LWD and no-coverslip Olympus objectives

Malcolm Storey · Post by **Malcolm Storey** » Fri Sep 18, 2015 6:08 am

I've been looking into no-coverslip objectives for my Olympus BX53. Olympus offer the following for no coverslip, or with adjustment for coverslip thickness:

LWD objectives:

LUCPLFLN 20x (LWD): WD 6.6-7.8mm, NA 0.45
LUCPLFLN 40x (LWD): WD 3-4.2mm, NA 0.6

Standard WD objectives:

MPLFLN 20x: WD 3.1 mm, NA 0.45
MPLFLN 40x: WD 0.63mm, NA 0.75

My current objectives, corrected for 0.17mm coverslips

UPlanFLN 20x NA 0.5
UPlanFLN 40x NA 0.75

A new LUCPLFLN 20x (LWD) came up on eBay, so I bought it. I'm now trying to decide what to do about 40x.

I'm wondering whether it's worth getting the 40x LWD given that the NA is only 0.6 - that's quite a drop from 0.75 (though it'll make stacking easier!) But will it actually give significantly more detail than 20x (NA 0.45) or would it just be a bigger image? (Image capture using Canon 1000D or 600D.) Has anybody tried this objective?

I'll be using it for small inverts and fungi etc, and close-ups of their parts.

Charles Krebs · Post by **Charles Krebs** » Fri Sep 18, 2015 4:10 pm

Malcolm,

It's a call only you can make. You will get about a 35% resolution increase from the 20/0.45 to the 40/0.60. I think you would see a difference.

A lot would depend on how you will be illuminating the subject and the possible "depth" of the subject. A NA of 0.75 would obviously be nice, but with a working distance of 0.63mm you would need to use either transmitted or epi illumination. And if the subject is not quite flat I could envision parts of it contacting the front of the objective if you go for a "deeper" section. The 3-4.2mm working distance of the LUCPLFLN is still pretty tight if you plan on using reflected light with external light sources, but way better than the 0.63mm, especially if you have a significantly 3-dimension subject.

BTW... the smallest resolvable detail based on NA is about as follows.

NA 0.75
.45 micron

NA 0.6
.56 micron

NA 0.45
.75 micron

Malcolm Storey · Post by **Malcolm Storey** » Sat Sep 19, 2015 2:24 am

Interesting that it gives only a 35% improvement in resolution.

Yes, I want them for epi and the difficulty of illuminating with sub 1mm clearance is a good point. (I generally surround with pieces of styrofoam cup and blast with flash from the outside. I guess a lot will depend on the shape of the objective nose and whether it's black or silvered! But it's hard to get enough light in at high mag without having low NA as well).

I was thinking in terms of completing my set of objectives rather than specific uses. In the past I've often bought kit for a specific purpose then it's opened up new avenues that are far more interesting than the original justification, so I now tend to go for "empowerment" and then work out what it's good for when I start using it. Actually, there aren't many cases in entomology where you need to go that small (but then so far I haven't been able to...) but I have always wanted to photograph fungal spores sitting in their 4's on toadstool gills - a nice flat subject - but it'd need a bit of an angle (and anyway, gills are often wedge shaped). May need to use LED as that many full power flashes might cook it.

I think I'll go for the LWD and perhaps get the other one too if it comes up cheap on eBay.

Thanks for your comments and especially for the resolution figures.

Malcolm Storey · Post by **Malcolm Storey** » Mon Sep 21, 2015 2:01 am

Interesting that the ratio of increase of resolution is a lot less than the ratio of successive magnifications. You could argue that either the lower mag objectives are over-engineered, or the higher mag obj's have empty magnification! I guess it's a mixture of compromise and value for money (you want to clearly see the detail at the top end, and it's cheap to add more NA at the bottom end).

But it does mean I'm justified in blowing up the low end a bit with extension tubes.

Pau · Post by **Pau** » Mon Sep 21, 2015 4:56 am

Malcolm Storey wrote:Interesting that the ratio of increase of resolution is a lot less than the ratio of successive magnifications. You could argue that either the lower mag objectives are over-engineered, or the higher mag obj's have empty magnification!

In fact it's more related with physics laws than with technology or economy, BTW expensive high corrected objectives like Pan Apos have higher NA and resolution than cheap achromats, but this relationship still applies to them. In most cases the high magnification objectives have higher NA and provide higher actual resolution at the subject side but the actual F number is higher, so the image is less sharp at the sensor/eyepiece side, ie you need less megapixels to capture all the detail present at the objective primary image. If you push the magnification too far you get empty magnification, and this is easier to happen with high mag. objectives
Play a bit with this interactive tutorial:
http://www.microscopyu.com/tutorials/ja ... index.html

LWD are more limited than short WD objectives just because NA is a function of the angle of the light

ChrisR · Post by **ChrisR** » Mon Sep 21, 2015 6:31 am

LWD are more limited than short WD objectives just because NA is a function of the angle of the light

As I understand it, that's back at the practicalities, rather than with the constraints of physics?
The longer WD objective would get to be larger and with its entrance pupil in a difficult-to-make place. I haven't seen any figures for theoretical limits.

Pau · Post by **Pau** » Mon Sep 21, 2015 9:58 am

As I understand it, that's back at the practicalities, rather than with the constraints of physics?

Yes and no: for moderate NA like a 10X 0.30 you can find objectives with 1mm WD and others with huge WD like the Mitu, but you are not going to find many ELWD 10X 0.45 (well, a Mitu 20X 0.42 pushed down will do). You can't find any 40X 0.95 with real LWD (a physics limitation), and all 20X 0.75 I've seen have small WD, an economy/technology/physics mixed case.

To make a high NA LWD you need to fill the angle a very large frontal lens to have large entrance pupil, and this will be limiting. The Mitus and similar objectives have nice NA for the low magnification models, pretty normal for the 20X and pretty small for higher magnifications. Engineers do know about the Abbe's law but make some compromises to maintain the versatility for industrial uses. Of course some improvement is physically posible.

This interactive tutorial: http://www.microscopyu.com/tutorials/ja ... index.html
can be instructive to visualize the topic

ChrisR · Post by **ChrisR** » Mon Sep 21, 2015 1:37 pm

That still doesn't indicate a maximum NA/WD set. They're holding the objective diameter constant in the applet.
If the maximum diameter constraint were relaxed completely, where would the limit be for, say a 20x objective?
We know from unusual designs - most extreme I think are the the Measuring Microscope objectives, that the positions of the optical planes can be in surprising places.
I don't suppose there would be much market for a 20x, NA 0.95 objective with a 20mm WD, at maybe 70mm diameter and the price of a house.

Malcolm Storey · Post by **Malcolm Storey** » Mon Sep 21, 2015 1:47 pm

Don't be so sure - shine a laser through it and you could etch silicon chips or trigger fusion in a deuterium pellet!

ChrisR · Post by **ChrisR** » Mon Sep 21, 2015 1:52 pm

Seeing the way the whole neighbourhood shakes when the deuterium "goes off" I'd fear for the alignment of the lens!

rjlittlefield · Post by **rjlittlefield** » Mon Sep 21, 2015 6:49 pm

ChrisR wrote:If the maximum diameter constraint were relaxed completely, where would the limit be for, say a 20x objective?

I don't know of any theoretical limit.

However, there's a big problem that for any particular design, the wavefront error scales directly with the size of the lens.

So, suppose you start with an excellent 1/4-lambda design for 2 mm FL, NA 0.70, and 6 mm working distance -- say, a Mitutoyo M Plan Apo 100X.

Then just make it 5 times bigger.

What you'd get is a 10 mm lens (20X), still at NA 0.70, now with 30 mm working distance, and with a completely dreadful 5/4-lambda wavefront error.

To get the aberrations back down into the acceptable range would require an even more elegant design with much tighter fabrication constraints.

Yep, "price of a house" seems about right.

--Rik

smith88g · Post by **smith88g** » Thu Mar 09, 2017 11:51 am

Sorry to bump an old thread but I was curious about something.
Aren't LUCPLFLN for inverted microscopes? Does this matter that the BX53 is an upright microscope?

What have you thought about it so far Malcolm?

Malcolm Storey · Post by **Malcolm Storey** » Thu Mar 09, 2017 1:08 pm

I've had the 20x and 40x objectives for a couple of years but hardly used them.

Both objectives have adjustable collars for slide/coverslip thickness, which I guess derives from their use from below so they peer thru the slide rather than the coverslip. Otherwise, it never ocurred to me that it would make any difference.

These are the only stacks I've taken so far (all with the 20x):

http://www.bioimages.org.uk/image.php?id=125968 (63 frames)
http://www.bioimages.org.uk/image.php?id=126158 (53 frames)
http://www.bioimages.org.uk/image.php?id=122012 (39 frames)

smith88g · Post by **smith88g** » Thu Mar 09, 2017 1:51 pm

Malcolm Storey wrote:I've had the 20x and 40x objectives for a couple of years but hardly used them.

Both objectives have adjustable collars for slide/coverslip thickness, which I guess derives from their use from below so they peer thru the slide rather than the coverslip. Otherwise, it never ocurred to me that it would make any difference.

These are the only stacks I've taken so far (all with the 20x):

http://www.bioimages.org.uk/image.php?id=125968 (63 frames)
http://www.bioimages.org.uk/image.php?id=126158 (53 frames)
http://www.bioimages.org.uk/image.php?id=122012 (39 frames)

What do you find yourself using the most these days at 20x/40x?

The images are cool. I don't know if the slight blur is just because of the lower NA or the inverted vs upright. I only wonder if the light path distance is different in upright vs inverted microscopes.
Does anyone know?

Malcolm Storey · Post by **Malcolm Storey** » Thu Mar 09, 2017 2:30 pm

At that magnification I'm usually working with (temporary) slides, so I use UPlanFLN objectives.

I assumed the slight blur was out-of-focus detail from adjacent frames in the stack. Or I didn't spend enough time retouching/in PhotoShop!

The camera is an EOS1000D (quite old now - I've had it since 09). The modern Canons seem to give crisper images (perhaps just in-camera sharpening?)

The camera adaptor is from Best Scientific. http://www.bestscientificweb.com/
I assume its lens is just achromat - there's more colour fringing in photos than by eye.

Keep hoping Rik might correct for it in ZS!!

LWD and no-coverslip Olympus objectives

LWD and no-coverslip Olympus objectives

Only a 35% improvement in resolution

Inverted microscope

Re: Inverted microscope

20x and 40x