Cell: the basic unit of structure and function of all living organisms; a bag in which chemical reactions take place, surrounded by a partially permeable membrane.
Tissue: A group of cells adapted for a particular function.
Organ: A group of tissues adapted for a particular function.
Estimation of size from microscope observation: first, calibrate an eyepiece graticule (small measuring scale on a round piece of glass that goes into the eyepiece and has a small eyepiece unit scale) using a stage graticule (similar thing, but a rectangular glass that goes on the stage and has a micrometre scale). Align the two scales to find the micrometre value of a small eyepiece unit under different magnifications. Remember, don’t just take the value of one eyepiece unit- take the value of 100 and divide to reduce uncertainty! When looking at an object under the microscope, use the eyepiece graticule to find out how many small eyepiece units it is under a certain magnfication. You already know how many micrometres on small eyepiece unit is, so with a little calculation, you can find out how many micrometres your object is.
Calculation of size from scale diagrams:
Observed size = natural size x magnification
Observed is the size you measure from the diagram, and natural size is the object’s real size.
Prokaryotic cells lack nuclei and membrane bound organelles. Bacteria and blue green algae are prokaryotic. Prokaryotic cells range in size from about 0.1-2 μm in size.
Eukaryotic cells possess a nucleus and membrane bound organelles. Plant and animal cells are eukaryotic. Sizes range from 10-100 μm.
Microscopes can be used to determine the structure of cells by magnifying them so that they can be perceived by the human eye. There are two types- light and electron, which use beams of light and electrons respectively.
Resolving power is the shortest distance apart two objects can be without appearing to be one object when viewed under a microscope. An electron microscope has a much greater resolving power than a light microscope because an electron beam has a shorter wavelength than a beam of light.
Using a light microscope: under the light microscope, living specimens can be observed. For clearer observation, cells can be prepared by embedding them in wax, or staining them using coloured dye. Sections (thin slices) of cell a few μm thick are cut using a metal knife. Above all, the light microscope is cheap and easy to use.
Using an electron microscope: specimens must be viewed in a vacuum because air molecules would deflect the electron beam, so obviously living specimens cannot be observed. Cells are prepared by embedding them in resin or staining them with a heavy metal salt such as lead citrate. A diamond or glass knife is used to cut sections a few nm thick. The electron microscope is more expensive, but the higher resolution allows us to learn about the structure of many different cell organelles.
Transmission electron microscope: in a transmission e.m., a stream of electrons is sent from a cathode to an anode, passing through a thinly sliced specimen. The electrons are focussed at the anode, where they then land on a photographic plate. The number of electrons that get through indicates the density of the specimen. This type of electron microscope has a high resolving power.
Scanning electron microscope: in a scanning e.m., a solid specimen is bombarded with an electron beam. This cause secondary electrons to be emitted from the specimen’s surface, which then land on a photographic plate. The scanning e.m. is more versatile than the transmission e.m., as solid structures can be investigated.
Buffer solutions: during cell preparation, unnecessary damage to the cell must be avoided if we want to make accurate observations. A good way to protect cells is to place them in a buffer solution, which can do the following:
- Resisting changes in pH (to an extent) by “mopping up” excess H+ ions or dissociating to provide more H+ ions as required. See Chemistry section for more details.
- If the solution is of the same concentration of the cell, it will prevent water loss/gain by osmosis, which could disrupt the cell.
Artefacts are false structures created during the preparation of cells. Artefacts can be recognised by looking at multiple slides of the same type of cell, as they will not appear in all cells.
Cell Fractionation and Ultracentrifugation: this is a process in which a cell is taken apart in order to examine the functions of its organelles. I will explain the process using the extraction of mitochondria from liver cells as an example.
- Liver tissue is chopped into small pieces and packed in ice to prevent enzyme activity.
- A buffer solution is added to resist pH changes and prevent osmotic activity.
- A homogeniser is used to break up the cells, forming a “soup”.
- The mixture is now spun in an ultracentrifuge. A particular speed will provide enough force to pull the large, dense nuclei down to the bottom. The remainder of the “soup”, called the supernatant, is decanted and centrifuged at a higher speed to pull the mitochondria down to the bottom. The supernatant is decanted once again, and could be centrifuged at even higher speeds to pull down first lysosomes, then cell membranes. Note: because membrane-type structures such as Golgi apparatus and plasma membranes have similar densities, they get pulled down at the same speed and mixed up together during centrifugation, making them more difficult to study.
- The mitochondria have now been largely separated from the other organelles, but they must be purified. To do so, they are suspended in a buffered sucrose solution whose density increases with depth. When this tube is centrifuged, mitochondria are puled down to the level of the tube where the solution density equals their own. Mitochondria are now easily separated from the impurities, which are pulled down to different depths.
- The purified mitochondria can now be examined under an electron microscope.
Cell Membranes: the current theory is the fluid mosaic model, in which the cell membranes consists of a phospholipid bilayer, with the hydrophilic heads on the outside, protecting the hydrophobic tails from exposure to water. Proteins are embedded in the membrane. Cholesterol molecules interact with the phospholipids, stabilising the membrane andmaking it less fluid.
Exocytosis: A process in which substances from inside the cell are released outside it. A Golgi vesicle from inside the cell fuses with the cell membrane, releasing its contents outside the cell.
Endocytosis: A process by which substances are taken into a cell. When a substance approaches the cell membrane, the membrane forms a flask shaped depression around that substance. The ‘neck’ of the flask then closes, forming a vesicle around the substance.
Diffusion is the movement of molecules down a concentration gradient, i.e. from an area of high concentration to an area of lower concentration until dynamic equilibrium (molecules evenly spread and diffusing back and forth between the two areas at equal rates) is reached. Diffusion is a passive process- no energy is required.
Fick’s Law governs the rate of diffusion:
so for a maximum rate of diffusion, surface area and concetraion difference must be high, and membrane thickness must be low (see Gas Exchange section). Also, increasing the temperature will give diffusing particles more energy and so increase diffusion rate. However, high temperatures will also cause damage to living systems.
Simple diffusion is where substances diffuse directly across the phospholipid bilayer of the cell membrane. Small, uncharged molecules move across the plasma membrane by simple diffusion
Facilitated diffusion: larger, charged molecules, cannot travel across the phospholipid bilayer. They diffuse across via the proteins embedded in the cell membrane. This is facilitated diffusion, and there are two main types:
- Channel proteins are water-filled pores that allow the passage of water-soluble substances across the cell membrane. Channels are selective, and only a particular ion/molecule can pass through a certain type of channel. Some channels are gated- they open and close only when triggered (see The Nervous System).
- Carrier proteins are believed to combine with a diffusing ion or molecule, carry it across the membrane and release it on the other side.
Active transport: sometimes a substance needs to be transported against a concentration gradient (from low concentration to higher concentration). This process is active transport. ATP is used as an energy source to allow this transportation to take place (for more on ATP, see later). Substances are actively transported via carrier proteins.
Water potential (Ψ -psi) is the tendency for a system to lose water, measured in Pascals, Pa. The less concentrated a solution, the higher its water potential. This is because there are fewer solute molecules to restrict the movement of water molecules, so they have a high free kinetic energy and can leave the system easily. Pure water has the highest water potential (0). Water potential is therefore always ≤0.
Solute potential (Ψs) is the contribution made to the water potential by the addition of solute (also measured in Pascals). In most cases Ψs=Ψ. Therfore, solute potential is always ≤0
Pressure potential (Ψp): when enough water enters a plant cell, the cytoplasm swells, and pushes against the cell wall. The cell wall then pushes back- the exertion of pressure on and by the cell wall is known as pressure potential (measured in Pascals). Pressure potential is always ≥0.
Hypotonic: less concentrated.
Isotonic: of equal concentration.
Hypertonic: more concentrated.
Osmosis is the movement of water from a region of less negative (higher) water potential to a region of more negative (lower) water potential, i.e. from a less concentrated solution to a more concentrated one. Osmosis is basically a form of diffusion, so it is a passive process.
Osmosis and animal cells: e.g. red blood cells placed in solutions of varying concentration-
Osmosis and plant cells:
Water potential equation:
Ψ =Ψs + Ψp
Determination of plant cell concentration: place cells in solutions of varying concentrations. Theoretically, the solution just concentrated to make the plasma membranes of the cells separate from the cell wall (incipient plasmolysis) has a solute potential equal to that of the cell.
Practically, cells will all plasmolyse at different rates, so incipient plasmolysis is taken as the point where 50% of the cells are visibly plasmolysed. A solution that has this effect is assumed to be the same concentration as the cell.
Relationship between water, solute and pressure potential at different stages of turgor and plasmolysis:
The Blue Drop Experiment to discover concentration, e.g. of a potato:
- Immerse slices of potato in solutions of varying concentration and leave for a few hours
- If the potato is more concentrated than the solution it is in, water will move from the solution to the potato. This means the solution will become denser and more concentrated.
If the potato is less concentrated than the solution it has been placed in, water will leave the potato and enter the solution. This means the solution will become less concentrated and more dense.
- When ready, decant off the solutions into separate test tubes. Add a few drops of dye to each solution. Also, set up test tubes containing samples of the solutions at their original concentrations.
- Take a drop of dyed solution and inject it into the corresponding original solution. Repeat for each solution (e.g. if you have some dyed solution that was 4M at the beginning of the experiment, you would inject it into fresh 4M solution).
- If the solution has gained water from the potato and become less dense, it will rise when injected into fresh solution. If the solution has lost water to the potato and thus become more dense, the dyed drop will rise. If the solution is isotonic with the potato, it will have remained at the same concentration and density, and will not rise or fall. Note: the faster the rise/fall, the greater the change in density.
- The concentration of the potato is between the highest concentration at which the drop falls and the lowest concentration at which it rises. To more accurately find its concentration, either time the speed at which the drop rises/falls, and use a graph to extrapolate which concentration would produce no rise/fall; or simply do repeat experiments to narrow the possible range of concentrations.
Determination of concentration by looking at changes in length:
- Place small chip-shaped pieces of potato several cm long in solutions of varying concentration for a fixed period of time (at least 20 mins).
- After this time, measure the changes in length of the potato chips.
- If the potato is more concentrated than the solution it was in, it will have gained water, and so the chip gets longer.
If the potato is less concentrated than the solution it was in, it will have lost water, and so the chip will have become shorter. There will be no change if the potato and solution were of equal concentrations.
Plot a graph of change in chip length against concentration. Use the graph to determine the concentration at which there would be no change in potato length. This is the concentration at which there would be no net water movement because the potato and solution would be isotonic. Therefore, this must be the concentration of the potato.