There are a variety of different factors that determine the achievable resolution. Some of these factors can not be actively influenced by the microscopist, others can. Some of the factors play a larger role, others a smaller one. In the following post, I want to summarize some of these factors.

Objective-related factors

• Correction of lens errors: In contrast to achromatic objectives, apochromatic objectives focus more colors of the spectrum to one point. This results in a sharper image.
• The numerical aperture of the objective: This value is printed on the objective. The higher the value, the higher the resolution. The numerical aperture is a dimension less value which represents the cone of light that can be caught by the objective.

Lighting system

• General color of light: The shorter the wavelength, the higher the resolution. If your microscope uses halogen or tungsten lamps (instead of LEDs), then the color of the light will shift towards the red end of the spectrum with increasing age. This will reduce the resolution. The color of the light also changes with its intensity. If you turn up the light to maximum intensity, then the color of the light will be more towards the blue end of the spectrum (shorter wavelength and higher resolution). LEDs do not change their color with age or brightness.
• Light spectrum (color range): The color range may also impact on resolution. In the case of monochromatic light, chromatic aberration does not play a role and the light can be focused on one point.

Specimen-related factors

• The correct thickness of the cover glass: The correct cover glass thickness is extremely important for high numerical-aperture objectives. For other objectives, the effect may not be noticeable.
• The correct refractive index of the cover glass: This is something that you do not have to worry about, this is the task of the cover glass manufacturer.
• The correct refractive index of the mounting medium: This one should be as close to the refractive index of glass as possible.
• Thickness of the mounting medium: the thinner the better.
• The presence of immersion oil: Objectives that carry the label “OIL” need the correct immersion oil for best resolution.

Adjustments of the microscope

• The correct condenser diaphragm setting: This setting must match the numerical aperture of the microscope in use.
• The correct setting of the correction collar: Some objectives have a correction collar (a turnable ring) to adjust to the cover glass thickness. Most objectives do not have one, however.

Maintenance-related factors

• The cleanness of the optical parts: Dust and dirt generally decrease image quality and are a big annoyance, especially if one uses dark-field microscopy.

Stability of the photomicrographic system

• Moving objects: Moving cells naturally cause a blurring when long exposure times are used. This decreases resolution of the moving object.
• Stability: A shaky photographic system generally decreases resolution of the image.

The checlkist: how to obtain the best image quality

• Use new light bulbs and turn up the light. This will reduce the wavelength of the light. Alternatively, use a blue filter.
• Use cover glasses of the correct thickness and make sure that the mounting medium has a refractive index which is close to the refractive index of glass.
• Adjust the condenser aperture diaphragm to the numerical aperture of the objective
• If you use oil immersion, make sure that the oil has the correct refractive index
• Use fresh light bulbs (low in red light, high in blue light)
• Keep the microscope free of dust
• Make sure that the objectives, eye pieces are clean

Stained nuclei of plant cells. Check if the optics deliver a crisp picture of high contrast. Is dust visible?

Before buying a second hand microscope, take care of the following points. The list is certainly not complete, but should give an overview of the things to look out for:

• Appropriate optics: Make sure that the microscope is equipped with the objectives that are needed for your task. Many microscopes from hospitals and research institutes are equipped with phase contrast or plan apochomatic objectives. These are expensive and not required for educational work.
• Are the focusing knobs easy to turn or has the lubrication oil already solidified? Do not force-turn the focus knobs, it will increase the wear on the gears.
• Does the condenser move freely up and down, or has the lubrication oil already solidified? Does the condenser stay up, or is it pulled downwards by its own weight?
• Does the iris diaphragm open and close without problems, or has the lubrication oil already solidified?
• Were the non-oil immersion objectives dipped into immersion oil? They are not designed for this.
• If the objectives are spring-loaded, does the front part of the objective retract properly when pushed in? Or was the objective covered completely with immersion oil, now solidified?
• Are there any fungi growing on the lens optics? This may be a problem of microscopes used in warm and humid areas.
• Does the stage stay where it is, or does it move down due to its own weight?
• Does the microscope generally make a good impression?

Additional points to consider for schools:

• If you want to equip a whole classroom with used (or new) microscopes, make sure that they are from the same series. Students are then confronted with the same device each time and do not have to re-learn the peculiarities of each instrument – more productive lab work. It also makes makes it easier for the teacher to explain the handling of the microscope. If you buy used instruments, then you may not be able to obtain a whole classroom set at once (unless another school, college or hospital replaces its equipment at once).
• Make sure that the used microscopes have the appropriate optics installed and not specialized objectives! You need achromatic bright-field objectives. Many research microscopes come with phase contrast objectives installed, however. This type of optics also can be made to work like bright-field objectives but they are more expensive and you don’t want to spend money on things that you don’t need.
• In recent years the so-called infinity optics became increasingly popular, especially in research. Be careful – infinity objectives are not compatible with the “traditional” DIN 160mm systems, and infinity optics from different manufacturers are also not compatible with each other. When purchasing infinity systems (if you can afford them…), be aware that the optics can possibly not be exchanged with other microscopes that you or your school owns.

A polyp. Inverted colors, not darkfield.

Occasionally people ask me for advice about which type of microscope to buy as a present for their children. I once responded to a newsgroup question making a very strong point in favor for stereo microscopes for young children (approx. 5 years of age), and I would like to reiterate these points below. Read the article “Different Types of Light Microscopes” for a description of similarities and differences between the different microscope types. The following section reflects my own personal opinion on this issue.

In any case, I do not recommend the purchase of “toy” microscopes. If you invest a little more you are able to obtain a “real” instrument with substantially better image quality and flexibility, one which will retain the interest of the child (and parent!!) for a longer time. And especially for children a good image quality is necessary. An experienced microscopist may be able to interpret the “dark washed-out blob” as a cell, but children need crisper and clearer images to maintain their fascination – my personal opinion. There is the danger of disappointment if they do not see similar images as those printed on the box, and I am almost certain that many “toy microscopes” are not capable of keeping their promise. But this is my personal opinion, and the quality of these devices certainly varies as well.

Some of these microscopes are also sold with unrealistic magnifications up to over 1000x. Please understand that toy microscopes are useless at this magnification, for a range of reasons:

• Resolution is too low: The object that you want to see is magnified 1000x for sure, but you only see a washed-out blob with no detail.
• Stability is low:. There is a good reason why microscopes are made of metal and why they are heavy. Every vibration (walking) is magnified as well and transferred to the microscope.
• Image is dark: A high magnification requires a high light intensity. Many of these microscopes are not capable of delivering the required light intensity.
• And: bad depth of field, optics not corrected for lens errors, etc. etc.

I have to admit that even “toy” microscopes vary greatly in quality (and price). If you want to buy one of these, then I would recommend you not to give magnifications above 200x or 400x much weight and to read appropriate reviews beforehand. A cheap plastic scope with 1000x magnification is unrealistic. I have already seen some better quality “toy” microscopes but the price difference to a microscope manufactured according to the international DIN standard was not too big. The bottom line is that the child should enjoy working with the instrument.

Compound or Stereo Microscope?

Instead of simply listing the pros and cons of each type, I’ll make life easy by giving you two simple rules:

The younger the child the more you should tend towards stereo microscopes.

and:

If you intend to purchase a compound microscope, make sure that it works with the DIN standard. This allows for an exchange of objectives and guarantees a minimum quality.

There are many points that speak for stereo microscopes for young children, they are not only more “child friendly”, and more forgiving and easier to handle:

• The subjective visual impression of 3D samples (flies, hair, rocks etc.) can be quite fascinating. The view is, in contrast to compound microscopes, upright (!). A big advantage for orientation.
• Little to no sample preparation required for many objects. There is no need to cut and slice the specimens into the required thickness, even though this is possible as well. There is no need to prepare specimen slides with cover-slips. We use stereo microscopes in our school, and as a first introduction, we gave our students a post card and made them look at the colored dots that compose the image – fast, simple with an immediate result.
• Stereo microscopes have a low magnification, often not more than 40x. This means that there is less abstraction “from the real world”. A fly looks like a fly, only much bigger and more impressive. For a compound microscope you need to take the fly apart first and examine the individual parts, it’s too thick otherwise to be observed.
• Stereo microscopes also allow for an observation of non-transparent objects like rocks, fingernails (the dirt is pretty interesting…), skin, plant leaves etc. Stick a whole earth worm under the microscope and see how it looks like. Directly observe a dish of pond-water. If the child is already collecting rocks, insects, stamps, coins, etc. then a stereo microscope is the natural extension to observe these collected items.
• Some stereo microscopes also allow for a change in magnification, by zooming. This is not a necessity, though.
• Decent stereo microscopes can be cheaper than compound microscopes, because they are less complex.
• Stereo microscopes need less time for instruction. More instruction time needed for compound microscope. With compound microscopes, if you use a higher magnification and then turn the coarse-focus-adjustment knob into the wrong direction, you run the risk of ruining both sample and objective because you smash the objective into the specimen. Stereo microscopes have a large sample-objective distance.
• Stereo microscopes have a higher depth of field. It is therefore much easier to find what you are looking for.

Stereo microscopes also possess certain disadvantages:

• There may be some samples that you or your child is interested in but requires a higher magnification. For example, if you want to watch the nucleus of cells, then you are better off with a compound microscope. It is also not possible to observe bacteria, they are simply too small. As a comfort, a compound microscope with regular bright-field optics also does not allow you to view living bacteria, as they are transparent, you need expensive phase-contrast objectives (labs use them). Alternatively the bacteria have to be stained first, and then I doubt that novices will be able to recognize them as bacteria.
• Sample preparation may indeed be one of the activities that a child may be interested in, but stereo-microscopes do not require much preparation, while it is necessary in compound microscopes.
• Some younger children may have problems viewing through both eye-pieces (they can look through one of them if they want to).
• And possibly the biggest “problem”: There is the myth that microscopes have to magnify very much in order for the person to see much. Children may be disappointed if they hear that their stereo microscope only magnifies up to 40x, if their friend has a department store (“toy”) microscope which magnifies 1250x. This is where education comes into play – magnification is not everything, and a high magnification does not mean that one sees more, resolution also counts. I want to guarantee you that you won’t be able to see much at 1250x.