Best size objective for Pond life?

[Joe Henry]

I mostly want to look at pond life, what are the most practical objectives to use for this purpose? Thanks..Joe.

[WWWW]

Hi,

I like looking to pond life too and use:
- 4x (overview, scanning)
- 10x 20x 40x
- 60x (difficult if the specimen is too tick)

WWWW

[deBult]

20* is by far my most favoured objective for pond life (4* for the overview, 10* to find your subject, 20* to observe its behaviour, 40* to identify).

[apochronaut]

Pond life is very diverse in all ways. Some non bacterial organisms are less than 5 microns. You will see only about 10% of what is going on with a 20X achromat objective. If you stay a keener, you will quickly find that at least 4 objective magnifications are necessary to get the big picture. Avoid a 60X objective unless you can afford a more advanced highly colour corrected one.Use a 15X eyepiece with your 40X instead.
4 or 5X, 10X, 40X and 100X are the ones but the 100X must have an adequate condenser and illumination.
Don't avoid oil immersion. It is necessary for a fulfilling adventure into the pond ecology and just requires a little precision. It is not dissimilar to learning to use gears on a bicycle once you already know how to ride.
Absurdly, one of the reasons some people avoid oil immersion is because they are too cheap to pay for the oil?!

[deBult]

Apo, you are right the 100* adds to the experience.

But, in my personal experience when using the 100* oil objective on pond sample slides, the coverglass often moves when scanning the slide.
And when using the 100* I often want to switch back to a lower enlargement, requiring a 40* or 50* oil, (and a 20* oil I do not own).

YMMV though.

[macnmotion]

Apo, you are right the 100* adds to the experience.

But, in my personal experience when using the 100* oil objective on pond sample slides, the coverglass often moves when scanning the slide.
And when using the 100* I often want to switch back to a lower enlargement, requiring a 40* or 50* oil, (and a 20* oil I do not own).

YMMV though.

I can routinely go back from 40x oil or 100x oil to 10x and 20x non oil without fear as my 10 and 20x objectives go nowhere near the slide. I had a similar problem with slipping cover glasses until I switched to the super long cover glasses. The additional friction holds everything in place.

[PeteM]

I'll second the desirability of including a 20x in the mix. It's often omitted in educational-level microscopes, leaving that big gap between 10x and 40x. However, it's often near perfect for having enough magnification to see detail, enough depth of field for a satisfying image, and not too high a magnfication to lose focus, have great difficulty in tracking moving critters, or sweep into oil as described above.

A better quality 20x may also have a numerical aperture of .50 or even .75 - meaning that it will support effective camera magnfications into the range of 400x and beyond.

[Tom Jones]

I agree that a 20x is highly valuable for pond life. I consider it a must have, and generally more useful and less trouble than a 100x oil. I have a 20x on every microscope I own that isn't an antique display scope. All of my outreach scopes have them as well. I even have mag changers on my BX series scopes. It's hard to have too many choices, particularly when framing for photography.

[apochronaut]

Of course a 20X is great, better a 25 but for a starter with only 4 holes, what do you leave out for a 20? Using a 15X eyepiece all of a sudden effectively adds a 6X, 15X and 60X to the existing 4 objectives with wider field and longer working distance.

[tpruuden]

I wonder, if the modern electrophysiology objectives at 20x and 40x would be the best option - optimized for water immersion, long working distances, quite high NA.

[charlie g]

I suggest, as does Phil/apo, learn to use your oil-100X objective..be selective..do what you enjoy with 20X, 40X, 60X dry...save 100X-oil for last excitement of your designated treat organisms on a wet mount slide.

As was suggested already..larger rectangular coverslips permit at least 80% of your wetmount slide observations..safe distance from coverslip border waters..so no 'mucking/ wetting your close working distance objectives.

So many diatoms, desmids, bacteria, smaller protozoa are wonderful under 100X-oil objective..save this 'comitting to oil for final wetmount slide observations. Learn to 'work that water-column film thickness to the minimum thickness/sweet spot film thickness..just before use of oil-objective..paper-towel slurp water from your wetmount slide with a snippet of this absorbant paper..learn that sweet spot of use of 100Xoil objective.

Much more use of 1.5% methylcellulose viscosity tool added 50/50 with pond water sample before you 'drop a coverslip'..this really opens observations with your: 40X, 60X, oil-100X objectives on larger protozoa, larger bacteria, etc..

As has been said here..the full complement of your objectives: ' offer the big picture/ context' of pond assemblages.'.enjoy these with the larger rectangular coverslips..to avoid the cover slip border waters.

charlie g

[deBult]

Apo, you are right the 100* adds to the experience.

But, in my personal experience when using the 100* oil objective on pond sample slides, the coverglass often moves when scanning the slide.
And when using the 100* I often want to switch back to a lower enlargement, requiring a 40* or 50* oil, (and a 20* oil I do not own).

YMMV though.

I can routinely go back from 40x oil or 100x oil to 10x and 20x non oil without fear as my 10 and 20x objectives go nowhere near the slide. I had a similar problem with slipping cover glasses until I switched to the super long cover glasses. The additional friction holds everything in place.

Great, as stated YMMV: my 20* is an Olympus SPlanApo with high NA: it is VERY close to the slide, and at risk touching the remaining oil residu when switching back from the 50* oil. A normal 20* Achromat would not suffer from this “handicap”. But love this 20* PlanApo: one of Olympus best from the 160 DIN era.

Will try and find some nr 0 super long cover glasses: thanks for bringing it to my attention.

[macnmotion]

Apo, you are right the 100* adds to the experience.

But, in my personal experience when using the 100* oil objective on pond sample slides, the coverglass often moves when scanning the slide.
And when using the 100* I often want to switch back to a lower enlargement, requiring a 40* or 50* oil, (and a 20* oil I do not own).

YMMV though.

I can routinely go back from 40x oil or 100x oil to 10x and 20x non oil without fear as my 10 and 20x objectives go nowhere near the slide. I had a similar problem with slipping cover glasses until I switched to the super long cover glasses. The additional friction holds everything in place.

Great, as stated YMMV: my 20* is an Olympus SPlanApo with high NA: it is VERY close to the slide, and at risk touching the remaining oil residu when switching back from the 50* oil. A normal 20* Achromat would not suffer from this “handicap”. But love this 20* PlanApo: one of Olympus best from the 160 DIN era.

Will try and find some nr 0 super long cover glasses: thanks for bringing it to my attention.

My 20X is a Nikon Plan Apo, and yes it's somewhat close to the cover glass but the layer of oil that trails from my oil objectives is so thin there's no way I'd hit it, but you're right I need to be mindful I don't turn the focus knob the wrong way. I make it a habit to move the objectives down a long way from the slide when switching back, then slowly coming back up to focus. Mostly, though, it's the 10X I go back to just to quickly reorient myself on the slide.

[macnmotion]

I suggest, as does Phil/apo, learn to use your oil-100X objective..be selective..do what you enjoy with 20X, 40X, 60X dry...save 100X-oil for last excitement of your designated treat organisms on a wet mount slide.
Much more use of 1.5% methylcellulose viscosity tool added 50/50 with pond water sample before you 'drop a coverslip'..this really opens observations with your: 40X, 60X, oil-100X objectives on larger protozoa, larger bacteria, etc..

When looking up "methyl cellulose" where I live, I only get results for carboxy methyl cellulose. Is that usable for this purpose?

[apochronaut]

Economics is always a prejudicial one. It would be nice to have access to any or all choices. I go back mentally to the 20 years or so of having only a 3 objective sub. R.M.S. hobby scope and then a 3 objective Spencer , both monocular with a 10 watt remote illuminator. The latter was bought in a pawn shop for 175.00. Pond water was an endless source of inspiration. Very insignificant creatures, so full of life and exhibiting such oddly decisive behaviour.
Ebay brought a range of economically feasable possibilities that I would have been in awe of in those days, especially those that a 4 or 5 place nosepiece brings. Ebay has afforded me the luxury of having a 6 place interchangeable nosepiece now but I don't forget the limitations of a single 4 place. Between the 3 objective years and the 3 x 6 objective years I used a 4 objective Spencer #3 with apochromats. 10X, 20X, 44X and 90X oil , and I looked at a lot of pond water in BF and DF. What I missed with that system was a low power scanning objective. Had I had the opportunity to acquire one, which objective would I have swapped for? I'm pretty sure I would have swapped for the 90X but after I had exhausted my viewing with dry objectives, would have finalized everything with 15X eyepieces , the oil immersion 90X 1.30 at 1350X and thus the widest field possible for that set up, never to go back to dry viewing for that sample. There was a nice 60X oil immersion for that system and I would gladly gave wanted that but try to find one. I have only ever seen a couple in many years of looking.
For anyone, if feeling the pinch on their nosepiece and if the instrument is equipped for it, try to outfit your scope with another nosepiece with a further hole, for all objective choices advance the study of pond water. I didn't have that option 40 years ago but would have grabbed it if I could have.
In the modern world there are other options too. In the modern W.F. eyepiece world, a pretty standard pair of 10X eyepieces are a 20mm f.n. Their compatible 15X counterparts are usually a 16mm f.n. This means that using a 60X objective with 10X F.N. eyepieces working at 600X yields a 333 micron field and a 40X objective with 15X eyepieces working at 600X yields a 417 micron field, with the working distance of a 40X objective. Although a .65 objective ( a real and good one) will give good results, a 40X .85 will give as good results as a 60X .85 objective plus there is the added advantage as well of adding 60X and 150X to your magnification capabilities, again both at wider fields than a 10X eyepiece would give you. You really can't use a 15X eyepiece with a 100X 1.25 objective. The only real drawback to this system is that usually the eye relief of 15X eyepieces is poorer than that of 10X. It can be but not always. It certainly wasn't the case using the pre-1955 Spencer compensating eyepieces, where the opposite is true and I haven't found it so with the AO cat. # 182 eyepieces made for the Microstar/Diastar, which is my principle microscope.
So, lets say you have a pretty common 4X .09, 10X .25, 40X .65 and 100X 1.25 equipped microscope. Swapping a 5 place nosepiece for the 4 allows you to add a 20X usually .45. , so you have 40X, 100X, 200X, 400x and 1000X, or if you are limited to a 4 place nosepiece you can swap a 20X in and out or replace it for another of the objectives.
Another option is to buy just the 15X eyepieces. This can be a bit tricky because there are various designs and the eyepieces need to be perfectly compatible with your objectives. It isn't unlike making sure you buy a compatible objective, so an eyepiece purchase takes some care. They are not that expensive, though. This adds essentially 2 free objectives to your system possibly 3. So, now you have 40X, 60X, 100X, 150X, 400X, and 1000X. There is a good possibility that you will get a good 600X image as well. Theoretically, you should but some 40X .65 objectives may not be up to it. The option still exists to upgrade your 40X to a higher N.A. 40X, which is usually not very expensive on the used market. That would ensure a good 600X dry image.

Lastly, there are now pretty decent options for water immersion 100X planachromats, both in the Olympus 180mm infinity corrected format N.A. 1.15 and the Nikon 200mm infinity format ( NIS) as either 45mm or 60mm parfocal at 1.10 N.A. I have found the NIS45 objective to be perfectly compatible with Reichert Austria infinity objectives. This a good option for those who have infinity microscopes of any of those types or similar stencil brands who for one reason or another don't want to get into oil immersion. The NIS45 objective is an excellent well built planachromat. They are in the $300.00 + range. I have been fleetingly working on a drop in DF stop for the NIS45 objective but at a native N.A. of 1.10, it needs little reduction in order to achieve DF.
P.M. me if you are interested in one of those.

[Polymerase]

Very often, inexperienced microscopists tend to think that it is all about the high power objectives and high magnification.
When starting histo, our professor started by stating that “you”ll be surprised how little time is spent on high power objectives.” While this is (mostly) true for histology sections, I am probably not the right person to proclaim how pond organisms are best viewed - but I firmly believe that most work on such specimens should pe performed in the low magnification range until something very specific turns up, that need the detail of higher resolution power.

As such, I would say the 4x, 10x and 20x objectives would be most useful. I’d rather spend money on higher n.a. low power objectives than going for the full range, if your budget demands choosing. 40x and above are a bit more of a hassle (especially planapos), but can be rewarding when great detail is required. And don’t be afraid of immersion objectives! A 50x or 60x oil is a good supplement to a 100x oil, allowing to switch back to lower magnification while using oil (do NOT forget to oil the condenser!)

Generally speaking, there is no such thing as a “best” objective, neither in quality, nor in magnification. It all boils down to what you are trying to achieve. What do you wish to study? What do you need to find out? If you can answer these questions, then asky ourself: “What kind of objective do I need to accomplish what I wish to study?” You may also ask which modality will yield the most useful results. Brightfield has limitations in living specimens. Phase contrast or darkfield may be of use, and may affect which objectives need to be obtained. DIC seems very glorified in the community. A wonderful modality, but very expensive. Is it worth it? What do you achieve, that phase contrast can’t give you?
Could staining specimens alleviate the need for costly equipment? For living specimens, maybe you would be better off with an inverted scope?

Systematic approaches yield the best results. Know what your intentions are. Then find out what equipment helps you get the results you desire.

Don’t forget that great detail and wonderful pictures can be produced with cheap achromats in the hands of a skillful microscopist. Your equipment doesn’t give you anything for free. Some things even get more difficult with more expensive and sophisticated systems.

Less is almost always more!

[apochronaut]

What about the myriads of critters under 10 microns not to mention their structures or even the structures of larger critters?

[Alexander]

What about the myriads of critters under 10 microns ....?

Most of them are so fast they pass the viewing field of a 40x within the fraction of a second. Not enough time to study any details.

[Polymerase]

What about the myriads of critters under 10 microns not to mention their structures or even the structures of larger critters?

A good question. However, viewing such creatures in a live specimen puts a heavy demand on slide preparation. Again, knowing what you intend to study is crucial for selecting which objectives to equip your scope with. There is no such thing as a “best size objective.” As a general advice, the lower power objectives will be the most rewarding for those not very experienced. That does not exclude the use of high power objectives for studying smaller organisms, or fine detail of the larger ones.

Being among those who prefer dead, fixed and stained samples, I may not be the one to pay the most attention to.

[Polymerase]

What about the myriads of critters under 10 microns not to mention their structures or even the structures of larger critters?

A good question. However, viewing such creatures in a live specimen puts a heavy demand on slide preparation. Again, knowing what you intend to study is crucial for selecting which objectives to equip your scope with. There is no such thing as a “best size objective.” As a general advice, the lower power objectives will be the most rewarding for those not very experienced. That does not exclude the use of high power objectives for studying smaller organisms, or fine detail of the larger ones.

Being among those who prefer dead, fixed and stained samples, I may not be the one to pay the most attention to.

On second thought: 10 microns is about the size of a lymphocyte. A 20x should let you observe those, although not in great detail.

[Javier]

Absurdly, one of the reasons some people avoid oil immersion is because they are too cheap to pay for the oil?!

In my case, not using the 100 x oil objective is a matter optical quality. I'm happy with the optical quality of my DIN achromatic objectives up to 40 x given the cost of my microscope, but the 100 x it is just not up to the task, even for a casual amateur like me. It is no secret that high resolution images are quite demanding in terms of optical quality and mechanical tolerances. In my opinion, beginners microscopes would be far more useful combining a 4, 10, 20 and 40 x in a 4 objectives turret.

[apochronaut]

" not in great detail " and I said under 10 microns . You must have been using a 20X .45 objective a lot .
Have a look at any protozoology text. Richerd Kudo's for example. All objectives were used and I doubt if any of them was a 20X .45 achromat because that is what most amateurs will be reaching for. The ex- gene sequencing M25 thread 20X .75 N.A. Nikon isn't going to be the tool of choice for too many of the curious readers of this thread.
Limiting yourself to 4X, 10X and 20X achromats with a maximum N.A. of .45 might be o.k. if you just want to sit and observe moving bodies but there is a whole lot more going on in an active slide of pond water than that and those objectives with higher N.A.'s give you a chance to explore that. Without N.A. , all you will see is " so many of them passing through the viewing field " to quote Alexander . Learning more precise technique and how to utilize all of your microscope is the key to becoming a microscopist and enjoying the instrument fully, not just looking at a screen of swarming blobs like a stoner staring at a fish tank.

[apochronaut]

Absurdly, one of the reasons some people avoid oil immersion is because they are too cheap to pay for the oil?!

In my case, not using the 100 x oil objective is a matter optical quality. I'm happy with the optical quality of my DIN achromatic objectives up to 40 x given the cost of my microscope, but the 100 x it is just not up to the task, even for a casual amateur like me. It is no secret that high resolution images are quite demanding in terms of optical quality and mechanical tolerances. In my opinion, beginners microscopes would be far more useful combining a 4, 10, 20 and 40 x in a 4 objectives turret.

If your 1.25 achromat isn't achieving it's stated specification, then it is likely one of the many poor examples that exist. Get another one. I realize that making that leap to fully utilizing a microscope is a big leap and that the optical quality of the lower N.A. objectives seems better than a poorly prepared fresh mount at high magnification but the opposite is in fact the case if the higher N.A. objectives are used according to their requirements. It's a skill worth learning.

[deBult]

When (ever, not that often see above) using a high NA or 100* oiled objective with a wet sample mount: I often use a number 0 [correction # 1] cover glass replacing the standard (Zeiss) 1.5 to compensate for the sample thickness.

As stated by Apo above when moving into the extreme magnification / large NA field good slide preparation is a must, the cheaper coverglasses are often not up to the task due to variation in thickness.

[Polymerase]

" not in great detail " and I said under 10 microns . You must have been using a 20X .45 objective a lot .
Have a look at any protozoology text. Richerd Kudo's for example. All objectives were used and I doubt if any of them was a 20X .45 achromat because that is what most amateurs will be reaching for. The ex- gene sequencing M25 thread 20X .75 N.A. Nikon isn't going to be the tool of choice for too many of the curious readers of this thread.
Limiting yourself to 4X, 10X and 20X achromats with a maximum N.A. of .45 might be o.k. if you just want to sit and observe moving bodies but there is a whole lot more going on in an active slide of pond water than that and those objectives with higher N.A.'s give you a chance to explore that. Without N.A. , all you will see is " so many of them passing through the viewing field " to quote Alexander . Learning more precise technique and how to utilize all of your microscope is the key to becoming a microscopist and enjoying the instrument fully, not just looking at a screen of swarming blobs like a stoner staring at a fish tank.

I do perfectly agree with you, and as stated, my experience is not in living pond samples. And I am spoiled objective-wise, as my primary 20x’es are of an n.a. of 0.46 and 0.70.
For the beginner, however, I believe higher power would yield a lot of frustration. Not to say one shouldn’t try, but if planning your purchases on a tight budget, I would prioritize lower power objectives with high n.a.

On the other hand, learning to use high power objectives is very rewarding when you get the grip of it. But it requires not only skill in using the objectives, but also in specimen preparation. I find it sad that people own fine instruments, but cannot take advantage of their full capabilities. Many 100x objectives are hardly used because one does not know how to use them properly.

I stand corrected by the way. Your knowledge far exceeds mine both in optics generally and in pond life samples specifically.

[Javier]

Absurdly, one of the reasons some people avoid oil immersion is because they are too cheap to pay for the oil?!

In my case, not using the 100 x oil objective is a matter optical quality. I'm happy with the optical quality of my DIN achromatic objectives up to 40 x given the cost of my microscope, but the 100 x it is just not up to the task, even for a casual amateur like me. It is no secret that high resolution images are quite demanding in terms of optical quality and mechanical tolerances. In my opinion, beginners microscopes would be far more useful combining a 4, 10, 20 and 40 x in a 4 objectives turret.

If your 1.25 achromat isn't achieving it's stated specification, then it is likely one of the many poor examples that exist. Get another one. I realize that making that leap to fully utilizing a microscope is a big leap and that the optical quality of the lower N.A. objectives seems better than a poorly prepared fresh mount at high magnification but the opposite is in fact the case if the higher N.A. objectives are used according to their requirements. It's a skill worth learning.

Doesn't a 100 x requires an extremely precise instrument in every aspect? I guess tolerances are extremely tight at that point. In my mind, until I get a far better microscope (not possible right now), getting another 100 x is not worthy, even considering that after a couple of years of doing microscopy my mount preparation skills are quite acceptable.

[apochronaut]

Not really. If you have a fine focus, a condenser and a mechanical stage, you should be able to use a 100X immersion objective quite easily. Failures are almost always a case of poor technique unless the objective is defective.
The biggest mistake people make is once they put oil on the coverglass , they focus down into the oil. What you do is bring the objective into the oil while looking from the side until the objective touches the cover, then observe while focusing slowly up. Once you gain focus, you only need a little tweaking wiyh the fine focus.

[Chas]

A lot of interesting things live on or around duckweed (lemna) so it is easy to drop a plant or two under a coverslip ..however this sort of deep/thick mounting makes a 'dangerous' combination with an x40 objective.
So I would go for (with a triple nose piece) 4x 10x 20x .

[charlie g]

Hi, all, many non-mobile, or low mobile ( diatoms, amoebae, etc.) are fascinating targets for : 60X, 100X oil-objectives.

For those active protozoans..learn to use methylcellulose 1.5%/ half pondwater/half this methylcellulose viscosity tool. We need to learn how to keep the water column/ water film thickness below the cover slip..in wetmount slide preps to the 'sweet spot minimal thickness'...please consider and try this: 'technique of wet mount slide setup'...before waxing sentiments on how wonderful living aquatic pond life enjoyment stops at a 20X, or 40X dry objective.

The use of proper: Kohler-illumination will enhance light microscopy observations..but simple wetmount slide usage ( large rectangular cover slips...say 22mmX44mm, No.1 slips..so you only cruise within the slides middle 80% of terrain...far from the coverslip border waters)...with use of viscosity tool, if you desire observations of active protozoans...this protocol is easy...low learning curve...try it....try it rather than opining on: 'no can do'.

It's odd to have pond life microscopy hobby...and say: ' not possible above 40X dry objective'...learn to enjoy your light microscopy of vibrant living assemblages..with all your objectives..just utilize the simple protocols to use your ''tediously close working distance " higher mag/ higher NA objectives. A lot of us learned how to paralell park in crowded urban settings...this wet mount live organism microscopy is so much easier to employ.

Please try it. charlie guevara/ finger lakes-USA

[deBult]

Not sure how we all ended up here : this is the beginners corner and a beginners question.

Naturally: with proper technique and practice you can use your higher Mag. and NA objectives: don’t see above discussion is helpful to a beginner.