Use a light microscope to: make observations of
(i) transverse sections of roots, stems, leaves;
(ii) plant tissues;
(iii) identify sclerenchyma fibres, phloem, sieve tubes, xylem vessels and their location
● To be able to use a microscope competently to observe biological specimens
● To learn how to draw and label plan diagrams accurately
● To be able to identify sclerenchyma fibre, phloem, sieve tubes and xylem vessels
● Wear eye protection.
● Avoid all skin contact with the toluidine blue stain; disposable gloves may be worn.
● Tell your teacher if you spill the stain on the bench or on your skin.
● Take care when using razor blades and mounted needles and ensure you hand them back to your teacher when you have finished with them.
● If you are using a microscope with daylight illumination – a mirror – you must not put the microscope in a position where direct sunlight might strike the mirror and be reflected into the eyes through the microscope. This could cause permanent retinal damage or blindness.
● eye protection
● protective gloves
● plant stem, at least 5 cm long
● root section, at least 3 cm long
● large plant leaf
● toluidine blue O stain
● two glass microscope slides
● two coverslips
● small paintbrush or tweezers
● single-sided razor blade
● white tile
● watch glass
● dropping pipette
● mounted needle
● lens tissue
● polystyrene strips
● absorbent paper (e.g. paper towel)
● stop clock
figure A Transverse section of a stem as seen through a microscope at low power
1. Collect a piece of plant stem. Add a few drops of water to the centre of the white tile and wet the razor to reduce friction. Hold the plant stem firmly, keeping your fingers away from the edge of the razor. Cut several transverse sections (across the stem), keeping them as thin as possible. Incomplete thin sections may sometimes be better than thicker complete ones. Use a brush to transfer the sections to water in a watch glass.
2. Select the thinnest section and place it on a slide. Add a drop of water. Remove excess water by carefully touching the edge with absorbent paper.
3. Wearing gloves and eye protection, add two drops of toluidine blue O stain and leave for 2–
4 minutes. Then add a coverslip and gently remove excess stain with a paper towel.
4. Turn the objective lens to low power. Examine the slide under the microscope. To do this, bring the lens as close to the slide as possible while watching it from the side. Then, looking through the eyepiece, focus using the coarse focusing knob, moving the lens away from the stage. This avoids damage to the slide and lens. Use the fine focus until a clear view of the section is established.
5. Still on low or medium power, draw and annotate a simple outline plan of the section. Show the arrangement of tissues within the stem but do not include any cell details. Use figure A to help you identify the different tissues and label them.
6. Repeat steps 1–5 using a piece of root.
7. To make a leaf cross section, place the leaf between two thin pieces of polystyrene. Gently stroke the razor blade across the top of the leaf and polystyrene to shave off four or five thin sections.
8. Select a thin section and place it on a glass slide with one or two drops of water. Cover with a coverslip.
9. Repeat steps 4 and 5 with the leaf section.
Note: You may not need to prepare your own specimens. You may instead be given preprepared slides to use. In this case, you will only need to follow steps 4 and 5 for the specimens provided.
● Before starting this practical, you may need to remind yourself about parts of the microscope and how to use the instrument as detailed in the learning tips on page 27.
● Whole sections are not needed to make the plan drawings. A thin partial section is better than a thick whole section.
● Toluidine blue O is a metachromatic stain, which means it reacts with different chemical components of cells to produce a variety of colours. This can provide information about the nature of the cell. Toluidine blue O stains lignin and tannins green to blue, pectins pinkish purple and nucleic acids purplish or greenish blue.
● Use a sharp HB pencil. Keep lines clear and continuous, not feathery or sketched.
● Draw only what you see. Do not draw stylised patterns and do not make it up.
● Start with an outline. Keep it large and think about proportions.
● Do not use shading or colour.
● Draw label lines in pencil with a ruler. Lines should not have arrowheads and should just touch the item to be labelled.