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The Compound Microscope
There would be little to do in a microbiology laboratory without a microscope, because the objects of our attention (bacteria, fungi, and other single celled creatures) are otherwise too small to see. Microscopes are optical instruments that permit us to view the microbial world. Lenses produce the magnified images that allow us to visualize the form and structure of these tiniest of living beings.
To use this important piece of equipment properly, it is helpful to know how a microscope works. A good place to begin is to learn the name and function of all of the various parts, because when we talk about the ways to improve microscopic images, terms like “ocular lenses” and “condenser” always come up.
Based on the picture of the binocular, compound light microscope in Figure 1, match the name of the major part (listed below) with its location on the microscope, and give a very brief description of what each is used for:
|Ocular lenses||_________________________________||Locate the parts on the microscope that allow you to:|
|Stage and stage clips||_________________________________|
|Course and fine focus knobs||_________________________________|
With a bright-field microscope, images are formed as a result of the interplay between light waves, the object, and lenses. How images of biological objects are formed is actually more physics than biology. Since this isn’t a physics course, it’s more important to know how to create exceptional images of the object than it is to know precisely how those images are formed.
Light waves that pass through and interact with the object may speed up, slow down, or change direction as they travel through “media” (such as air, water, oil, cytoplasm, etc.) of different densities. For example, light passing through a thicker or denser part of a specimen (such as the nucleus of a cell) may be reflected or refracted (“bend” by changing speed or direction) more than those waves passing through a thinner part. This makes the thicker part appear darker in the image, while the thinner parts are lighter.
For a compound microscope, the optical path leading to a detectable image involves two lenses – the objective lens and the ocular lens. The objective lens magnifies the object and creates a real image, which will appear to be 4, 10, 40, or 100 times larger than the object actually is, depending on the lens used. The ocular lens further magnifies the real image by an additional factor of 10, to produce a vastly larger virtual image of the object when viewed by you.
Light from an illuminator (light source) below the stage is focused on the object by the condenser lens, which is located just below the stage and adjustable with the condenser adjustment knob. The condenser focuses light through the specimen to match the aperture of the objective lens above, as illustrated in Figure 2.
Appropriate use of the condenser, which on most microscopes includes an iris diaphragm, is essential in the quest for a perfect image. Raising the condenser to a position just below the stage creates a spotlight effect on the specimen, which is critical when higher magnification lenses with small apertures are in use. On the other hand, the condenser should be lowered when using the scanning and low power lenses because the apertures are much larger, and too much light can be blinding. For creating the best possible contrast in the image, the iris diaphragm can be opened to make the image brighter or closed to dim the light. These adjustments are subjective and should suit the preferences of the person viewing the image.
When the light waves that have interacted with the specimen are collected by the lenses and eventually get to your eye, the information is processed into dark and light and color, and the object becomes an image that you can see and think more about.
The microscope you’ll be using in lab has a compound system of lenses. The objective lens magnifies the object “X” number of times to create the real image, which is then magnified by the ocular lens an additional 10X in the virtual image. Therefore, the total magnification, or how much bigger the object will actually appear to you when you view it, can be determined by multiplying the magnification of the objective lens by 10.
The magnifying power of each lens is engraved on its surface, followed by an “X.” In the table below, find the magnification, and then calculate the total magnification for each of the four lenses on your microscope.
|Magnification of objective lens||Total magnification of viewed object|
|Low Power Lens|
|High Power Lens|
|Oil Immersion Lens|
Let’s say you wanted to look at cells of Bacillus cereus, which are rod-shaped cells that are about 4 µm long. If you were observing B. cereus with a microscope using the high power lens, how big would the cells appear to be when you look at them? ___________________________
HomeResolution Limits Magnification
So the microscope makes small cells look big. But why can’t we just use more or different lenses with greater magnifying power until the images we see are really, really big and easier to see?
The answer is resolution. Consider what happens when you try to magnify the fine print from a book with a magnifying glass. As you move the lens away from the print, it gets larger, right? But as you keep moving the lens, you notice that while the letters are still getting larger, they are becoming blurry and hard to read. This is referred to as “empty magnification” because the image is larger, but not clear enough to read. Empty magnification occurs when you exceed the resolving power of the lens.
Resolution is often thought of as how clearly the details in the image can be seen. By definition, resolution is the minimum distance between objects needed to be able to see them as two separate entities. It can also be thought of as the size of the smallest object that we can clearly see.
The ability of a lens to resolve detail is ultimately limited by diffraction of light waves, and therefore, the practical limit of resolution for most microscopes is about 0.2 µm. Therefore, it would not be practical to try to observe objects smaller than 0.2 µm with a standard optical microscope. In addition, cells of all types of organisms lack contrast because many cellular components refract light to a similar extent. This is especially true of bacteria. To overcome this problem and increase contrast, biological specimens may be stained with selective dyes.
HomeThe Oil Immersion Objective
The lens with highest magnifying power is the oil immersion lens, which achieves a total magnification of 1000X with a resolution of 0.2 µm. This lens deserves special attention, because without it our time in lab would be frustrating.
The resolving power of this lens is dependent on “immersing” it in a drop of oil, which prevents the loss of at least some of the image-forming light waves because of refraction. Refraction is a change in the direction of light waves due to an increase or decrease in the wave velocity, which typically occurs at the intersection between substances through which the light waves pass. This is a phenomenon you can see when you put a pencil in a glass of water. The pencil appears to “bend” at an angle where the air and water meet (see Figure 3). These two substances have different refractive indices, which means that light passing through the air reaches your eye before the light passing through the water. This makes the pencil appear “broken.”
The same thing happens as the light passes through the glass slide into the air space between the slide and the lens. The light will be refracted away from the lens aperture. To remedy this, we add a drop of oil to the slide and slip the oil immersion objective into it. Oil and glass have a similar refractive index, and therefore the light bends to a lesser degree and most of it enters the lens aperture to form the image.
It is important to remember that you must use a drop of oil whenever you use the oil immersion objective or you will not achieve maximum resolution with that lens. However, you should never use oil with any of the other objectives, and you should be diligent about wiping off the oil and cleaning all of your lenses each time you use your microscope, because the oil will damage the lenses and gum up other parts of the instrument if it is left in place.
HomeUsing the Microscope
If you are new to microscopy, you may initially feel challenged as you try to achieve high quality images of your specimens, particularly in the category of “Which lens should I use?” A simple rule is: the smaller the specimen, the higher the magnification. The smallest creatures we observe are bacteria, for which the average size is a few micrometers (μm). Other microscopic organisms such as fungi, algae, and protozoa are larger, and you may only need to use the high power objective to get a good view of these cells; in fact, using the oil immersion objective may provide you with less information because you will only be seeing a part of a cell.
This brings us to two additional concepts related to microscopy—working distance and parfocality. Working distance is how much space exists between the objective lens and the specimen on the slide. As you increase the magnification by changing to a higher power lens, the working distance decreases and you will see a much smaller slice of the specimen. Also, once you’ve focused on an object, you should not have to make any major adjustments when you switch lenses, because the lenses on your microscope are designed to be parfocal. This means that something you saw in focus with the low power objective should be nearly in focus when you switch to a high power objective, or vice versa. Thus, for viewing any object and regardless of what lens you will ultimately use to view it, the best practice is to first set the working distance with a lower power lens and adjust it to good focus using the coarse focus knob. From that point on, when you switch objectives, only a small amount of adjustment with the fine focus knob should be necessary.
Here is a final consideration related to objective lenses and magnification. Look at the lenses on your microscope, and note that as the magnification increases, the length of the lens increases and the lens aperture decreases in size. As a result, you will need to adjust your illumination to compensate for a darkening image. There are essentially three ways to vary the brightness; by increasing or decreasing the light intensity (using the on/off knob), by moving the condenser lens closer to or farther from the object using the condenser adjustment knob, and/or by opening/closing the iris diaphragm. Don’t be afraid to experiment to create the best image possible.
Guidelines for safe and effective use of a microscope:
1. Carry the microscope to your lab table using two hands, and set it down gently on the bench. Once placed on the bench, do not try to slide it around on its base, because this is extremely jarring to the optical system.
2. Clean all of the lenses with either lens paper or Kimwipes (NOT paper towels or nose tissues) BEFORE you use your microscope, AFTER you are done, and before you put it away.
3. When you are finished with the microscope, check the stage to make sure that you don’t leave a slide clipped in the stage. Make sure to switch the microscope OFF before you unplug it. Gently wrap the cord around the base and cover your microscope with its plastic cover.
4. Return the microscope to the cabinet before you leave the lab. Make sure that the ocular lenses are facing IN.
Together we will review how to effectively achieve an exceptional image using a standard optical microscope. This will include not only locating and focusing on the object, but also using the condenser lens and iris diaphragm to achieve a high degree of contrast and clarity.
We’ll start by looking at a prepared slide of a “rectal smear,” which is quite literally a smear of feces on a slide stained with a common method called the Gram stain. You will observe several different types of bacterial cells in this smear that will appear either pink or purple. While the main purpose of this is to develop proficiency in use of the oil immersion objective lens, it also provides the opportunity to look at bacteria, observe the differences in cell shapes and sizes, and note that when Gram stained they turn out to be either purple or pink.
When you have achieved an exceptional image of the fecal bacteria at 1000X, consider the following questions.
- In a single field, approximately what proportion of the bacterial cells are circular (the microbiology term for circular bacterial cells is “cocci”)? _____________________________
- Among the cells that are cocci, can you see any specific types of arrangements, like chains of cocci (called “streptococci”) or clusters (called “staphylococci”)? Sketch examples in the space below:
- In a single field, approximately what proportion of the bacterial cells are rod-like (the microbiology term for rod-shaped cells is “bacilli”)? _________________________________
- Among the cells that are bacilli, can you see any specific types of arrangements, like pairs (diplobacilli), chains (streptobacilli) or parallel clusters (palisades)? Sketch examples in the space below:
- Based on the shape/arrangement of the bacterial cells, and now including color (whether they are pink or purple), estimate how many different types of bacteria you are able to see in a single microscopic field. ___________________________________________________
HomeWhat’s in YOUR Mouth?
The human mouth is home to numerous microbes, which persist no matter how many times you brush your teeth and use mouthwash. Since these microbes generally inhabit the surface layers of the oral mucosa, we humans have evolved ways to keep their numbers under control, including producing antibacterial chemicals in saliva and constantly turning over the outer layer of epithelial cells that line the inside of the mouth.
Obtain a prepared slide labeled “mouth smear.” On this slide you will see large cells with a nucleus, clearly visible with both the low power and high power objective lenses. These are squamous epithelial cells that form the outermost layer of the oral mucosa. At high power, you should start to see small cells on the surface of the larger epithelial cells. With the oil immersion objective lens, you will be able to tell the smaller cells are bacteria.
Locate and focus on a single squamous epithelial cell with obvious bacteria on its surface. Create a sketch of the “cheek” cell (as squamous epithelial cells are sometimes called) in the circle provided. Then label the cell membrane, cytoplasm, and nucleus of the “cheek” cell, which should be easily observed.
Add to your illustration the bacterial cells which you should see on or near the larger larger cheek cells. Try to keep the size of the bacterial cells to scale with the size of the cheek cell.
- How would you describe the shape and arrangement of the bacterial cells (using the microbiology terms you used to describe the bacteria in the rectal smear)?
- The nucleus of a squamous epithelial cells is approximately 10 µm in size. Compare the size of the cell’s nucleus with the size of the bacterial cells. Based on this comparison, what is the approximate size of the bacterial cells in the image?
Once you’ve looked at the prepared slide, obtain a glass slide and a sterile swab. Collect a sample of your oral mucosa by gently rubbing the swab over the inside of your cheek. Smear the swab over the surface of the slide (this is known as making a “smear” in microbiology). Allow the smear to dry, and then heat fix by passing the slide through the flame of a Bunsen burner, as demonstrated. Discard the swab in the biohazard waste.
Once the sample is heat fixed, stain it with safranin. This is a pinkish-red colored stain, and all cells (both bacterial and your mouth cells) will take up the stain and increase the contrast in the image.
Observe your mouth smear with the microscope. When you get to the oil immersion objective, locate and focus on a single cheek cell. As you did with the prepared slide, sketch the larger cheek cell in the circle provided and label the membrane and nucleus . Add the bacterial cells to your sketch, and try to keep the size scale accurate.
- Below, describe the shape and arrangement of the different types of bacterial cells you observe in the smear.
HomeWill Yogurt Improve your Health?
Yogurt is produced when lactic acid (homolactic) bacteria that naturally occur in milk ferment the milk sugar lactose and turn it into lactic acid. The lactic acid accumulates and causes the milk proteins to denature (“curdle”) and the liquid milk becomes viscous and semi-solid.
Within the past few years, positive health benefits have been correlated with eating fermented foods containing “live” cultures. Although several types of bacteria are known to ferment milk and produce yogurt, two genera in particular, Lactobacillus and Bifidobacterium, have been singled out as promoting good digestive health and a well-balanced immune response. Both of these are bacilli arranged in pairs or short chains. Streptococcus spp., which are cocci arranged in chains, are also usually involved in the process of making the milk into yogurt, but these are not directly associated with positive health benefits to the person who eats the yogurt.
Obtain a prepared slide labeled “yogurt smear” and view it with the microscope. The milk proteins in the yogurt will be visible as lightly stained amorphous blobs. By now you should have a pretty good idea of what bacteria look like, so locate and focus on areas where you see bacterial cells.
- HomeUsing microbiology terms, describe the shape and arrangement of the different types of bacterial cells you see in the smear
Once you’ve made the observations using the prepared slide, obtain a glass slide and a sterile swab. Collect a sample from the container of commercially prepared yogurt by swirling the swab in the yogurt, then scraping of the excess on the edge of the container. Smear this over the surface of the slide, making sure that you leave only a thin film of yogurt on the surface. Make a second smear from the container of freshly prepared homemade yogurt, if available. Allow both smears to air dry, and then heat fix them.
Once the sample(s) are heat fixed, stain them with crystal violet. This is a purple colored stain, and although both the milk proteins and cells will stain this color, the milk stains faintly and the bacteria will appear dark purple. Keep in mind that the probiotic bacteria are bacilli. Below, sketch a representative field as seen with the oil immersion objective for each of the yogurt samples.
Move your stage so you observe 10 different microscope fields. Keep track of the number of different types of bacterial cells you encounter during your survey, and record that information below:
- Cell Count of Bacteria in Commercial Yogurt:
- Number of cocci in 10 microscope fields: ____________
- Number of bacilli in 10 microscope fields: ___________
- Cell Count of Bacteria in Homemade Yogurt:
- Number of cocci in 10 microscope fields: ____________
- Number of bacilli in 10 microscope fields: ___________
- Does the container of commercially prepared yogurt state that there are “live, active cultures” in the yogurt? If the container lists the name(s) of the bacteria, write them below, followed by whether they should be bacilli or cocci.
- Considering the relative number and type(s) of bacteria you saw in the stained smear of the commercially prepared yogurt, what do you conclude about the health benefits of eating products such as this as a probiotic?
- Considering the relative number and type(s) of bacteria you saw in the homemade yogurt, what do you conclude about the health benefits of eating a fresh, homemade type of yogurt as a probiotic?
Molecular make up of cells
The practical Life Science techniques are an important part of the Life Science assessment program. This section addresses some of these important skills. Learners are introduced to the various microscopes and this enables the skills of drawing, labelling and annotating diagrams and micrographs, using microscopes as well as calculating magnification of cells. (This will be covered later in this Chapter).
Cells are the basic structural and functional units of all living organisms. Cells are made up of the compounds you learnt about in the previous chapter: carbohydrates, fats, proteins, nucleic acids and water. The word 'cell' was first used by the 17th century scientist Robert Hooke to describe the small pores in a cork that he observed under a microscope. Cells are very small structures. The human body is made up of ( ext<10>^< ext<13>>) cells. Each of these is too small to see with the human eye and it is through the development of microscopic techniques that we have been better able to visualise and understand them.
Early attempts to magnify images of objects through grinding of glass lenses eventually gave rise to the earliest microscope. In 1600, Anton van Leeuwenhoek, a Dutch microbiologist used a simple microscope with only one lens to observe blood cells. He was the first scientist to describe cells and bacteria through observation under microscope. By combining two or more lenses, the magnification of the microscopes was improved, thus allowing scientists to view smaller structures.
The dissecting microscope is an optical microscope used to view images in three dimensions at low resolution. It is useful for low-level magnification of live tissue. The development of the light microscope, (Figure 2.5) which uses visible light to magnify images allowed for up to 1000X magnification of objects through which scientists were able to view individual cells and internal cell structures such as the cell wall, membrane, mitochondria and chloroplasts. However, although the light microscope allowed for 1000X magnification, in order to see even smaller structures such as the internal structure of organelles, microscopes of greater resolving power (with up to 10 000X magnification) were required.
With the development of electron microscopes the microscopic detail of organelles such as mitochondria and chloroplasts became easier to observe. The Transmission Electron Microscope (TEM) was developed first, followed by the Scanning Electron Microscope (SEM). TEM is used to view extremely thin sections of material. Beams of electrons pass through the material and are focused by electromagnetic lenses. In SEM the electrons are bounced off the surface of the material and thus produce a detailed image of the external surface of the material. They produce a 3D image by picking up secondary electrons knocked off the surface with an electron collector. The image is then amplified and viewed on a screen. Examples of each of the image types produced by these microscopes are given in Figures Figure 2.1 to Figure 2.3.
Sections for TEM have to be so thin that they have to be prepared using a special piece of equipment called an ultramicrotome.
SEM: A natural community of bacteria growing on a single grain of sand.
SEM: These pollen grains show the characteristic depth of field of SEM micrographs.
TEM: Image of chloroplast, showing thylakoid discs within a eukaryotic cell.
Transmission electron microscopes can magnify an image 50 million times.
Figure 2.4: Transmission electron microscope in use.
The apparatus most commonly used in lab microscopy exercises is a simple light microscope. Table 2.1 shows an annotated diagram of a light microscope with a description of the function of each part. The main parts are described in the table that follows and the function of each part is explained.
Figure 2.5: Light microscope
- A cylinder containing two or more lenses.
- These lenses are held at the correct working distance.
- The ocular lens/eyepiece helps to bring the object into focus.
The revolving nose piece holds the objectives in place so that they can rotate and can be changed easily.
The objective magnifies the objects.
There are normally three objectives present:
The coarse adjustment screw is used for the initial focus of the object. By moving the stage up and down, bringing the object closer to or further away from the objective lens.
The fine adjustment screw is used for the final and clear focus of the object.
- A rigid structure for stability.
- The frame is supported by a U-shaped foot leading to the base of the microscope.
The diaphragm and condenser control the amount of light which passes through the slide.
- The microscope slide is placed here.
- The stage contains a clip or clips to prevent the slide from moving around.
- There is a hole in the stage which allows light through.
The ocular, rotating nosepiece and objectives are held above the stage by the arm.
How to use a microscope correctly
- When handling or carrying the microscope, always do so with both hands. Grasp the arm with one hand and place the other hand under the base for support.
- Turn the revolving nosepiece so that the lowest power objective is in position.
- Place the microscope slide on the stage and and fasten it with the stage clip(s).
- Look through the eyepiece and adjust the diaphragm for the greatest amount of light.
- While looking at the slide on the stage from the side, turn the coarse adjustment screw so that the stage is as close to the objective lens as possible. WARNING: Make sure you do not touch or damage the slide.
- Slowly turn the coarse adjustment screw until the image comes into focus.
- Now use the fine adjustment screw to move the stage downwards until the image is clearly visible. Never move the lens towards the slide.
- You can readjust the light source and diaphragm for the clearest image.
- When changing to the next objective lens use the fine adjustment screw to focus the image. WARNING: Never use the coarse adjustment screw for the strongest objective lens.
- Do not touch the glass part of the lenses with your fingers.
- When finished, move the stage and objective as far away from each other as possible and remove the slide.
- Disconnect the power source and cover the microscope.
- Carry the microscope by holding it firmly by the "arm" and "base" and when walking it should be near your chest.
Remember that microscopes are expensive scientific equipment and need to be handled with care to prevent damaging them. Proper lens paper should be used when cleaning dust or dirt off any lenses. Avoid getting moisture on the objective lenses. Dust and moisture are the biggest enemies of microscopes.
If using a mirror for illumination instead of a light bulb, never reflect direct sunlight as you could damage your eyes.
Differences between the light microscope and transmission electron microscope
Bacterial spores as seen under light microscope.
Chlamydomonas reinhardtii, a single celled green algae, as seen under the transmission electron microscope.
Calculating magnification (ESG4R)
Microscopes magnify an image using a lens found in the eye-piece, which is also known as the ocular lens. The image is further magnified by the objective lens. Thus the magnification of a microscope is: magnification power of the eye-piece x the power of the objective lens
Example: if the eyepiece magnification is 5X and the objective lens' magnification is 10X, the image of the object viewed under the microscope is 50X bigger than the object:
Calculating the field of view
When viewing an object through a microscope, the diameter of the circle through which you view the object is known as the field of view.
As the magnification increases, the field of view decreases.
To measure the field of view, use a microscope slide with a tiny ruler printed on it. For example, the size of the field of view shown below under low power magnification is approximately 1 mm.
Figure 2.6: Field of view is approximately 1 mm.
Once the size of the field of view is known, we can estimate the size of the objects being viewed under the microscope. At 10 X magnification, the field of view is ( ext<1,0>) mm. If the magnification is increased to 100 X, what will the new field of view be?
( ext<1,0>) mm at 10 X magnification
(x) mm at 100 X magnification
If magnification is increased 10-fold, the field of view will decrease 10-fold. Thus it will become 0.1 mm. What this means is that at higher magnification, we are able to see objects of smaller and smaller size within our field of view. This is why at higher magnification, the field of view becomes smaller.
At 500 X magnification, the field of view of a microscope is ( ext<0,05>) mm. What will the field of view be at 100X magnification?
Calculating magnification and using scale bars
When drawing cells or cellular structures, your diagrams will usually be much larger than the actual size of the structures you will be drawing. The magnification is given by:
When a scale bar is provided with the diagram, the magnification is given by:
Antireflective surface inspired from biology: A review
Optical anti-reflection means the decrease of reflection as much as possible, which has been used in many fields such as solar cells, diodes, optical and optoelectronic devices, screens, sensors, anti-glare glasses and so on. Over millions of years, natural creatures have been uninterruptedly combating with extreme environmental conditions. In particular, some biology has evolved a diversity of antireflective functional surfaces gradually. More importantly, as a result of the same order of magnitude in the ingenious structures and the wavelength of visible light, these structures can interact strongly and present excellent antireflective performance. It is worth to be mentioned that these wonderful architectures lead to a perfect performance on antireflection. This review mainly covers recent progress on the bionic antireflective structures. Then, the mechanism of the structure-based antireflective properties of some biology is analyzed. Besides, some typical models and the basic theory of these bionic structures for antireflection have been reported to facilitate mechanism analysis. At last, the prospects and the challenge researchers may faced with are also addressed. It is hoped that this review could be beneficial to provide some innovative inspirations and new ideas to the researchers in the fields of engineering, and materials science.
A microscope is an instrument that can be used to observe small objects, even cells. The image of an object is magnified through at least one lens in the microscope. This lens bends light toward the eye and makes an object appear larger than it actually is.
Though modern microscopes can be high-tech, microscopes have existed for centuries – this brass optical microscope dates to 1870, and was made in Munich, Germany.
Photograph by Martin Shields / Alamy Stock Photo
A microscope is an instrument that is used to magnify small objects. Some microscopes can even be used to observe an object at the cellular level, allowing scientists to see the shape of a cell, its nucleus, mitochondria, and other organelles. While the modern microscope has many parts, the most important pieces are its lenses. It is through the microscope&rsquos lenses that the image of an object can be magnified and observed in detail. A simple light microscope manipulates how light enters the eye using a convex lens, where both sides of the lens are curved outwards. When light reflects off of an object being viewed under the microscope and passes through the lens, it bends towards the eye. This makes the object look bigger than it actually is.
Over the course of the microscope&rsquos history, technological innovations have made the microscope easier to use and have improved the quality of the images produced. The compound microscope, which consists of at least two lenses, was invented in 1590 by Dutch spectacle-makers Zacharias and Hans Jansen. Some of the earliest microscopes were also made by a Dutchman named Antoine Van Leeuwenhoek. Leeuwenhoek&rsquos microscopes consisted of a small glass ball set inside a metal frame. He became known for using his microscopes to observe freshwater, single-celled microorganisms that he called &ldquoanimalcules.&rdquo
While some older microscopes had only one lens, modern microscopes make use of multiple lenses to enlarge an image. There are two sets of lenses in both the compound microscope and the dissecting microscope (also called the stereo microscope). Both of these microscopes have an objective lens, which is closer to the object, and an eyepiece, which is the lens you look through. The eyepiece lens typically magnifies an object to appear ten times its actual size, while the magnification of the objective lens can vary. Compound microscopes can have up to four objective lenses of different magnifications, and the microscope can be adjusted to choose the magnification that best suits the viewer&rsquos needs. The total magnification that a certain combination of lenses provides is determined by multiplying the magnifications of the eyepiece and the objective lens being used. For example, if both the eyepiece and the objective lens magnify an object ten times, the object would appear one hundred times larger.
The dissecting microscope provides a lower magnification than the compound microscope, but produces a three-dimensional image. This makes the dissecting microscope good for viewing objects that are larger than a few cells but too small to see in detail with the human eye. The compound microscope is typically used for observing objects at the cellular level.
Though modern microscopes can be high-tech, microscopes have existed for centuries &ndash this brass optical microscope dates to 1870, and was made in Munich, Germany.
Caenorhabditis elegans: Cell Biology and Physiology
Amy S. Maddox , Paul S. Maddox , in Methods in Cell Biology , 2012
Inverted microscopes are so called because the objective lens points upward at the sample and focusing is accomplished by moving the objective lens and not the stage. Because the stage is generally heavier than the objective turret, inverted microscopes are less prone to focal drift due to gravity. Fluorescence studies are well suited to inverted microscopes because the imaging light path is below the specimen, within the microscope body where it has a low center of gravity and is less susceptible to vibration-induced aberrations. For this reason, biophysics labs generally prefer inverted microscopes. Correspondingly, transmitted light imaging modes (e.g., DIC) are normally less stable on inverted microscopes as the illuminating light path is elevated, poorly supported, and prone to vibration. One benefit of inverted microscopes is the ease of access to the specimen such as for microinjection at a step angle (as preferred for injecting C. elegans this is not easily accomplished on an upright microscope).
This project is part of the research work undertaken by the Greek group for the ‘History and Philosophy of Science in Science Teaching’ (HIPST) project w8 , funded under the 7 th Framework Program, Science in Society-2007-126.96.36.199 – teaching methods.
The author would like to thank the coordinator of the Greek research group of the HIPST project, Fanny Seroglou (associate professor at the Aristotle University of Thessaloniki) for her support on the project.
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Cytology, the study of cells as fundamental units of living things. The earliest phase of cytology began with the English scientist Robert Hooke’s microscopic investigations of cork in 1665. He observed dead cork cells and introduced the term “cell” to describe them. In the 19th century two Germans, the botanist Matthias Schleiden (in 1838) and the biologist Theodor Schwann (in 1839), were among the first to clearly state that cells are the fundamental particles of both plants and animals. This pronouncement—the cell theory—was amply confirmed and elaborated by a series of discoveries and interpretations. In 1892 the German embryologist and anatomist Oscar Hertwig suggested that organismic processes are reflections of cellular processes he thus established cytology as a separate branch of biology. Research into the activities of chromosomes led to the founding of cytogenetics, in 1902–04, when the American geneticist Walter Sutton and the German zoologist Theodor Boveri demonstrated the connection between cell division and heredity. Modern cytologists have adapted many methods of physics and chemistry to investigate cellular events. See also cell.
Under the Microscope: Get Forensic with Hair Analysis
Life was long gone from the cold, bloody corpse when the crime scene investigators arrived. The seasoned team soon confirmed the death was a murder, but no footprints, no fingerprints, no weapons were found𠅊 few strands of hair caught in the dead woman’s broken fingernails were the only evidence the killer left behind.
Who’s the culprit? Ask your students. Teaching basic physiological science concepts is interesting from a forensics point of view. By incorporating a problem-solving approach to science education, teachers engage their students in exciting and innovative ways. Forensic labs also provide “real-world” applications of science and math.
Trichology is what?
Trichology is the scientific study of the structure, function, and diseases of human hair. Medical professionals, beauticians, and forensic scientists, among others, practice occupations within trichology. Hair is a valuable tool for forensic scientists. It is more resistant to decay than most other body tissues and fluids, thus remaining intact far longer than other evidence. This durability makes hair one of the most frequently found pieces of evidence at crime scenes.
A hair shaft is composed of 3 layers. The outer layer, or cuticle, consists of overlapping scales, with the free ends of the scales directed toward the tip of the shaft. Just beneath the cuticle is the cortex, made up of compact, elongated cells and often containing pigment granules. The central core of a hair shaft is the medulla, composed largely of air spaces.
Forensic scientists perform 3 major types of hair analysis: (1) testing the hair shaft for drugs or nutritional deficiencies in a person’s system, (2) analyzing DNA collected from the root of the hair, and (3) viewing hair under a microscope to determine if it’s from a particular person or animal. They usually study the hair’s scale pattern and appearance of the medulla to identify a hair of unknown origin.
Studying scale patterns
scientists study a cast of the hair shaft for determining scale pattern. The arrangement and shape of hair scales can vary greatly from species to species and are often very distinctive. Scientists usually classify scales into 1 of 3 categories:
- Coronal𠅌ompletely encircling the hair shaft
- Spinous—Long, narrow, and not encircling the hair shaft
- Imbricate—Short, wide, and not encircling the hair shaft
Turn your students into a CSI team and let them solve a 𠇌rime” using hair analysis. The forensic mystery will both engage and intrigue them while they learn science concepts.
Making a cast mount of a hair shaft
- Place a drop of latex near one end of a clean slide. Note:If you do not have latex, an alternate casting medium is nail polish. Brush a thin layer of nail polish in the middle of a clean slide.
- Tilt a second slide over the first (at approximately a 30° angle), ensuring that the slide ends farthest from the drop of latex are touching.
- Slowly pull the tilted slide over the first until it touches the drop of latex.
- Allow the latex to run along the edge of the tilted slide.
- With a smooth motion, push the tilted slide back along the first slide to spread the latex into a thin film.
- Immediately place several strands of hair on the film of latex or nail polish.
- Let the slide sit undisturbed for 10 to 15 minutes allowing the latex to harden. If using nail polish, wait until the polish is tacky-dry.
- Once the latex is hard or polish is nearly dry, use forceps to pull as much hair as possible off the slide (it is not necessary to remove every strand of hair).
- Examine the slide using only the low-power objective of a microscope. Note:Do not attempt to examine scale casts under the high-power objective. Look for impressions of individual scales and note the following features:
- Whether or not individual scales completely surround the hair shaft
- The general shape of an individual scale
- Whether the exposed edge of a scale is smooth or jagged
Studying the medulla
A whole mount allows study of the appearance of the medulla however, a medulla is not always present in a hair. When medullae are present, they often show distinctive variations between species. The appearance of a medulla is classified as continuous (unbroken), intermittent (regular intervals), or fragmented (irregular intervals).
Making a whole mount of hair
- Microscope Slide
- Mountant or Water
- Strands of Hair
- Paper (for drawing)
- Pencil (for drawing)
- Obtain a clean microscope slide and place a drop of mountant or water on it.
- Place several strands of hair on the drop of mountant or water.
- Use forceps and slowly lower a coverslip onto the drop of mountant or water.
- Examine the slide under the low- and high-power objectives of a microscope. Examine several different sized hairs while noting any internal features such as granules or air spaces. Draw the hair showing the observed features.
Note: If you are using a mountant, keep the finished slide flat until the mountant hardens. To harden the mountant, heat the slide in an oven at 60° C for 1 day or leave at room temperature for several days.
Identifying whether the hair is human or animal is the first step in forensic hair analysis. Human hairs usually have a thin (less than ⅓ of the hair’s diameter) or an absent medulla region. Animal hairs usually have thick medullae (more than ½ of the hair’s diameter). Compare the photos below to what you see under the microscope.
The next step is comparing the found hair to known suspects or animals present at the crime scene. This analysis can be subjective based on the person’s experience at identifying the shape of the medulla. For beginning forensic students, simple identification of animal or human is sufficient. Using characteristics such as color, curliness, and thickness may help identify different human hairs.
All supplies needed to make cast mounts and whole mounts of hair, including hair from animals and humans, are in the Hair Analysis Kit (item 699870) .
Extend your inquiry
Carolina offers many forensic kits that teach biology concepts in a fun, hands-on “ real-world” scenario. Following are some kits you may want to consider.
Microscopic Forensics Kit, item 699880. Students undergo forensic “training,” observing labeled slides of hair, blood, and textiles—materials forensic investigators typically find at a crime scene. After studying the known slides, students receive a mock murder mystery and crime scene 𠇎vidence.” They then examine the evidence, compare it to the suspects’ testimonies, and pinpoint the culprit.
The Case of the Murdered Mayor Kit, item 699830. Students become crime scene investigators using their observational skills and deductive reasoning to solve this realistic crime scenario. Kit includes the following activities: dusting and lifting fingerprints, examining hair samples, analyzing impressions of tire tracks, blood typing with synthetic blood, examining entomological evidence and reviewing a police interview log.
Carolina™ Forensic Dissection Kit, item 221489. Students conduct a pig dissection, modeling the protocols a pathologist uses for a human autopsy. Upon completing the forensic dissection, students return the organs to the body cavity and suture the incisions. A Carolina’s Perfect Solution ® adult pig heart and pig kidney are also included for comparison with the dissected specimen. Students use a set of 7 prepared slides, extending and enhancing the dissection by examining tissue types found in each system.
Carolina ’s extensive line of forensic science materials is ideal for teaching various concepts in science and math. These activities are also an engaging and innovative way to enhance your science curriculum. For more information about our vast array of kits and supplies, call 800.227.1150 or visit www.carolina.com.
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Objectives: View objects at 40x, 100x, and 400x magnification!
2 LED Lights: Two light sources allow this microscope to magnify both slides and solid objects!
Portability: Light-weight construction and battery operation allow kids to take this scope and explore their surroundings, rather than being tied to a desk!
DUO SCOPE MICROSCOPE
The Duo Scope is our most popular microscope in the My First Lab series! Designed with two light sources, this microscope can magnify both slides and solid objects. View slides, coins, plants, stamps, insects, jewelry, and more!
No longer do you need one basic biological microscope to look at specimens on a slide (light shines up from under the slide and through the tissue) and a traditional stereo or dissecting microscope to view solid objects (light shines down onto the specimen to be observed). Instead, equipped with dual lighting, the Duo Scope allows for both applications!
Cordless battery power makes it possible for kids to take this microscope into the field. Portability, combined with durability, makes for a perfect option for explorers! This microscope is an affordable, high-quality option for your STEM-focused child!
Light Microscope Vs Electron Microscope
Using an instrument the size of his palm, Anton van Leeuwenhoek was able to study the movements of one-celled organisms. Modern descendants of van Leeuwenhoek's light microscope can be over 6 feet tall, but they continue to be indispensable to cell biologists because, unlike electron microscopes, light microscopes enable the user to see living cells in action. The primary challenge for light microscopists since van Leeuwenhoek's time has been to enhance the contrast between pale cells and their paler surroundings so that cell structures and movement can be seen more easily. To do this they have devised ingenious strategies involving video cameras, polarized light, digitizing computers, and other techniques that are yielding vast improvements, in contrast, fueling a renaissance in light microscopy.