Investigating specimens using microscopes

In a nutshell

Cells and other microscopic specimens can be viewed using a light microscope. It is also important to be able to draw good scientific drawings and identify the structures that you can see under the microscope. Using the power of the microscope lenses, the total magnification of the image can be calculated.

Equations

These are the equations that you will need to know for these microscopy practicals.

$total\space magnification = eyepiece \space lens \space magnification\times objective \space lens \space magnification$

$magnification = \frac{image\space size}{real\space size}$

Quantitative units

The specimens you will be examining are very small so you will need to know how to express their size using the following units.

Unit name	Unit	Standard form
$millimetre$	$mm$	$\times10{^-}{^3}\space m$
$micrometre$	$\mu m$	$\times10{^-}{^6}\space m$
$nanometre$	$nm$	$\times10{^-}{^9}\space m$
$picometre$	$pm$	$\times10{^-}{^1}{^2}\space m$

Equipment

The following equipment can be used to view microscopic specimen.

Equipment	Use
Microscope	To view the specimens.
Microscope slide	To hold the specimen so it can be viewed under the microscope.
Water	To place a drop on the slide and prevent the specimen drying out.
Cotton swab	To collect cheek cells.
Pipette	Used to put a drop of water on the slide.
Tweezers	To transfer the specimen onto the slide.
Stain	A stain is used if the specimen is colourless. Staining the specimen will make it easier to see.
Coverslip	A small piece of glass that is put on top of the specimen on the slide to flatten the specimen and stop the slide from drying out.
Filter paper	To absorb excess stain.

Experiment 1: Preparing a specimen on a microscope slide

Specimens must be prepared before they are viewed under a microscope. You must be careful when you do this as you don't want to damage the specimen. Onion cells and cheek cells are most commonly used for plant and animal cells respectively.

Experimental variables

Every experiment will have an independent, dependent and constant variable. The independent variable is the variable that you change. The dependent variable is the variable which depends on the independent variable, therefore it is the one you measure. The control variable is the variable that is kept the same during the experiments.

Independent variable	$The\space specimen\space you\space are\space examining.$
Dependent variable	$Magnification.$
Control variable	$The\space type\space of\space dye\space used\space and\space the\space stage\space micrometer.$

Safety precautions

When carrying out experiments it is very important to consider the safety precautions so nobody gets hurt.

Hazard	Risk	Safety measure
Stains	The stains may stain your skin.	Wear gloves when using the stains or just be careful not to get the stain on your skin or clothes.
Coverslips and microscope slides	These are made of glass and may cut you.	Be careful when handling the coverslips and microscope slides.

Method for preparing an animal cell specimen

Biology; Biology practicals; KS4 Year 10; Investigating specimens using microscopes

1.	Collect the specimen. Use a cotton swab to swab the inside of your cheek.
2.	Using a pipette, add a drop of water to the slide.
3.	Gently rub the cotton swab on the slide.
4.	Add a drop of methylene blue stain to the slide. This stain turns the nucleus of the cell blue.
5.	Using tweezers, place a coverslip over the specimen.
6.	Use filter paper to absorb any excess stain.

Method for preparing a plant cell specimen

Biology; Biology practicals; KS4 Year 10; Investigating specimens using microscopes

1.	Collect the specimen. Using the tweezers, peel away a layer of skin from an onion.
2.	Using a pipette, add a drop of water to the slide.
3.	Using the tweezers, place the onion skin layer on the slide. *Note:* Make sure it is laying flat on the slide and there are no air bubbles.
4.	Add a drop of iodine stain to the slide.
5.	Using tweezers, place a coverslip over the specimen.
6.	Use filter paper to absorb any excess stain.

Experiment 2: Viewing the specimen using a microscope

Parts of a microscope

Biology; Biology practicals; KS4 Year 10; Investigating specimens using microscopes

	Part of the microscope	Explanation
1.	Eyepiece	The eyepiece is what you look through to see the magnified image. The eyepiece contains the eyepiece lens which magnifies the specimen to produce the image you can see.
2.	Body tube	The body tube separates the eyepiece and objective lenses.
3.	Rough focusing knob	The rough focusing knob is used to move the stage to bring the image into focus.
4.	Fine focusing knob	The fine focusing knob is used to bring the image into focus after the rough focusing knob is used.
5.	Objective lenses	The objective lens magnifies the specimen. There are usually three lenses with increasing power. The highest-powered lens will create a greater magnification.
6.	Handle	The microscope should be held by the handle.
7.	Stage	The stage is where the slide is placed.
8.	Mirror	The mirror should be angled towards a light source and up through the hole in the stage. This provides the light that enables you to see the image. *Note:* In some microscopes there will be a lightbulb instead of a mirror.

Method

1.	Place the prepared slide onto the stage and clip it in place so the specimen is positioned where the light will come through.
2.	Select the lowest powered objective lens. Rotate the lens so it is over the slide.
3.	Use the coarse adjustment knob to move the stage. Move the stage up so the slide is just underneath the objective lens.
4.	Look down the eyepiece lens and use the coarse adjustment knob to move the stage downwards until the specimen is almost in focus.
5.	Use the fine adjustment knob to bring the specimen into focus. The image should now be clear.
6.	If you need a greater magnification, switch to a higher powered lens and refocus the image.

Analysis

To report on what you have observed through the microscope, you need to calculate the size of the specimen, report on the magnification and be able to draw biological drawings.

Here are two methods for calculating the size of the specimen.

Method one

1.	Place a clear ruler on the stage.
2.	Measure the diameter of the circle that you can see when looking down the eyepiece. This is your field of view (FOV).
3.	If you increase the magnification, recalculate your FOV. Note: The FOV allows you to estimate the size of the specimen.

Method two

Biology; Biology practicals; KS4 Year 10; Investigating specimens using microscopes

1.	Place an eyepiece graticule (A) on the eyepiece lens. If you look down the lens you should now see a scale. Before being able to measure the cells, you must calibrate the eyepiece graticule. This means you need to find out the distance that each division on the eyepiece graticule represents. *Note:* Each objective lens will give a different value so you must calibrate the eyepiece graticule each time you use a new lens.
2.	Remove the slide with your specimen and place a stage micrometer (B) onto the stage. Note: A stage micrometer is a slide with an accurate scale on it.
3.	Line up the stage micrometer and the eyepiece graticule. Select a set measurement on the stage micrometer and count the number of divisions on the eyepiece graticule that fit in this set measurement. Using the stage micrometer, calculate the distance (in micrometers) of one division on the eyepiece graticule.
4.	Remove the stage micrometer and use the calibrated eyepiece graticule to measure the cell on your original specimen slide.

Calculating the magnification

You can use the equation below to work out the total magnification if you know the magnification of both the eyepiece lens and the objective lens.

total\space magnification = eyepiece \space lens \space magnification\times objective \space lens \space magnification

If you do not know the magnification of the lenses, you can still work out the magnification. To do this, you will need to measure the image and know the real size of the specimen.

magnification = \frac{image\space size}{real\space size}

Note: The units for the image size and the real size should be the same. The units shown in the table at the top of this summary can be used.

Creating biological drawings

Biological drawings are needed to show what you have seen down the microscope. These are different to normal drawings and it is important that you follow the steps below.

1.	Use a sharp pencil to draw the outline of all the main features you can see.
2.	Make sure your drawing takes up at least half of the space you are given and that all of the parts are proportionally the correct size.
3.	Draw straight lines from each structures and write the name of the structure.
4.	Include the magnification you have used and a scale bar.

Example

Biology; Biology practicals; KS4 Year 10; Investigating specimens using microscopes

Evaluation

After completing the experiment, you must comment on the quality of your data. You should also think about how your method could be improved. If you have calculated the magnification from the actual size of the specimen and the size of the image created, you can compare this to the actual magnification you used. The two values for magnification should be the same.

The most common mistake with calculating the magnification is to reverse the formula. In order to calculate the correct magnification, it is also crucial to correctly calibrate the eyepiece graticule (or calculate the FOV). If you do not do this correctly, the sizes will be incorrect. Therefore, your result for the magnification will also be incorrect.