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Microscopy: 1. Microscopes - uses, history and development - optical light and electron

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There are various sections to work through, after 1 they can be read and studied in any order.

Sub-index of biology notes: investigations using microscopes


(1) Microscopes - uses, history and development

 What is an optical light microscope? How is it constructed?

 How does a microscope work? How can we measure the size of a cell?

 What is the advantage of studying structures with a microscope?

 What do we mean by the resolution/resolving power of a microscope?

 What is the formula for magnification? How do you do magnification calculations?

What is the difference between a light microscope and an electron microscope? Which is the most powerful?

What do we use a microscope for in biology? and why is it such a useful investigative tool?

A microscope is an important instrument for studying cells e.g. the type of cell and the structure of cells.

Microscopes enable to see structures that we cannot see with the unaided naked eye.

Plant and animal cells can be studied in greater detail with a light microscope by magnifying the image.

Microscopes use a glass lens system to magnify images - with a bigger image you see more detail.

You can increase the resolution of an image by using more powerful and better quality lenses. Resolution means how good a microscope is at distinguishing between two points that are close together on an image. The higher the resolution the more clear is the image, especially when looking for fine details e.g. in a cell.

Microscopes enable you to see objects (like microorganisms) which you cannot see with the naked eye.

Microscopes using the visible part of the electromagnetic spectrum (visible light) were invented in the 16th century and the optical lens systems of light microscopes have been improved through the following centuries even until today.

With these microscopes, by passing light through a specimen up into a lens system, you can see individual cells and smaller details such as nuclei and mitochondria in all cells, and chloroplasts in plant cells.

Optical light microscopes

Light microscopes using visible light and lenses to form a magnified image of the object under investigation e.g. cells of plant or animal tissue. With a light microscope you can see individual cells and large subcellular structures like the nucleus, but not internal cell structures such as ribosomes or plasmids. The best light microscopes can give a magnification of 2000 times of a specimen's length.

The resulting image on a photographic plate, book or a computer screen is called a light micrograph.

Very high magnification is not possible with optical light microscopes. The limitation is due to the light gathering ability of the microscope and the short working distance of the lenses. This limits the total magnification of light microscope to about x 1500.

BUT, even with a high magnification, details may still not be that clear. The microscope must have a high resolving power - this is the resolution of the microscope. The resolving power is the smallest distance between two points that can be clearly distinguished. For optical light microscopes the best resolution is about 0.2 m (200 nm).

However, unlike electron microscopes (described below), light telescopes can be used t observe living cells.

Electron microscopes - a means of looking at cells in more detail

Changes in microscope technology have enabled us to see cells with more clarity and detail than in the past, including simple magnification calculations.

Over time the design and usefulness of microscopes has improved, particularly using new technology in the 20th century and into the 21st century.

In the 20th century, with advances in atomic physics, the electron microscope (EM) was invented in the 1930s which uses beams of electrons instead of visible light photons. Electron microscopes use beams of electrons instead of beams of visible light photons. They have a much greater magnifying power and resolving power than optical light microscopes - larger sharper images.

So, using an electron microscope, using electrons instead of light photons, you can form images of very small subcellular structures such as ribosomes, plasmids and the internal structure of mitochondria (site of respiration) and chloroplasts (site of photosynthesis), because they have a much higher resolving power, but they are much more expensive!

Electron microscope images have a higher resolution than light microscopes - a higher resolving power increases the distinction between points on an image, i.e. you get a much sharper image of the fine detail of cell structure.

Electron microscopes can produce much greater magnified images (compared to light microscopes) of up to ten million (109) times the real length of the specimen under investigation.

So electron microscopes are superior to optical light microscopes in terms of both magnification and resolution.

The resulting image on a photographic plate or a computer screen is called an electron micrograph.

Electrons do not form a colour spectrum, so all images are in black and white.

Electron microscopes can't be used to look at living cells - the electron beams would damage the function of living cells.

The transmission electron microscope (TEM) is particularly good at looking at very thin layers of biological materials e.g. a layer of cells and investigate in great detail the sub-cellular components of a cell e.g. plasmids and mitochondria and also viruses.

A transmission electron microscope is a large instrument, not very portable and very expensive. To prepare specimens for examination is a complicated process, and, unlike light microscopes, cannot be used to examine living tissue.

In a TEM, as the electron beam passes through the sample, some electrons are scattered and those that pass through are focussed by electromagnetic coils (instead of lenses) to produce an image on an electronic screen.

A TEM can examine very thin sections of cells up to a magnification of 106 (million x) and with a resolution of less than 1 nm (10-9 m). This is 200 x greater resolution than the best light microscopes.

The scanning electron microscope (SEM)

A scanning electron microscope works by bouncing beams of electrons off the surface of a specimen. The specimen must be first coated in an ultra-thin layer of a heavy metal like gold. The scattered electrons are again focussed by electromagnetic coils to produce an image on an electronic screen.

A SEM is used to produce images (micrographs) of the surface shape of structures of e.g. of individual cells or small organisms.

This has enabled the magnification produced by a microscope to be considerably increased to the point where you can see even smaller structures such as the internal detailed structure of mitochondria, chloroplasts and plasmids (hoops of DNA) so as to give a better understand of their structure and how their role in cell behaviour - in other words a powerful tool for better understanding how a cell works and the function of sub-cellular structures.

e.g. in animal cells

or plant cells

where you can observe the fine detailed complex internal structure of important subcellular structures such as mitochondria (where aerobic respiration takes place), chloroplasts (where photosynthesis takes place), ribosomes (where protein synthesis takes place) as well as the detailed structure of specialised (differentiated) cells e.g. red blood cells (oxygen carriers) or white blood cells (immune defence system). In other words, electron microscopes allow more detailed studies of some pretty important structures and their functions!

Mitochondria and ribosomes can only be adequately viewed using an electron microscope and the 3D structure of biological specimens requires the use of an scanning electron microscope.

Extra note on microscopy methods

There is a technique called 'super-resolved fluorescence microscopy' which allows a much higher resolution than normal light microscopy. Since this is based on light, it means you can study living cells, which you can't do with electron microscopes - electron beams kill cells!

e.g. the size and shape of cells and subcellular structures are important, they are also variable, and such differences can be important e.g.

The complexity of mitochondria can indicate how active a cell is.

You can measure the ratio of the area-volume of the cytoplasm to that of the cell's nucleus.  A high ratio of cytoplasmic area-volume : nucleus area-volume can show a cell is about to divide. A low ratio can indicate a cancer cell.

 

You need to be able to use the following microscope formula:

For any microscope: magnification = length of image / real length of object, and for light microscopes:

total magnifying power = magnification of object lens x magnification of objective lens

and with a variety of units e.g. micro, nano etc. as well as expressing small numbers in standard form!


Keywords, phrases and learning objectives for this part on the history of microscope development

Know that optical light microscopes are used in biology and have been very important in the history and development of biological science.

Know that the electron microscope is much more powerful than an optical telescope and can investigate cells in much more detail from the higher magnification to produce high quality micrographs.

Know the advantage of electron microscopes over optical light microscopes.

Know that the transmission electron microscope TEM and scanning electron microscope SEM are further improved technological developments in the science of microscopy.


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