Light then passes up through the slide and into the objective lens where the first magnification of the image takes place. Magnification increases the apparent size of an object. In the compound light microscope two lenses, one near the stage called the objective lens and another in the eyepiece, enlarge the sample. The magnifying power of an objective lens is engraved in the lens mount. Microscopes in most microbiology laboratories have three objective lenses: the low power objective lens (10X), the high-dry objective lens (40X) and the oil-immersion objective lens (100X). The desired objective lens is rotated into working position by means of a revolving nosepiece.

F stops generally range from f/1.4 (let in lots of lot for darker areas) to all the way to f/22 (let in little light for bright areas). A wider aperture will keep less of the scene in focus. A narrower aperture will give a crisp focus to more of the scene. Portrait photographers prefer wider apertures like f/2.8 or even f/4 — they can focus on the subject and blur the background.

That’s also why landscape photographers typically shoot in the f/11 to f/22 range — they want more of the landscape in focus, from the foreground to the distant horizon. Take a look at the graphic below to see it in action.

F number camerafor beginners

A couple of important things happen when you change your f-stop. First of all, a wider aperture (think f/1.4 to f/2.8) will let a lot more light in through the lens and on to the sensor. This allows you to shoot with a much faster shutter speed. A narrower aperture (think f/16 to f/22) will let in much less light and require a slower shutter speed.

First let us consider a primary feature of all microscopes, the light source. Proper illumination is essential for effective use of a microscope. A tungsten filament lamp usually serves as the source of illumination. If reflected illumination is used, a separate lamp provides a focused beam of light which is reflected upward through the condenser lenses by a mirror.

F-stops may seem a bit counter-intuitive to learn, but once you get the hang of it, you’ll know it for life. The key takeaway is this: the smaller the number, the bigger the physical opening of the aperture. Conversely, the larger the number, the smaller the physical opening. So f/1.4 is a very wide opening (or larger aperture) while f/22 is a much smaller opening (or smaller aperture).

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When the camera is in Auto mode, it automatically decides the aperture and other settings based on the available light. If you want to set shutter speed and f-stop (aperture), the camera has to be in Aperture mode or Manual mode.

f-number formula

If you really want to blur the background for macro or wildlife shots, you’ll need to look for a lens with a maximum aperture of f/2.8 or even f/1.4. If you’re shopping for a lens that zooms, it’s normal for the lens to have a wider maximum aperture when it’s zoomed out than when it zooms in. That’s why some lenses have two apertures listed in their titles, such as 18-55mm f/3.5-5.6.

On both sides of the base of the microscope are the course and fine adjustment knobs, used to bring the image into focus. Rotation of these knobs will either move the specimen and the objectives closer or farther apart. The coarse adjustment moves the nosepiece in large increments and brings the specimen into approximate focus. The fine adjustment moves the nosepiece more slowly for precise final focusing. In some microscopes, rotation of the fine and course adjustment knobs will move the stage instead of the nosepiece.

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The compound microscope used in microbiology is a precision instrument; its mechanical parts, such as the calibrated mechanical stage and the adjustment knobs, are easily damaged, and all lenses, particularly the oil immersion objective, are delicate and expensive. Handle the instrument with care and keep it clean.

When the oil-immersion objective lens is in use, the difference between the light-bending ability (or refractive index of the medium holding the sample) and the objective lens becomes important. Because the refractive index of air is less than that of glass, light rays are bent or refracted as they pass from the microscope slide into the air, as shown in Figure 3-9. Many of these light rays are refracted at so great an angle that they completely miss the objective lens. This loss of light is so severe that images are significantly degraded. Placing a drop of immersion oil, which has a refractive index similar to glass, between the slide and the objective lens decreases this refraction, and increases the amount of light passing from the specimen into the objective lens. This results in greater resolution and a clearer image.

The image of the specimen continues on through a series of mirrors and/or prisms that bend it toward the eyepiece. A further magnification takes place at the eyepiece producing what is called a virtual image. Total magnification is equal to the product of the eyepiece magnification and the objective magnification. Most often eyepiece lenses magnify 10-fold resulting in total magnifications of 100, 400, or 1000X, depending upon which objective is in place. Many modern microscopes will also have focusable eyepieces to compensate for differences between individuals and even between individual's eyes. The adjustment of these is important and is described below.

You can also set the aperture manually and allows the camera to decide the best shutter speed for the available lighting conditions. To use a fixed f stop, the camera has to be in the Aperture Priority mode.

The same wheel or slider controls shutter speed and aperture on most cameras. To change the aperture, you have to hold down the AV button and then adjust the wheel. Both the AV button and aperture/shutter speed control wheel are on the top-right of the camera. You have a wheel for both aperture and shutter speed on more advanced cameras.

F-stop vs aperture

We’ve already referred to the lens opening as “f-stop” and “aperture,” but it’s also sometimes called the “f number” or “f-stop number.” When shopping for a new lens, you’ll often see apertures referred to as “f/” with a number. For instance, if you see “f/1.4” or “f/5.6,” those numbers indicate the diameter of the aperture.

Magnification alone is not the only aim of a microscope. A given picture may be faithfully enlarged without showing any increase in detail. The true measure of a microscope is its resolving power. The resolving power of the lens is its ability to reveal fine detail and to make small objects clearly visible. It is measured in terms of the smallest distance between two points or lines where they are visible as separate entities instead of one blurred image. The resolving power of the objective lens, engraved on the lens, allows us to predict which objective lens should be used for observing a given specimen. However, having good resolution in the microscope does not guarantee a visible image, the resolving power of the human eye is quite limited. Often further magnification is needed to obtain a good image.

F-number lens

Aperture also influences the depth of field, determining how much of the image will be in focus. Small apertures produce a deep depth of field and allow you to have the entire frame in focus. Large apertures produce a shallow depth of field and let you blur a busy background and make the subject more visible.

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The microscope, as shown in Figure 3-1, is one of the most important instruments utilized by the microbiologist. In order to study the morphological and staining characteristics of microorganisms such as bacteria, yeasts, molds, algae and protozoa, you must be able to use a microscope correctly.

When it comes to photography, there are a lot of new terms to learn: exposure (internal redirect), shutter speed (internal redirect), ISO, and f-stop. Some of these terms are more self explanatory than others. For many beginners, f-stop is one of the trickier terms in their new hobby, so let’s go back to basics and give you a solid understanding of f-stops.

The light from the illuminating source is passed through the substage condenser. The condenser serves two purposes; it regulates the amount of light reaching the specimen and it focuses the light coming from the light source. As the magnification of the objective lens increases, more light is needed. The iris diaphragm (located in the condenser), regulates the amount of light reaching the specimen. The condenser also collects the broad bundle of light produced by the light source and focuses it on the small area of the specimen that is under observation.

Before the light from the scene can hit your digital camera’s sensor and help create an exposure, it must first travel through the camera lens. The amount of light that reaches the sensor depends on your lens and the settings you’ve chosen.

F-stop, or aperture, specifically controls the size of the opening in the lens. With a larger opening (or a smaller f-stop number) you’ll allow more light to enter. With a smaller opening (or a larger f-stop number) you’ll allow less light to enter. By controlling the amount of light entering the lens, you’re also controlling the overall exposure of the image. Images with a wider aperture opening will be more prone to overexposure, while images with a narrower aperture opening will be more prone to underexposure. You’ll need to adjust your shutter speed or ISO accordingly to balance out changes in f-stop and to get the ideal exposure.

F-stop chart

Every lens has what’s called a maximum aperture, meaning the widest the aperture can possibly go. Traditionally, less expensive kit lenses don’t feature the wider range of apertures. They might only go as wide as f/4 or even f/5.6.

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Aperture controls the amount of light that reaches the sensor. Together with shutter speed and ISO, the aperture controls the efficient use of available light and the luminosity of the image. When the aperture is wide open (small f-stop), more light reaches the sensor and creates a brighter image. When the aperture is narrow (large f-stop), less light reaches the sensor and creates a darker image.

Figure 3.1. The light microscope. A modern light microscope. This is an example of the kind used in the teaching labs at the University of Wisconsin-Madison. The various parts of the microscope are labeled. Please take the time to become familiar with their names.

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Figure 3.9. Refraction of light at 100X. Light passing out of the slide, into the air, toward the objective lens is refracted, due to the different in refractive index between air and glass. While the bending cause by this difference is not important at 100X and 400X, at 1000X this refraction is problematic, causing blurring of the image and significant loss of light. Immersion oil has a refractive index very similar to that of glass. Placement of a drop of oil between the objective lens and the slide prevents the bending of light rays and clarifies the image. The blue dashed line represents a potential light ray if immersion oil is not present. The red dashed line represents a light ray if immersion oil is present.

The lowest f-stop available is f/0.7. It belongs to Zeiss Planar 50mm, a lens used by NASA to film the moon, and Stanley Kubrick to make a film at candlelight. However, most photo lenses go as low as f/2, fewer go to f/1.2, and only a few reach f/0.95.

Fstop symbol

The largest f-stop available is f/45. It belongs to Sigma 105mm f/2.8 EX DG Macro and provides the maximum depth of field possible for a macro lens. There is also an f/40 aperture in Itorex 50mm, a lens that recreates the pinhole lens effect. Most commercial lenses have the minimum aperture at f/32 or f/16.

Generally, subjects such as macro, which value heavily blurred backgrounds, are best shot with the widest apertures (f/1.4-f/4). This ensures that all elements of the scene are in focus, from the foreground to the distant background.Practice working in these general f-stop ranges until you get more comfortable with the available apertures on each of your lenses. Then you can experiment to get exactly the look you’re going for!

Figure 3.8. The path of light through a microscope. Modern microscopes are complex precision instruments. Light, originating in the light source (1), is focused by the condensor (2) onto the specimin (3). The light then enters the objective lens (4) and the image is magnified. Light then passes through a series of glass prisms and mirrors, eventually entering the eyepiece (5) where is it further magnified, finally reacing the eye.

f-number calculator

Learning the parts of your camera and how to use them is the most important step in your photography journey. Experiment with settings, shots, and styles, and you’ll soon be mastering your new hobby.

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The f-stop refers to the aperture opening of the lens through which light can pass to the sensor or film. By looking at the current f-stop listed, you can quickly get an idea of how much light you’re allowing in, and how much depth of field you will have in your image.

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The microscope is basically an optical system (for magnification) and an illumination system (to make the specimen visible). To help understand the function of the various parts of the microscope, we will follow a ray of light as it works its way through a microscope from the light source, through the lenses, up to the eye. Figure 3-8 traces the path of light through the parts of the microscope

Inside every lens is a mechanical aperture. Consisting of several connected blades (generally between six and nine), the aperture opens and closes to let in more or less light. If you look at older manual aperture lenses, you can see the aperture opening and closing with a twist of a dial. In modern lenses, you change the aperture by adjusting the settings in your camera body.

Figure 3.1. The light microscope. A modern light microscope. This is an example of the kind used in the teaching labs at the University of Wisconsin-Madison. The various parts of the microscope are labeled. Please take the time to become familiar with their names.