro.com/primer/tech ... chome.html

Transmitted DIC only makes sense when your objects are phase objects (it's explained on the website). Other techniques for enhancing phase differences are also available, like oblique illumination and Zernicke phase contrast, which are much cheaper to implement.

Reflective DIC is rarely used on biological specimens, as you will see on the website.

Ichthy

[dolmadis]

Hi

Thanks for the quick reply and point of technical reference.

My growing interest is returning to the study of single cell algae and protozoans which is a long time post university days now that I am almost retired from paid work.

Since then we have digital imaging which is new to me.

I was given a reflective light Nikon Optiphot with a range of BD Objectives which are nice.

I have on old Lomo from Uni days with the Aplanatic Oblique Condenser and makeshift Polarisation.

Also recently I have acquired a Ortholux and a Zetopan where the latter has DF and Phase Contrast Anoptral and Heine.

Both need some work on lubrication and cleaning together with some work on the lighting.

Well that is the story and I will appreciate some thoughts !!

Regards


John

[Ichthyophthirius]

Hi John,

I would recommend using the Ortholux and/or Zetopan for brightfield and phase contrast for now. You wrote that you have a "Heine" for the Zetopan. Is this the "Polyphos"? http://www.microscopy-uk.org.uk/mag/ind ... topan.html That's an amazing piece of equipment, allowing phase contrast and circular oblique illumination (COL) which is very useful for protozoa.

While you do that, you can look out for a DIC set. It often takes people months and years to get a full set for their microscope. Unless you have $3000 that you can spend for a full set right now. DIC is available for the Zetopan https://www.flickr.com/photos/frezmicro/8848513362/ , the Ortholux (very expensive!) http://www.science-info.net/docs/leitz/ ... _Newer.pdf , and the Optiphot (don't confuse the transmitted setups with the setup for reflective DIC).

If you need to save money, you have to search long and hard for getting good deals. Some people try mixing components from different manufacturers (search "hybrid DIC") but there are so many differences between the manufacturers and even different product generations (different objective exit pupils, different distances of the prisms from the objectives, different angles of the interference fringes and of the additive/substractive positions) that this is very hit-and-miss and can cost you a lot of money.

There are no real shortcuts, otherwise everybody would do it. Some people here get excellent results with the PZO DIC system and Nikon objectives http://www.photomacrography.net/forum/v ... hp?t=24367 . The PZO system http://www.laboratorium.dp.ua/books/pzo-biolar.pdf allows horizontal movement of the DIC prism, which means that it can be used with a wide variety of different objectives, not just those from PZO.

Zeiss produced a lot of DIC sets, so they are on offer all the time. They can be quite cheap (relatively speaking), especially if they have delamination damage. There are various compatibility issues which you can read up on here: http://www.the-ultraphot-shop.org.uk/faq.htm

So really, the recommendation is to be patient, read up on the literature and wait until something suitable turns up. Use the Zetopan while you wait Be a microscopist, not a microscope collector.

Ichthy

[dolmadis]

Hi Ichthy

Thanks for the information on the stands that I have on the bench right now.

You are right about avoiding being a collector. That was one of the notions behind seeking advice.

My evaluation of the kind gift of the Optiphot was that the Nikon BD Plan Objectives could be usefully cycled into a DSLR/Microscope Objective project.

The Ortholux and the Zetopan came my way at the same time before I had considered my needs but were hard to turn away.

My thoughts then turned to which one to concentrate on to meet my interest and neither seemed to support my deduced desire for transmitted DIC.

So my thanks to you for the pointers of what to look for at an acceptable cost for each; whatever comes first perhaps.

As to the Heine it is the B/F,D/F, PV, Leitz condenser for the Ortholux not a Polyphos for the Zetopan.

You have reminded me that I read somewhere, fairly recently, that someone had successfully used a Heine on a Ortholux with Reichert (Zetopan) Anoptral Phase objectives. This will not involve me in extra cost because all the components are to hand.

I also have a condenser dovetail adapter for the Ortholux so I might try the Lomo Aplanatic Oblique Condenser as well.

So with you help I have now concluded that I will lube the Ortholux and vamp up the light source power because my tests to date indicate that this is necessary.

The PZO Biolar DIC is of interest if one ever turns up because I seem to recall that PZO components may have some compatibility with LOMO.

Thanks for the help.

John

[Ichthyophthirius]

The Ortholux and the Zetopan came my way at the same time before I had considered my needs but were hard to turn away.

My thoughts then turned to which one to concentrate on to meet my interest and neither seemed to support my deduced desire for transmitted DIC.

Actually, the set for Nikon Optiphot might be the easiest to find for a reasonable price. Here is a link to an Ebay offer, just to show you what it looks like (it's too expensive in my opinion that's why I didn't link it earlier) http://www.ebay.co.uk/itm/Nikon-Microsc ... 4ad8d45e5c You'd obviously also need a set of matching Nikon objectives, which tend to be quite expensive.

As to the Heine it is the B/F,D/F, PV, Leitz condenser for the Ortholux not a Polyphos for the Zetopan.

You have reminded me that I read somewhere, fairly recently, that someone had successfully used a Heine on a Ortholux with Reichert (Zetopan) Anoptral Phase objectives. This will not involve me in extra cost because all the components are to hand.

That's probably true. The Heine has a very narrow light ring, so it can be used with most phase contrast objectives, regardless of the manufacturer, except for the lowest magnification objectives (where the diameter to the Heine light ring is too large).

The PZO Biolar DIC is of interest if one ever turns up because I seem to recall that PZO components may have some compatibility with LOMO.

I have the Biolar set and it only really fits on PZO stands (which are not expensive). It definitely won't fit on Lomo. The dovetails are similar to Zeiss West Germany but slightly excentric. PZO is quite unique in everything they did.

The Lomo dovetail and some other components are compatible with pre- and post 1950 Carl Zeiss Jena!

The Ortholux and the Zetopan came my way at the same time before I had considered my needs but were hard to turn away.

They are such magnificent instruments!

[dolmadis]

Thanks Ichthy for the links to help with my hardware puzzle.

With the Optiphot my light train does not come from the base but above the objectives so I do not think that the Nikon condenser will be a solution though.

The Leitz Condenser seems to be for the Ortholux II rather than the Ortholux?

I have seen reflective DIC attachments for the Zetopan but not transmitted DIC.

The PZO Biolar seems to be the best option even though I will need a new stand (but that would be added interest anyway !! because I do like the kit aspect of microsopy too.)

But I have never seen one on Ebay (and only ever one CZ Phako Interference where I bid high but not high enough) so as the owner of a Biolar where would you suggest I look please?

Google is not much help either so I assume that the market is in Poland?

I know we have some excellent practitioners on this Forum from Poland so I wonder if anyone could help me here?

Many thanks for the input and guidance..........but where did your Biolar spring from?

Regards


John

[JohnyM]

PZO DIC is very rare, but quite cheap. The cheapest set i saw was an trinocular biolar scope with basic DIC configuration for about 750 USD - which is standard price for complete DIC set alone. But that was some time ago - now the prices are higher with every passing day.
The set alone is very versatile - i've managed to use it on Olympus BHB, Zeiss Standard, and all PZO scopes. With little metal work it can be accomodated to all 160/170 scopes.
I've used it with achromatic, planachromatic, plan fluor and plan apochromatic lenses - with slightly better and slightly worse effects. Due to prism regulation in the MPI head it's very Universal.

[Ichthyophthirius]

Hi guys,

The Heine will fit wherever its dovetail fitting fits. Or do you have one with a circular fitting?

The Zetopan transmitted DIC does exist. You can see it on the Flickr link if you click through the album https://www.flickr.com/photos/frezmicro ... otostream/

The Biolar is a good stand (not as great as the Ortholux or Zetopan, but still) and quite cheap (< $300; not to be confused with the PZO Studar) though the phototubes are hard to get. This is why people try to adapt the set to other microscopes as Johny has.

However, it's not that easy. The tube with the DIC prisms contains a Telan lens system. For this system to work as designed, I think, the tube must have the same distance from the objectives as on a PZO stand, which is 47mm. The Zeiss Standard microscope, which is superficially compatible with PZO, has a distance of 40mm from objective to dovetail. This must affect the Telan system function (the Telan system doesn't create the infinity light path it's supposed to, instead creating a converging light path). So you change the microscope tube length and negatively affect the optical correction. You won't notice this easily unless you run rigorous optical tests with high magnification dry objectives.

I can't prove this but I suspect this is why some people are dissatisfied with the PZO system. They run it on a Zeiss stand. In my opinion it's not worth doing physical modifications on the PZO DIC system (modifying fittings, removing levers and adjustment pins) when Biolars are so easy to get. Unless you can to the metal work yourself, the costs will quickly approach the costs of a Biolar stand anyway.

PZO doesn't exsist as a manufacturer anymore, I think. There is a company of this name but they sell "Chinese" microscopes. You can only get second hand DIC sets which come up on Ebay from time to time, maybe once or twice a year.

Ichthy

[dolmadis]

Hi

The Heine is fitted with the Leitz Dovetail and from inspection it does not seem removeable and be re-fitted with the Zetopan Dovetail. But that might not be necessary if I mount the Reichert Phase Objectives on the Ortholux with the Heine.

Thank you for confirming that there definitely is transmitted DIC for the Zetopan. Do you think that it might be very rare to find?

Thanks for the tips on the PZO DIC which I understand.

I will seek a PZO DIC unit and then a stand if one pops up at all.

Thanks Again


John

[Ichthyophthirius]

Do you think that it might be very rare to find?

Judging from the lack of images on the web, I would think it is very rare.

[JohnyM]

Yes. MPI has telan lenses - like any other bit of equipment for finite system (with exception of some epi-illuminatrors for dedicated, longer tube lenght lenses). It'll work fine with a couple millimeter differences - effect will be a slightly difference of magnification and if You exceed objective tolerance - some abberation.
Objective tolerance from M.Pluta "Advanced optical microscopy":
Achromatic:
10x - 27,3mm
20x - 15,8mm
40x - 7,6mm
60x - 2,0mm
100x Oil - 6,5mm
Apochromatic +/- 2mm

As you can see the tolerance is quite large with lenses up to 40x, and for higher magnification immersion lenses. Only problematic objective is dry 60x.

About biolar stand - it's really grat, and trinocular tube is easier to get than DIC but still rare. "PZO" was killed by modern capitalism... the new company is "PZO Warszawa" and only service old PZO microscopes and sell rebrand chineese one's .

Edit:
I've even managed to "adapt" PZO DIC to Olympus infinite BX40 microscope. It was held together with ducttape etc. and reqired some extra optical components and knowledge - but it could be succesfully adapted. Some photos are on our polish forum.

Also keep in mind about DIC heads (if you'll have choice) MPI5>MPI3>MPI2

[dolmadis]

Thanks Johnny.

A forum member has contacted me via Equipment Exchange with an Offer of a PZO DIC Head together with PZO Objectives.

I would need to find a KP12?

I really need to be able to read up on what the PZO DIC Head, KP12 Condenser and Objectives can deliver in each configuration and may be just without KP12.

The only "Manual" I have found is either in Russian or Polish from the link put up from Icthy.

Do you have anything you can share please?

Thanks


John

[JohnyM]

You can ask me, i know all about it. No english manual, sorry. But in Pluta "Advanced light microscopy" is quite a lot about PZO DIC.

Here are photos of set that i sold some time ago:
https://plus.google.com/photos/11021621 ... 5441625905

Kpi2 is a condenser for DIC with four prisms: 10x 20x 40x and 100x. 100x and 40x work pretty good with most 60x lenses and 100x and 10x can be used with a slightly obligue light at 5x lens - best with lassaix lens under the condenser.

Objectives (PiV probably - with red band - photo nr4 - 6) are useless to you. They contain an additional nomarski prism which with conjunction with DIC head and slit condenser (photo 9-12 ) provide an splitted (dual) image for microinterferometry.

Basically, for DIC you need:
- MPI head
- Kpi2 condenser
- Polarizer (slide in under the slit and KPI2 condenser)
- Any 160/170 lenses and stand that can accomodate KPI2 and MPI.

Member from our polish forum (from Germany) managed to take prisms out of KPI2 and fit them in Zeiss phako condenser. He use it and MPI head on zeiss phtotomicroscope. I can provide his photos via link to our forun if U interested.


Edit: Keep in mind that if you'll manage to get an MPI head and no condenser - You can obtain pretty good DIC images with slit condenser or any other condenser and self made slit (razor blades are excellent).

[dolmadis]

Thanks Johnny.

That is a most helpful review which although useful to me specifically at this stage of my hunt for DIC I am sure that it will helpful to many others in due time.

I had read a reference to the PZO objectives in a Google Search not being something I might need but as your reply explains why this is most helpful.

Yes, I am interested in the link please you mention.

Rather embarrassingly I have to say that I had never heard about a slit condenser before today !!

Are there proprietory slit condensers out there? Leitz, Zeiss, Olympus, Nikon etc?

I will Google but a shortcut is always welcome.

Thanks

John