Onion epidermal cells are commonly used for studying the basic structure of a plant cell. These cells are obtained from the thin inner epidermis of onion bulb. The accepted scientific name of onion is Allium cepa L. and it is listed as an accepted species.
The inner epidermis is a very thin and almost transparent layer. It can be peeled from the inner surface, placed in water or stain and mounted directly below a coverslip. Sectioning of the tissue is not required for this simple preparation.
Under a compound light microscope, onion peel shows closely arranged elongated and rectangular cells. A clear cell wall surrounds every cell. The cytoplasm is mainly present as a thin peripheral layer and a large central vacuolar region occupies most of the cell interior. After proper staining, the nucleus is generally seen near the cell side, between the vacuole and the cell wall.
All internal details are not equally clear in every slide. It depends on the stain, magnification, correct focusing and the quality of prepared slide. A folded peel, excess stain and trapped air bubbles can make the observation less clear.
Quick Answer
Appearance: Elongated rectangular cells arranged side by side, with clear walls, peripheral cytoplasm and a large clear central region.
Stain: Dilute iodine in potassium iodide solution is commonly used for increasing the contrast of onion epidermal cells.
Recommended magnification: Start at about 100× to locate the cells. Then use around 400× for observing the nucleus, cytoplasm and other cell details.
What Are Onion Epidermal Cells?
Onion epidermis is the thin surface tissue present over an onion bulb scale. The peel used in laboratory work is generally removed from the inner concave surface of a fresh fleshy scale. This surface is also called the adaxial epidermis. Onion scale epidermis is usually only one cell thick.
The thin peel is commonly called the epidermal membrane or onion peel. Here, membrane does not refer to the plasma membrane of one cell. It is a complete sheet of epidermal tissue, formed of many cells remaining joined with each other.
This epidermal membrane is different from the brown and papery outer skin of onion. The dry outer skin develops from the older outer bulb scales. These tissues undergo drying, browning and cell death, and later form a protective covering around the bulb. The epidermal peel for microscopy is taken from a moist fleshy scale, not from this dry skin.
Onion epidermal tissue contains many closely arranged eukaryotic plant cells. The cells form a continuous layer and have a nucleus, cytoplasm, vacuole, plasma membrane and a surrounding cell wall. Some of these internal parts are not clearly seen without proper staining. Plants are multicellular eukaryotes and the epidermis represents one of their main tissue systems.
The peel is thin and nearly transparent. It can be spread directly in a drop of water on a glass slide. Transmitted light can pass through this one-cell-thick preparation, therefore the cells can be viewed by a compound light microscope without cutting a tissue section.
Why Is Onion Peel Used for Microscopy?
Onion peel is thin and can be removed easily from the fleshy bulb scale with forceps. It generally comes out as a continuous epidermal layer. The peel can be placed directly on a glass slide and spread in a single flat layer.
The epidermis consists of closely arranged cells. These cells are comparatively large and show a regular arrangement. Onion epidermis is therefore used as a monolayered and homogeneous plant tissue for different microscopic studies.
No complicated tissue sectioning is required. A small peel is removed, mounted in water or stain and covered with a coverslip. Since the specimen is only about one cell thick, transmitted light can pass through it. The cell layer is then observed by a compound light microscope.
The cell walls are clear and easy to recognise. They form visible boundaries between the neighbouring cells. Onion epidermis has also been widely used for studying the structure and arrangement of primary plant cell walls.
It is suitable for beginner microscope training. Students can practise temporary slide preparation, focusing, changing objective lenses and making a biological drawing. The specimen usually gives many cells in one field, so the basic plant cell arrangement can be observed without a difficult preparation.
Onion bulbs are readily available and only a very small amount of peel is required. One onion can supply enough epidermal tissue for several preparations or even a class practical. This makes it an inexpensive biological specimen for laboratory work.
Onion peel can be used for more than the simple observation of cells. Different stains are used to increase the contrast of the cellular structures. It is also used in osmosis and plasmolysis experiments, because the solution surrounding the living cells can be changed easily. The large cells are suitable for cell-size measurement and microscope calibration exercises also.
What Does Onion Peel Look Like Under a Microscope?
Onion peel under a microscope shows many elongated and rectangular cells. Some cells look like bricks due to their straight sides. They remain arranged side by side and generally form regular rows. The shape may not be exactly same in every region of peel.
The cell walls form the most conspicuous boundaries. These walls separate one cell from another and produce the brick-like appearance. Inside the wall, a thin layer of cytoplasm occurs mainly toward the cell periphery.
A large central vacuole occupies most part of the mature epidermal cell. Due to this, the cytoplasm and nucleus are pushed toward the cell side. The central region may appear clear, although it is not actually an empty space. It contains cell sap and is enclosed by the tonoplast.
The nucleus appears as a small rounded or oval structure, commonly near the cell margin. It becomes easier to locate after a suitable stain is added. In an unstained preparation it can remain very faint and sometimes is not identified.
Chloroplasts are normally not visible in the inner epidermal cells of a white onion bulb. These cells are obtained from non-green fleshy scales and do not show the numerous green chloroplasts seen in a photosynthetic leaf cell. This is an important difference between onion epidermis and cells of Elodea or other green leaves.
Appearance Without Staining
The unstained onion epidermis is transparent or nearly colourless. Under correct illumination, the cell outlines can be seen because the walls produce differences in light refraction. Internal structures give much less contrast.
The vacuolar region generally looks clear and may appear empty. The cytoplasm is present as a thin peripheral layer, but it can be difficult to recognise. The nucleus also remains pale or invisible when the preparation is thick, folded or supplied with excess light.
The plasma membrane is generally not separated clearly from the cell wall in a normal water-mounted cell. Turgor pressure presses the membrane and peripheral cytoplasm against the inner side of the wall. It is more easily distinguished when the protoplast pulls away from the wall during plasmolysis.
Too much illumination can wash out the transparent specimen. In this condition only a faint arrangement of cells may be obtained. The condenser iris or diaphragm is adjusted to reduce the light and produce better contrast. Closing it too far, however, makes the field dark and reduces the useful detail.
Appearance After Staining
Staining increases the contrast between different parts of onion epidermal cell. The cell wall remains distinct, while the nucleus becomes darker. Peripheral cytoplasm may also appear more clearly, producing a visible difference between the stained cell material and the transparent vacuolar region.
Iodine solution commonly gives a yellowish-brown or brown appearance and improves the visibility of cell structures. It can darken the nucleus. When starch grains are present, iodine produces a blue-black colour, but starch grains are not always found in onion epidermal preparations.
Safranin generally produces pink to red staining of plant material. Methylene blue gives a blue colour and can increase the contrast of the nucleus and other stained cell contents. The final shade depends on stain strength, staining period and the thickness of peel.
Excess stain should be removed by filter paper or washing, according to the method. Overstaining makes a large part of the preparation very dark. The nucleus, cytoplasm and other boundaries can then merge in colour and become less clear, instead of showing more detail.
Aim of the Onion Peel Experiment
The main aim of the onion peel experiment is to prepare a stained temporary mount of onion epidermis and study its cells by using a compound light microscope.
The following are the aims-
- To prepare a thin and flat temporary slide of onion epidermal peel without folding of the tissue.
- Observation of the shape and regular arrangement of onion epidermal cells under low and higher magnification.
- To identify the visible parts of a plant cell, mainly the cell wall, cytoplasm, stained nucleus and large central vacuolar region.
- The staining is done to increase contrast and make some cellular parts more clear during observation.
- To practise the correct use of microscope, focusing of specimen and changing the objective lenses.
- It also helps in preparing a labelled biological drawing of the onion epidermal cells as actually seen through the microscope.
Materials Required for the Onion Peel Experiment
The following are the materials required for preparation and observation of onion epidermal peel-
| Material | Purpose |
|---|---|
| Fresh onion bulb | It provides a fresh inner epidermal peel from the fleshy scale of Allium cepa L. The peel should remain thin and moist. |
| Forceps | It is used to hold and gently remove the transparent epidermal layer without much damage. |
| Scalpel or blade | Used for cutting a small piece of onion scale and trimming the peel into required size. It should be handled carefully. |
| Fine brush or mounted needle | The brush helps in transferring the delicate peel. A mounted needle is used to spread it flat and lower the coverslip. |
| Watch glass | It holds the peel in water or stain. This prevents early drying and folding of the specimen. |
| Distilled water | It is used for keeping the peel hydrated, rinsing excess stain and preparing a simple water mount. |
| Iodine, safranin or another suitable stain | Staining increases contrast in the nearly transparent cells. Iodine in potassium iodide, safranin or methylene blue can be selected according to the practical method. |
| Glycerine or water | These are used as mounting medium. Water is suitable for immediate observation, whereas glycerine reduces drying of the temporary preparation. |
| Dropper | It is used to add a controlled drop of water, stain or mounting medium on the slide. |
| Glass slide | The onion peel is placed and spread over its clean central surface for microscopic observation. |
| Coverslip | It covers the peel and keeps the specimen in a thin flat layer. It is lowered slowly to avoid trapped air bubbles. |
| Blotting paper | It is used to remove excess water, stain or glycerine from the edge of coverslip. |
| Compound light microscope | It is used to observe the mounted epidermal cells, first at low power and then under higher power. |
| Gloves and eye protection | These are used where required by the stain safety sheet and local laboratory rules. They reduce skin or eye contact with chemicals. Sharp blades are used only with proper instruction. |
How to Prepare a Temporary Mount of Onion Peel
The temporary mount is prepared by removing a thin epidermal peel, staining it and placing below a coverslip. The following are the steps-
A. Obtaining the Epidermal Peel
- Separate one fresh and fleshy scale from the inner part of onion bulb.
- Cut a small piece of the scale, about 1 cm × 1 cm is sufficient. The thin membrane present on its inner concave surface is selected.
- Hold one edge of the membrane with forceps. Lift it gently from the scale. It should come out as a thin and transparent epidermal peel.
- Cut a small manageable piece of this peel. Do not include the thick fleshy tissue below it, because it produces an uneven and less transparent preparation.
- Immediately place the peel in water or keep it in a watch glass containing water. This prevents drying of the cells.
- The peel should not be allowed to curl or fold. A dried peel becomes difficult to spread and may produce damaged cells.
B. Staining and Mounting the Specimen
- Take a clean glass slide. Place one drop of water or suitable mounting medium at its centre.
- Transfer the onion epidermal peel into the drop with forceps or a fine brush. Do not leave it exposed for a long period.
- Spread the peel gently by using a mounted needle. It should form a flat single layer without folded edges.
- Add a small and controlled amount of suitable stain. Iodine solution, safranin or another selected stain may be used according to the experiment. A large amount is not required.
- When stain is added after placing the coverslip, keep one drop at one edge of coverslip. Touch filter paper to the opposite edge. The stain is drawn below the coverslip with the liquid.
- Excessive stain can be removed with blotting or filter paper. This is done carefully from the slide edge.
- Hold the coverslip with a mounted needle. Place one edge first and lower it slowly at an angle over the peel. This method reduces the trapping of air bubbles.
- Air bubbles should not remain over the specimen. They interfere with observation and can be confused with cell structures.
- Blot the excess water or stain from around the coverslip. Do not press over it, as pressing can crush or displace the epidermal cells.
Schematic diagram showing Staining and Mounting procedure of onion peel
C. Focusing the Slide Under the Microscope
- Rotate the nosepiece and select the lowest-power objective lens first.
- Place the prepared slide on the microscope stage. Secure it with the stage clips.
- Move the slide until the onion peel is centred above the stage opening and light path.
- While looking from the side, bring the low-power objective close to the slide by using the coarse adjustment. The objective must not touch the coverslip.
- Look through the eyepiece. Slowly move the objective away from the slide with the coarse focus until the cells become visible. Coarse adjustment is used only during the initial low-power focusing.
- Use the fine adjustment knob to obtain a clear image. Adjust the light source and iris diaphragm also. Transparent onion cells are difficult to see when the illumination is excessive.
- Move the slide and centre a clear group of cells in the field. A flat region without folds and air bubbles is selected.
- Rotate the nosepiece to a higher-power objective. The selected cells should remain near the centre.
- Use only fine adjustment for detailed viewing under higher power. The cell wall, cytoplasm and stained nucleus can then be examined more clearly.
- Never force the objective downward while looking through the eyepiece. Contact of the lens with coverslip can break the slide and may damage the objective.
Onion Cells at Different Magnifications
Total magnification = eyepiece magnification × objective magnification
When a 10× eyepiece is used, the 4× objective gives 40× total magnification. The 10× objective gives 100× and the 40× objective produces 400× total magnification.
As magnification increases, a smaller area of onion peel remains in the field of view. The image becomes larger, but all cell organelles do not become visible only by increasing magnification. Staining, contrast and resolving power are also required.
View at 40× Total Magnification
- At 40×, a broad sheet of onion epidermal tissue is observed.
- Many cells remain present within one field. They occur side by side and form rows.
- The general rectangular or elongated pattern can be recognised. Individual cells look small at this magnification.
- Cell-wall boundaries may be visible as broad outlines. The overall arrangement is easier to examine than the internal parts.
- The field of view is large. It is useful for locating a flat area without folds and air bubbles.
- The nucleus, peripheral cytoplasm and vacuolar region are generally indistinct. These may remain faint even in a stained slide when its contrast is poor.
View at 100× Total Magnification
- Individual onion cells become more distinct at 100×.
- The cell walls show clearer separation. Cell shape, size difference and arrangement in rows are better observed.
- Fewer cells occur in the field as compared to 40×. This is due to the smaller field of view at greater objective magnification.
- A thin cytoplasmic region may become noticeable near the cell wall. It is not equally clear in every cell.
- Properly stained nuclei may begin to appear as darker rounded or oval bodies. Their visibility depends much on stain uptake and correct focus.
- The central region usually looks clear. It represents the area occupied mainly by the large central vacuole.
View at 400× Total Magnification
- At 400×, only a small number of epidermal cells remain within the microscope field.
- A properly stained nucleus is more readily recognised. It commonly occurs toward the side of the mature cell.
- The cytoplasm is seen mainly as a thin peripheral layer. Sometimes cytoplasmic strands can also cross the vacuolar region, but they are not always visible in a routine preparation.
- The large clear central vacuolar area occupies most of the cell interior. Mature plant-cell vacuoles can occupy a major proportion of the complete cell volume.
- Fine adjustment is required at this magnification. The selected cells should be centred before changing to the 40× objective.
- Folded peel, thick underlying tissue and uneven staining have a stronger effect at higher power. Only a narrow depth of the specimen remains sharply focused at one time.
- The cell wall, stained nucleus, peripheral cytoplasm and central vacuolar region may be recognised. It should not be stated that mitochondria, ribosomes, Golgi bodies and all other organelles become visible at 400×. Many of these structures need special staining or advanced microscope methods.
| Total magnification | Typical view | Structures likely to be recognised | Main limitation |
|---|---|---|---|
| 40× | Large sheet containing many rows of small rectangular cells | General tissue arrangement and broad cell-wall outlines | Internal cell details are mostly indistinct. |
| 100× | Fewer and more distinct individual cells | Cell walls, cell shape and sometimes stained nuclei or peripheral cytoplasm | The nucleus may still remain faint. |
| 400× | Detailed view of a few cells | Cell wall, stained nucleus, peripheral cytoplasm and large central vacuolar area | Small field, fine focus and good staining are required. Not all organelles can be seen. |
The exact view can change with the microscope, onion variety, stain, illumination and preparation quality. Low power is used first, and then a clear group of cells is examined under high power.
Structures Visible in an Onion Epidermal Cell
The structures observed depend on staining, focusing and the quality of onion peel. A routine compound microscope mainly shows the larger cellular parts. Very small organelles are not separately visible.
Cell Wall and Plasma Membrane
- The cell wall is the most conspicuous structure of onion epidermal cell. It forms a thick-looking rectangular boundary around every cell.
- Onion epidermal walls contain cellulose microfibrils as a major structural part. Pectins and hemicelluloses are also present in the wall matrix.
- The wall gives a fixed shape and mechanical support to the cell. It also resists the pressure formed by water inside the cell. Onion epidermal wall is strong but it can expand during cell growth.
- The plasma membrane is present immediately inside the cell wall. It surrounds the living content or protoplast of the cell.
- In a normal turgid onion cell, the plasma membrane remains pressed closely against the wall. Therefore, the two boundaries are difficult to distinguish in a simple water-mounted slide.
- The membrane can be demonstrated more clearly during plasmolysis. In a concentrated solution, water moves from the cell and the protoplast becomes smaller. The plasma membrane then separates from some regions of the cell wall.
Cytoplasm, Nucleus and Central Vacuole
- Cytoplasm– The cytoplasm is the living material present within the plasma membrane. It contains the different cellular components.In mature onion epidermal cells, cytoplasm occurs mainly as a thin peripheral layer. It surrounds the large vacuole and may also form narrow strands across it.Cytoplasm is transparent and sometimes difficult to identify without staining. It can look like a thin layer present near the cell wall.
- Nucleus– The nucleus is generally rounded or oval in shape. In an unstained preparation it may remain faint.After suitable staining, the nucleus takes a darker colour than its surrounding cytoplasm. It is then easier to recognise.The nucleus is commonly positioned toward the side of the cell. This peripheral position occurs because the large central vacuole occupies much of the cell interior. Studies of onion epidermis also show that its nuclei may have grooves and internal folds, but these fine structures are not visible in an ordinary school slide.
- Central Vacuole– The central vacuole occupies most of the volume of a mature onion epidermal cell. Under a light microscope, it generally appears as a large and clear central region.It contains cell sap, which consists mainly of water with dissolved ions, sugars and other substances. In red onion, anthocyanin pigment may also be stored in the vacuole.The vacuole takes part in storage of water. Its enlargement also contributes to cell expansion and maintenance of the internal turgid condition. A vacuolar membrane called the tonoplast surrounds it. The tonoplast has been demonstrated in onion epidermal cells by fluorescent labelling and confocal microscopy. It is not normally resolved as a clear separate boundary in a routine school preparation.
Why Are Chloroplasts Absent?
- The onion peel generally used in the experiment is taken from the inner epidermis of a fleshy bulb scale.
- These bulb scales are mainly storage tissues and remain nonphotosynthetic. Their transparent epidermal cells do not normally show visible green chloroplasts.
- It does not mean that every cell of the onion plant lacks chloroplasts. The green aerial leaves of Allium cepa contain photosynthetic tissues and chloroplasts.
- Different types of plastids occur in onion tissues. Research on onion has also reported chloroplasts in some cells situated near vascular bundles of white fleshy leaves. Thus, the absence is mainly related to the transparent inner bulb epidermis selected for the practical.
- Lack of numerous green chloroplasts keeps the bulb epidermal cells nearly transparent. The cell wall, stained nucleus and peripheral cytoplasm are not hidden by many chloroplast bodies.
Which Stain Is Used for Onion Cells?
Onion epidermal cells are nearly colourless. A stain is used to increase contrast between the cell structures and surrounding liquid. Iodine solution, safranin and methylene blue can be used. There is not only one correct stain for every onion peel experiment.
Iodine or Lugol’s Iodine
- Iodine in potassium iodide solution, also called Lugol’s iodine, is commonly used in beginner microscopy. It is added below the coverslip or directly over the epidermal peel.
- The iodine increases contrast. Onion cells generally get a yellow-brown colour and some regions may appear darker brown.
- The nucleus and peripheral cell material can become easier to locate. Iodine-potassium iodide has also been used for darkening nuclei in onion bulb epidermis.
- Iodine should be added in a small controlled amount. A drop can be drawn below the coverslip by keeping filter paper at its opposite edge.
- Excess iodine is removed using filter paper. When too much stain remains, the specimen gets a nearly uniform dark colour and internal differences become less clear.
- Any blue-black granule produced by iodine represents starch. It should not be confused with the nucleus. Starch grains are not always present in onion epidermal preparations.
Safranin
- Safranin is another stain commonly given in school practical work. NCERT cell observation activity uses safranin during preparation of a temporary onion-peel mount.
- It gives pink to red colour to the stained cell material. The cell boundaries become easier to follow. A properly stained nucleus may also look more conspicuous.
- Safranin does not colour every part with equal intensity. The final colour depends upon concentration of stain, condition of peel and the exposure period.
- After staining, extra safranin should be washed or blotted away. This gives a lighter background and prevents the entire peel becoming deep red.
- A very concentrated solution or longer staining time can hide the cytoplasmic region and nucleus. Dilute stain is generally sufficient for this temporary preparation.
Methylene Blue
- Methylene blue produces a blue contrast. Nuclear material usually takes a darker blue colour than the surrounding cell material.
- This stain is more commonly associated with human cheek-cell practicals. In this preparation, greater stain uptake by the nucleus makes it dark blue while the cytoplasm remains pale blue.
- Methylene blue can also be used for onion epidermal preparations. An IGNOU laboratory manual gives methylene-blue staining of onion epidermal cells for about 2 to 3 minutes, followed by removal of extra stain.
- The stain may make the nucleus more conspicuous and gives some contrast to peripheral cytoplasm. It should not be described as the only correct stain for onion cells.
- Excess methylene blue produces a strongly blue field. The peel is then washed or extra solution is drawn away with filter paper.
Stained vs Unstained Cells
| Feature | Unstained onion cells | Stained onion cells |
|---|---|---|
| Overall colour | Transparent or nearly colourless. | Yellow-brown with iodine, red or pink with safranin and blue with methylene blue. |
| Cell-wall visibility | Cell outlines can be recognised under correct light, but contrast may be low. | Boundaries generally look more distinct against the coloured cell material. |
| Nuclear visibility | The nucleus may remain faint or completely unnoticed. | It commonly appears darker and is easier to locate. |
| Cytoplasmic contrast | The thin peripheral cytoplasm is difficult to separate from the clear cell interior. | Peripheral material may become more noticeable, depending on stain uptake. |
| Vacuolar appearance | The large central vacuolar region looks clear or empty. | It normally remains paler than the stained nucleus and surrounding cytoplasm. |
| Advantage | Living cells can be observed with little chemical interference. | The main cell structures are easier to recognise and draw. |
| Limitation | Internal structures have very low contrast. | Overstaining may make the slide uniformly dark and hide the structures. |
Observation and Result
Observation
The following observations were recorded from the stained temporary mount of onion epidermis-
- Shape: The cells were rectangular or elongated in shape. Some cells appeared more or less brick-like. Onion bulb epidermal cells commonly show a rectangular form in the plane of the tissue.
- Arrangement: The cells were closely arranged and present side by side. They formed a continuous sheet with a regular row-like pattern.
- Cell wall: A distinct cell wall was observed around each cell. It formed the clear boundary and maintained the definite shape of the cells.
- Cytoplasm: The cytoplasm appeared as a thin peripheral layer near the inner side of cell wall. It was not equally clear in all cells.
- Nucleus: A stained nucleus was seen as a round or oval and darker body. It was generally present toward the side of the cell. The actual nucleus can have a more irregular surface, but this is not normally resolved in a routine school preparation.
- Vacuole: A large and clear central vacuolar region occupied most part of the cell. Due to this large vacuole, cytoplasm and nucleus remained mainly near the cell periphery.
- Chloroplasts: Green chloroplasts were not normally observed. The inner bulb epidermal cells are nonphotosynthetic cells and chloroplasts generally do not develop in this tissue. This should not be applied to the green leaves of onion plant.
- Intercellular spaces: Large intercellular spaces were not conspicuous in the mounted view. The cells appeared closely fitted with their neighbouring cells. Small spaces or damaged regions may occur when the peel is torn or folded.
The nucleus and cytoplasm may remain faint when staining or focusing is not proper. A clear slide gives the better observation of these parts.
Result
A sheet of closely arranged plant epidermal cells was observed under the compound light microscope.
Each cell had a definite rectangular or elongated boundary formed by the cell wall. A stained nucleus, thin peripheral cytoplasm and a large central vacuolar region could be identified in the properly prepared specimen.
The observed characters are consistent with onion epidermal cells. They show the common structural features of mature plant cells, but all smaller cell organelles were not visible by the routine preparation.
Onion Peel Compared With Other Common Microscope Specimens
Onion Peel Cell vs Human Cheek Cell
| Feature | Onion peel cell | Human cheek cell |
|---|---|---|
| Organism | It is obtained from onion, Allium cepa L. | It is obtained from human, Homo sapiens. |
| Cell type | A plant epidermal cell from the fleshy bulb scale. | A squamous epithelial cell obtained from inner cheek surface. Human buccal mucosa consists of non-keratinized stratified squamous epithelium. |
| Shape | The cells are elongated, rectangular or brick-like. Onion epidermal cells show a more or less rectangular shape in the tissue plane. | The cells are flat and generally irregular or polygonal. The superficial cheek cells have a squamous form. |
| Arrangement | Cells are closely joined. They occur in regular rows and form a continuous sheet. | Cells are released during scraping and spread separately over the slide. Some cells may overlap. |
| Cell wall | A distinct cell wall is present. It produces the fixed and regular boundary. | Cell wall is absent. |
| Plasma membrane | The plasma membrane is present immediately inside the cell wall. It is difficult to separate from the wall in a normal turgid cell. | The plasma membrane forms the outer cell boundary. This boundary is flexible and less regular. |
| Nucleus position | The nucleus is generally toward the side. The large vacuole occupies much of the cell and pushes the nucleus into peripheral cytoplasm. | The stained nucleus is commonly seen near the central region of the flattened cell. Its exact position may vary. |
| Vacuole | A large central vacuole is present. | A large central vacuole is absent. |
| Chloroplasts | Chloroplasts are not normally visible in the colourless inner bulb epidermis. | Chloroplasts are absent because it is an animal cell. |
| Recommended stain | Iodine-potassium iodide solution is commonly used. Safranin can also be used. | Methylene blue is commonly used to increase the contrast of nucleus. |
| Main educational use | It is used for studying the cell wall, regular plant-cell arrangement, nucleus and large vacuolar region. | It is used for studying a basic animal cell and comparing plant and animal cells. |
Onion Peel Cell vs Onion Root-Tip Cell
| Feature | Onion peel cell | Onion root-tip cell |
| Tissue type | It belongs to the mature epidermal tissue of a fleshy bulb scale. | It belongs mainly to the actively growing root apical meristem. |
| Cell condition | The cells are differentiated and perform a protective surface role. | The meristematic cells remain capable of active cell division. |
| Cell size and shape | Cells are comparatively large, elongated and rectangular. They form a flat epidermal sheet. | Cells are generally smaller and more compact. Their shape is less elongated in the dividing region. |
| Main observation | It is used for studying ordinary plant-cell structure. The cell wall, nucleus and vacuolar area can be identified. | It is used mainly for studying mitosis and chromosome behaviour. Onion root tips show interphase, prophase, metaphase, anaphase and telophase cells. |
| Preparation method | A thin epidermal peel is removed and mounted directly. No squash preparation is normally required. | The root tip is fixed, stained and generally prepared by the squash technique. Squashing spreads the meristematic cells. |
| Nucleus during interphase | A stained round or oval nucleus is usually present toward the side of the cell. | An interphase nucleus is also present in many root-tip cells. It may occupy a larger proportion of the smaller meristematic cell. |
| Chromosomes | Condensed individual chromosomes are not normally visible in an ordinary peel cell. | Condensed chromosomes become visible in the cells passing through mitosis. Their arrangement changes with different stages. |
| Mitotic stages | Mitotic stages should not be expected in a normal bulb epidermal peel. A darkly stained nucleus is not automatically a prophase nucleus. | Several mitotic stages can be found because the root meristem contains actively dividing cells. |
| Main educational use | Used to demonstrate basic plant-cell structure and epidermal arrangement. | Used to demonstrate the cell cycle, mitotic stages and chromosome separation. |
Stained vs Unstained Onion Cells
| Feature | Unstained onion cells | Stained onion cells |
| Overall contrast | Contrast is low. The cells are transparent or nearly colourless. | Contrast is increased. The cellular regions appear darker against the background. Iodine is used to make onion-cell structures easier to see. |
| Cell-wall visibility | Cell-wall outlines can be seen under correct illumination. They may look faint when excess light is used. | Cell boundaries generally appear more distinct due to increased contrast. |
| Nuclear visibility | The nucleus may remain faint or cannot be located in some cells. | The stained nucleus generally becomes darker and is more easily recognised. Iodine-potassium iodide can be used to darken onion epidermal nuclei. |
| Cytoplasmic contrast | The peripheral cytoplasm is difficult to separate from the clear central region. | The thin cytoplasmic region may become more noticeable. It is not equally stained in all cells. |
| Natural colour | The natural pale or colourless appearance is retained. | The natural appearance is changed by the selected stain. Iodine gives yellow-brown colour, safranin gives pink or red and methylene blue gives blue contrast. |
| Vacuolar appearance | The large vacuolar region looks clear and sometimes appears empty. | It generally remains lighter than the stained nucleus and peripheral cell material. |
| Risk of artefacts | Stain deposits are absent. Folds, drying and air bubbles can still affect the observation. | Excess or uneven stain can make the specimen too dark. Extra iodine is normally drawn away or absorbed with paper. |
| Suitable observation | It is suitable for observing the natural cell outline and living-cell changes, including plasmolysis when a suitable solution is added. | It is more suitable for identifying the nucleus and other low-contrast cell regions during a beginner practical. |
| Main limitation | Internal structures remain difficult to identify. | Staining can change the natural appearance and does not make every organelle visible. |
Common Problems and Troubleshooting
Some problems occur due to improper slide preparation. Others are related with focusing, illumination or staining. The following problem–cause–solution tables are used for correcting them.
Cells Are Blurry
| Problem | Possible cause | Solution |
|---|---|---|
| The complete field remains blurred. | The specimen is not focused correctly. | Begin with the low-power objective. Use coarse adjustment first and then fine adjustment until the cell walls become sharp. |
| Dark marks or hazy areas move with the slide or eyepiece. | The slide, coverslip, eyepiece or objective lens is dirty. | Clean the slide and coverslip. Optical surfaces should be cleaned only with lens paper and suitable lens cleaner. |
| Some cells are clear, while other layers remain blurred. | Thick underlying onion tissue was included with the peel. | Prepare another slide using only the thin epidermal membrane. Thick specimens contain different focal planes and produce more out-of-focus detail. |
| Cell outlines cross over each other. | The epidermal peel is folded. | Remove the coverslip when possible and spread the peel into one flat layer. A badly folded specimen should be replaced. |
| The peel moves and cannot be focused properly. | Too much water, glycerine or other mounting medium is present. | Draw excess liquid from the coverslip edge with blotting paper. Do not press over the specimen. Excess liquid below a coverslip should be removed carefully. |
| Only part of the field is illuminated or clear. | The objective is present between two positions on the revolving nosepiece. | Rotate the nosepiece until the selected objective clicks completely into the light path. |
Large Circular Structures Cover the View
| Problem | Possible cause | Solution |
| Large round structures with dark edges are seen. | These are generally trapped air bubbles, not onion cells or cell organelles. Air bubbles in wet mounts appear rounded and may have a dark boundary. | Observe another region when only one or two bubbles are present. |
| Many bubbles occur below the coverslip. | The coverslip was dropped directly over the water or mounting medium. | Keep one edge of coverslip in contact with the drop. Lower its opposite side slowly at an angle by using a needle. This method reduces trapping of air. (isu.edu) |
| Bubbles cover the main specimen area. | The mounting procedure was not proper or insufficient liquid remained below the coverslip. | Prepare another slide. A new mount is better when bubbles hide most onion cells or disturb focusing. |
Cells Appear Too Dark
| Problem | Possible cause | Solution |
| The complete peel looks uniformly dark. | Excessive stain was added. | Draw extra stain from one coverslip edge with blotting paper. The peel can also be rinsed before mounting. Standard onion-peel methods remove excess stain before observation. |
| The background and cells have nearly the same deep colour. | The stain solution is too concentrated. | Use a more dilute stain or prepare a fresh mount. A controlled amount should provide contrast, not colour every region equally. |
| The nucleus and cytoplasm cannot be separated. | The peel was exposed to stain for a long period. | Reduce the staining time. Rinse away the free stain before adding mounting medium. |
| Dark overlapping bands are present. | The peel is thick or folded into several layers. | Select a thin single epidermal layer. Spread it flat before keeping the coverslip. |
| Removing stain does not improve the view. | The specimen has become heavily overstained or damaged. | Prepare a fresh peel and use less stain for a shorter period. Staining is used for increasing contrast, but too much coloured material reduces useful differences in the field. |
The Nucleus Cannot Be Seen
| Problem | Possible cause | Solution |
| Cell walls are visible but the nucleus is absent. | No stain was used or contrast is insufficient. | Add a suitable stain such as dilute iodine, safranin or methylene blue according to the practical method. Stains increase contrast in nearly transparent specimens. |
| The nucleus looks too small to identify. | Observation is being made only under low magnification. | Locate the cells under low power. Centre a clear region and then change to the higher-power objective. |
| The whole cell appears soft and unclear. | The microscope is not focused on the plane containing the nucleus. | Turn the fine adjustment slowly. The nucleus may lie slightly above or below the plane where the wall appears sharp because a microscope has a limited depth of field. |
| The transparent specimen looks washed out. | Illumination is excessive or the diaphragm is opened too far. | Reduce the light and adjust the iris diaphragm until the cell contents show better contrast. Do not close it completely. |
| The complete interior appears very dark. | The specimen is overstained. | Remove excess stain using blotting paper or prepare a fresh lightly stained mount. |
| The nucleus is not found in the current cell. | It is outside the present focal plane or hidden by a fold. | Use fine focus and inspect another properly spread cell. Not every nucleus appears equally distinct in one focal setting. |
| Cells look shrunken, broken or colourless. | The peel was damaged or allowed to dry before mounting. | Use a fresh moist peel. Keep it in water during preparation and observe it before the mount starts drying. Wet specimens can lose image quality and living condition after drying. |
Cells Overlap or Appear Distorted
| Problem | Possible cause | Solution |
| Two or more cell patterns are present over each other. | The epidermis is folded. | Spread the peel with a fine needle or brush before lowering the coverslip. Use a fresh piece when the fold cannot be removed. |
| Several cell layers remain in one region. | Multiple pieces of peel were placed together. | Mount only one small epidermal piece. It should form a single flat layer. |
| Cells appear compressed or crushed. | Excess pressure was applied over the coverslip. | Do not press the coverslip. Lower it gently and blot liquid only from its edges. Pressure can crush or distort a wet-mounted specimen. |
| Cell walls are broken and the arrangement is irregular. | The peel was torn by forceps, blade or needle. | Lift a new peel gently from its edge. Avoid scraping the epidermis from the fleshy scale. |
| A thick irregular mass is present below the cells. | Fleshy underlying bulb tissue was included accidentally. | Remove only the transparent inner epidermal membrane. Thick tissue should be discarded and another thin peel is prepared. |
Precautions
The following precautions should be followed during onion peel experiment-
- A fresh and thin onion epidermal peel should be selected. Thick fleshy tissue below the epidermis should not remain attached.
- Do not use the dry and papery outer onion skin. The transparent epidermis from the inner surface of a fresh fleshy scale is used.
- Keep the epidermal peel moist from the time of removal. It should be placed immediately in water. Drying can damage the cells and makes the peel difficult to spread.
- A clean glass slide and coverslip should be used. Water, dust or old stain over their surfaces can interfere with observation.
- Spread the peel gently into a flat single layer. Folds and overlapping regions produce an unclear image.
- Only a small amount of suitable stain should be added. Excess stain makes the complete preparation dark and the cellular parts become less distinct.
- Extra water or stain is removed carefully from the coverslip edge by using filter paper. Do not allow the liquid to reach the objective lens.
- Lower the coverslip gradually at an angle with the help of a mounted needle. This reduces the formation of trapped air bubbles.
- The coverslip should not be pressed strongly. Pressure may displace, crush or distort the onion epidermal cells.
- Observation is started with the lowest-power objective. A clear part of the peel is located and centred first.
- Under high power, only the fine adjustment knob should be used for obtaining the clear details. Sudden coarse focusing is avoided.
- Do not permit the objective lens to strike the slide or coverslip. The lens position should be checked from the side while bringing it near the slide. Otherwise the coverslip can break and the objective may be damaged.
- Blade or scalpel should be handled carefully and only under proper instruction. Stains are used according to their safety information. Eye protection should be worn where required, especially when iodine solution can splash. Broken glass should not be collected with bare hands.
- Hands should be washed properly after completing the experiment and after removal of gloves. Any stain present on skin is washed without delay.
Uses of Onion Epidermal Cells in Biology
Onion epidermal cells are used as a common experimental material in plant biology. The peel is thin, transparent and formed of large cells. Some of the important uses are-
- Plant Cell Structure– Onion epidermal cells are used to study the basic structure of a plant cell. The cell wall, stained nucleus, peripheral cytoplasm and large central vacuolar region can be observed in a properly prepared slide.
- Microscope Training- It is used for learning the correct use of a compound microscope. Students can practise slide preparation, staining, low-power focusing and higher-power observation.
- Cell Measurement– The large onion epidermal cells are suitable for measurement of cell length and width. It is also used for learning microscope scale, field diameter and calculation of actual cell size.
- Osmosis Study– Onion peel is used to demonstrate the movement of water across the plasma membrane. The cells are placed in solutions having different concentrations and their changes are then observed.
- Plasmolysis– The epidermal cells are commonly used for studying plasmolysis. In a concentrated solution, water moves out from the cell and the protoplast separates from the cell wall. Deplasmolysis can also be observed after placing it again in water.
- Membrane Observation– In a normal turgid cell, the plasma membrane remains closely attached with the cell wall. During plasmolysis, its separation becomes more clear. Thus, onion peel is used for differentiating the cell wall and plasma membrane.
- Cell Wall Study– Onion epidermis is used for studying the structure and mechanical properties of primary plant cell wall. Cellulose arrangement, wall stretching and changes in wall strength can be examined by different microscopic methods.
- Cell Expansion– These cells are used for studying the process of plant cell enlargement. The relation between cell-wall arrangement and elongated cell shape can be observed in onion bulb epidermis.
- Turgor Pressure– Onion cells are used in experiments related with turgor pressure and cell stiffness. Their response during water gain and water loss gives information about the combined function of cell wall and vacuole.
- Gene Expression – Onion epidermal cells are used for transient gene-expression studies by introducing foreign DNA into the cells.Protein Localisation – Fluorescent proteins such as green fluorescent protein (GFP) are used to locate proteins in nucleus, cytoplasm or plasma membrane.Stain Testing – The transparent peel is used for testing biological stains and fluorescent probes in different cell regions.
Further Experiments With Onion Epidermis
Onion epidermis can also be used for some additional experiments. These are related with osmosis, natural pigment and cell measurement.
Plasmolysis in Salt or Sugar Solution
- Turgid Cell– In water, the onion cell remains turgid. The vacuole is filled and the protoplast remains pressed against the cell wall.
- Hypertonic Solution– The peel is placed in a concentrated salt or sugar solution. This solution has lower water potential than the cell sap.
- Water Loss– Water moves out from the cell by osmosis. The vacuole and cytoplasm decrease in volume.
- Plasmolysis– The protoplast becomes smaller and pulls away from the cell wall. The rigid cell wall still maintains its external shape.
- Deplasmolysis– The plasmolysed peel is again placed in water. Water enters the cell and the protoplast may return near the wall, when the cell is not permanently damaged.
Red Onion Epidermal Cells
- Vacuolar Pigment– Red onion cells may contain anthocyanin inside the vacuole. It produces red or purple colour.
- Natural Contrast– The pigment gives natural contrast. The vacuole and protoplast can be observed without adding a strong artificial stain.
- Pigment Variation– Pigmentation is not same in every bulb layer or epidermal surface. Outer red layers generally contain more anthocyanin.
- Inner Epidermis– The inner epidermis may remain pale or nearly colourless. Every red onion peel does not show strong pigmentation.
- Plasmolysis View– Red onion epidermis is useful for plasmolysis observation. The coloured vacuole makes the shrinking protoplast easier to follow.
Estimating Onion Cell Size
- Field Method– Determine the diameter of microscope field. Count the number of cells present across it and divide the field diameter by this number.
- Graticule Method– An eyepiece graticule can be used for measuring the length or width of onion cells.
- Calibration– The eyepiece graticule is calibrated with a stage micrometer. Calibration should be done at the same objective magnification.
- Approximate Value– Several cells should be measured because all onion cells are not exactly equal in size. An approximate mean is then calculated.
- Result Reporting– Cell size is reported in micrometres (µm). The magnification and measurement method should also be mentioned.
Classroom Viva Questions
| Question | Answer |
|---|---|
| Why are forceps used in this experiment? | Forceps are used to lift the delicate epidermal peel without touching it by hand. |
| What is the function of a watch glass? | It is used to keep the peel in water or stain and prevents its drying. |
| Why should thick onion tissue be removed? | Thick tissue blocks transmitted light and produces overlapping focal planes. |
| What happens when the peel is folded? | Cells overlap with each other and a clear single-layer view is not obtained. |
| Why is a small piece of peel selected? | A small peel can be spread and mounted more easily below the coverslip. |
| What is the use of a mounted needle? | It is used to spread the peel and lower the coverslip slowly. |
| Why is blotting paper used? | It removes excess stain or mounting liquid from the coverslip edge. |
| What happens when excess mounting liquid is present? | The coverslip may float and the specimen can move during focusing. |
| Why should the coverslip not be pressed? | Strong pressure can crush, tear or distort the epidermal cells. |
| What indicates a properly mounted peel? | The peel remains flat, moist and free from major folds or air bubbles. |
| How is a suitable viewing area selected? | A flat region having clear rows of cells and no thick tissue is selected. |
| What is the function of the diaphragm? | It controls the amount of light passing through the specimen. |
| Why is excess illumination avoided? | Excess light reduces contrast and makes transparent cell parts difficult to see. |
| Why should the objective click into position? | A correctly positioned objective remains aligned with the light path. |
| What happens when the objective is between two positions? | The field may become dark, partly illuminated or unclear. |
| Why is the specimen centred before high power? | High power gives a smaller field, so an uncentred specimen may disappear from view. |
| What is the use of coarse adjustment? | It is used for initial focusing under the low-power objective. |
| Why is coarse adjustment avoided under high power? | It may move the objective into the coverslip and damage the slide or lens. |
| How can an air bubble be recognised? | It appears circular with a dark border and does not show rectangular cell walls. |
| Why should the slide remain clean? | Dust, fingerprints and old stain can be mistaken for specimen structures. |
| What happens when the peel dries during observation? | The cells may shrink, become distorted and lose their normal appearance. |
| Why is a fresh onion preferred? | Its epidermis separates easily and the cells remain in a better hydrated condition. |
| What should be done when the slide is uniformly dark? | Excess stain should be drawn out or a fresh lightly stained mount is prepared. |
| What should be done when only one part is focused? | Fine adjustment is used slowly because different parts may occur at slightly different focal planes. |
| Why should the blade be handled carefully? | It has a sharp edge and can cause injury during cutting of the onion scale. |
Frequently Asked Questions
1. What does onion peel look like under a microscope?
Onion peel shows elongated or rectangular cells arranged side by side. The cells appear like a brick wall, with distinct cell-wall boundaries.
2. What type of cells are present in onion peel?
Onion peel contains eukaryotic plant epidermal cells. These cells have a cell wall, plasma membrane, cytoplasm, nucleus and a large central vacuole.
3. Why are onion cells rectangular?
The firm cell wall maintains their definite shape. Directional cell expansion and close arrangement of the epidermal cells also produce the elongated rectangular pattern.
4. Which onion layer should be used for the experiment?
Use the thin transparent epidermis from the inner concave surface of a fresh fleshy bulb scale. The dry brown outer skin should not be selected.
5. Why is onion peel suitable for microscopy?
The peel is thin, transparent and easy to remove. It provides a flat layer of comparatively large cells without complicated tissue sectioning.
6. What structures can be seen in an onion cell?
The cell wall, stained nucleus, peripheral cytoplasm and large central vacuolar region can generally be identified. Visibility depends on stain, focus and slide quality.
7. Can the cell membrane be seen clearly?
The plasma membrane is usually difficult to distinguish in a normal turgid cell. It remains closely pressed against the inner side of cell wall. It becomes more clear during plasmolysis.
8. Why is the nucleus near the edge of the cell?
A large central vacuole occupies much of the cell volume. It pushes the cytoplasm and nucleus toward the cell periphery.
9. Why does the vacuole appear empty?
The vacuole contains mostly transparent cell sap. Its membrane or tonoplast is also not resolved clearly in a routine school preparation, so the region looks empty.
10. Do onion peel cells contain chloroplasts?
The inner bulb epidermal cells normally do not show visible chloroplasts. This statement applies to the non-green bulb peel, not to every onion tissue.
11. Why are chloroplasts absent from onion bulb epidermis?
The examined bulb scales are mainly nonphotosynthetic storage tissues. Green onion leaves contain photosynthetic tissues and chloroplasts, but the inner bulb epidermis generally does not.
12. Can onion cells be observed without staining?
Yes. The cell shape, arrangement and cell-wall outlines can be observed without staining. The nucleus and peripheral cytoplasm may remain faint.
13. Why is iodine added to onion peel?
Iodine solution increases contrast in the transparent specimen. It makes the cell contents and nucleus easier to locate.
14. Can safranin be used instead of iodine?
Yes. Safranin can be used as a plant-cell stain and gives pink or red contrast. The excess stain should be removed before observation.
15. Can methylene blue be used for onion cells?
Yes. Methylene blue can provide blue contrast and may make nuclear material more conspicuous. It is more commonly used for cheek cells, but iodine is not the only possible onion-cell stain.
16. What magnification is required to see the nucleus?
A stained nucleus may begin to appear at about 100× total magnification. It is generally easier to recognise near 400×, using correct illumination and fine focus.
17. What is visible at 40× magnification?
At 40×, a broad sheet containing many rows of rectangular cells is seen. Cell arrangement is clear, but internal structures generally remain indistinct.
18. What is visible at 100× magnification?
At 100×, individual cells and their cell walls become more distinct. A stained nucleus may begin to appear in some properly focused cells.
19. What is visible at 400× magnification?
At 400×, fewer cells are seen in greater detail. The stained nucleus, thin peripheral cytoplasm and large central vacuolar region may be recognised. Not all cell organelles become visible.
20. What is the observation of the onion peel experiment?
Closely arranged rectangular plant cells are observed. Each cell shows a distinct wall, and a properly stained specimen may show a nucleus, peripheral cytoplasm and large clear vacuolar region.
21. What is the final result of the experiment?
A continuous sheet of plant epidermal cells was observed. The visible structures and regular arrangement are consistent with mature onion bulb epidermis.
22. Why should the coverslip be lowered at an angle?
Lowering it slowly at an angle allows air to move out before the coverslip settles. It reduces the formation of air bubbles.
23. Why do air bubbles appear in the slide?
Air bubbles become trapped when the coverslip is dropped directly over the liquid or insufficient mounting medium is present. They appear as large circular structures with dark edges.
24. Why can I not see the onion cell nucleus?
The specimen may be unstained, poorly focused or supplied with excessive light. Overstaining, drying and the nucleus being outside the present focal plane can also make it invisible.
25. Is onion peel one cell thick?
The inner bulb epidermal peel generally separates as a single layer of intact cells. A preparation may look thicker when it is folded or contains attached underlying tissue.
26. What is the difference between onion peel and cheek cells?
Onion cells are rectangular plant cells with a cell wall and large central vacuole. Cheek cells are irregular or flattened animal epithelial cells, and a cell wall is absent.
27. What is the difference between onion peel and onion root tip?
Onion peel contains mature epidermal cells used for plant-cell structure. The root tip contains smaller meristematic cells and is used for studying mitosis through a stained squash preparation.
28. Do onion peel cells undergo mitosis?
Mature bulb epidermal cells are differentiated and do not normally show mitotic stages in the practical slide. Onion root-tip cells are selected when mitosis has to be observed.
29. What happens to onion cells in concentrated salt solution?
Water moves out from the cells by osmosis. The vacuole and protoplast shrink, and the plasma membrane pulls away from the cell wall. This is called plasmolysis.
30. Are red onion cells naturally coloured?
Some red onion epidermal cells contain red or purple anthocyanin inside the vacuole. Pigmentation varies between bulb layers and surfaces, so every inner peel is not strongly coloured.