If you’re new to the world of microscopy, it can feel as though there are a lot of things to learn. You may have already purchased your microscope, but when it comes to learning advanced topics, the path forward may not seem clear. One great place to start would be learning more about magnification. After all, magnification is obviously an important topic given the subject matter. Read below for a few pointers on where to get started with advanced magnification knowledge.
How it Works
Before you can learn advanced topics in magnification, let’s quickly review how it works. A simple microscope or magnifying glass (lens) produces an image of the object upon which the microscope or magnifying glass is focused. Simple magnifier lenses are bi-convex, meaning they are thicker at the center than at the periphery. Light from an object that is very far away from the front of a convex lens will be brought to a focus at a fixed point behind the lens. This is known as the focal point of the lens. The first lens of a microscope is the one closest to the object being examined and, for this reason, is called the objective. Light from either an external or internal (within the microscope body) source is first passed through the substage condenser, which forms a well-defined light cone that is concentrated onto the object (specimen). Light passes through the specimen and into the objective (similar to the projection lens of the projector described above), which then projects a real, inverted, and magnified image of the specimen to a fixed plane within the microscope that is termed the intermediate image plane. A fun exercise to teach this concept is to have students examine printed text on paper through the microscope. They’ll find that the letters that they are familiar with appear upside down in the microscope, and it’s a great way to orient and give a sense of scale to the specimen.
Now that we’ve covered the basics, let’s talk about resolution. When you’re looking at your current microscope or buying a new one, what does the description mean when it says, for example, a microscope lens has a resolution of 400x? It’s important to note that a microscope’s total magnification is a combination of the eyepieces and the objective lens. In our example, a 400x magnification could come from a microscope with 10x eyepieces and 40x objectives. So more magnification means a better microscope, right? Not in all cases. There are limits to a microscope’s resolution.
Alas, your microscope is not all powerful: eventually you will reach a point called “empty magnification.” Empty magnification occurs when the image continues to be enlarged, but no additional detail is resolved. This is often the case when higher magnification eyepieces are used. In order to avoid empty magnification, there are a few simple steps that are helpful to follow.
You’ll need to pick the right microscope for the job. When selecting a combination of eyepieces and objective lenses, you’ll need to consider the numerical aperture of the objective lens. In short, the numerical aperture is the objective’s resolution, and each microscope has a minimum and maximum magnification necessary for the details in an image to be resolved. When you surpass the maximum magnification, it will result in empty magnification.
Another limitation to be aware of is known as airy disks. At very high magnifications with transmitted light, point objects are seen as fuzzy discs surrounded by diffraction rings. These are called Airy disks. The resolving power of a microscope is taken as the ability to distinguish between two closely spaced Airy disks (or, in other words the ability of the microscope to reveal adjacent structural detail as distinct and separate). It is these impacts of diffraction that limit the ability to resolve fine details. There is therefore a finite limit beyond which it is impossible to resolve separate points in the objective field, known as the diffraction limit. While there are techniques for surpassing the resolution limit, these topics are quite advanced and best suited for veteran microscope technicians.
If you have questions about which microscopes are best for your lab, contact our friendly sales team. We would be more than happy to assist you and your lab in finding the right microscope for the job.