1. Carry the microscope in an upright position with both hands and place the base of the microscope 5cm from the edge of the bench

Transcription

1 The Microscope Operating the compound light microscope 1. Carry the microscope in an upright position with both hands and place the base of the microscope 5cm from the edge of the bench 2. Check that lenses are clean and undamaged -Dirt on the slide: move the slide, the dirt will stay in the same place relative to the image -Dirt on the eyepiece: rotate the eyepiece and the image of the dirt rotates -Dirt on the objective lens: change the objective lens and the dirt disappears -The eyepiece and objective lens are cleansed using the lens cleaning tissue and 70% ethanol cleaning solution 3. Swing Low Power objective into line -The objectives are par-focal; changing the objective only requires a small focal adjustment with the fine focus 4. Switch on light source (0 = off, I = on) 5. Adjust light intensity (4-5 is adequate for most specimens) using the light adjustment 6. Position slide on stage, with the coverslip uppermost and centering over the light source 7. Focus on the specimen by rotating the coarse focusing knob. Watch from the side while raising the stage to 5mm from the tip of the LP objective then look down eyepiece while slowly lowering the stage to focus on specimen 8. Raise condenser and open iris diaphragm fully, move specimen aside 9. Lower the condenser using the condense knob while looking through eyepiece until a piece of paper on light source is in focus -Condenser lens Moving up and down, the condenser lens controls the focus of the light by controlling the diameter of the cone of light passing to the objective. The diameter must match the size of the lens of each objective to obtain maximum resolving power. If the condenser is too high, the cone of light does not fill the objective lens, lowering resolution. If the condenser is too low, some light is scattered outside the objective lens, lowering resolution.

2 10. Adjust iris diaphragm by removing eyepiece and closing the diaphragm until 3/4 of the field of view is illuminated -Iris diaphragm Iris diaphragm controls the amount of light reaching the objective lens by varying the angle of the cone of light which reaches the specimen and passes into the objective. Opening up = wide angular aperture = increase resolution, decrease contrast, brighter field of view Closing down = narrow angular aperture = decrease resolution, increase contrast, darker field of view It is adjusted every time the objective is changed. 11. Replace specimen, fine focus with the smaller central knob 12. Centre specimen 13. Position High Power objective 14. Adjust iris diaphragm 15. Fine focus Dissecting microscope Microscope theory -Magnification = enlargement of an image -Minimum resolved distance = minimum distance at which two objects can be identified as distinct from each other -The image seen through the eyepiece is inverted and magnified. Scale -Scale is used so the representation can be related to the size of the specimen rather than a magnification as it distorts the photo. -Horizontal scale bars have whole number units which represent about a 1/4 or 1/3 of the size of the drawn object -When the diameter of the field of view of an objective lens is known, the size of an object viewed on a slide can be determined by estimating the number of times the object fits across the diameter (usually 1mm/1000μm) -drawn size of object = drawn length of scale bar actual size of object actual length represented by scale bar Drawing -All drawings are done in pencil -All structures are labeled and indicated by a ruled label line, drawn parallel to the bottom of the page and without arrowheads -All drawings include a title at the top and a scale bar to indicate the size of the specimen -Use double lines to indicate hollow tube-like structures

3 Cell Structure Cell Theory: -A cell is the basic structural and functional unit of living organisms. So when you define cell properties you are defining the properties of life. -The activity of an organism depends on both the individual and the collective activities of its cells. -According to the principle of complementarity of structure and function, the biochemical activities of cells are dictated by the relative number of their specific subcellular structures. -Continuity of life from one generation to another has a cellular basis -Eukaryotic cell has membrane bound nucleus + organelles -Organelles (small organs) compartmentalize chemical (enzyme) reactions so they don t negate each other. -Cellular respiration: Glucose + O 2 cellular respiration CO 2 + H 2O + ATP -ATP when broken down to Adenosine diphosphate, releases energy Structures and functions of a cell Organelle Function Nucleus Nuclear membrane Nucleolus Cytoplasm Cytoskeleton Cell membrane (plasma membrane) Centriole Mitochondrion Rough endoplasmic reticulum Smooth endoplasmic reticulum Golgi body Lysosome Ribosome (organelle) Directs cell's life processes, contains the chromosomes, contains one or more nucleoli. -Separates the nucleus from the cytoplasm. Controls the highly-selective two-way exchange between the nucleus and cytoplasm. -Contains nuclear pores which controls mrna movement from nucleus to cytoplasm Ribosomal RNA is synthesised and combined with proteins. Ribosomes assemble Semi-fluid material (Cytosol) filling the space between plasma and nuclear membranes in which the organelles are found. Cellular scaffolding/ skeleton: -Microtubules: produced by centrioles, found in cilia and flagella -Intermediate filaments -Microfilaments -Regulates movement of substances into and out of the cell. -Phospholipid bilayer: hydrophilic phosphate molecules form the outer layer while the hydrophobic lipid tails are towards the middle clear distinction from the extracellular environment to prevent the high concentration of proteins from being dissipated by the water -Protein channel: passive channel which doesn t require energy when channeling ions or molecules Produces microtubules during mitosis and meiosis Produces ATP (Adenosine triphosphate) by cellular respiration Site of protein synthesis and transport. Site of lipid synthesis to allow detoxification of poisons (drug, alcohol) Storage, modification and packaging of secretory products. Contains digestive enzymes; cellular digestion. Translation of mrna to form protein precursors; protein synthesis

4 Surface area to volume ratios in relation to cells -The rate of diffusion into or out of a cell decreases as the cell increases in size because there is less surface area per unit of volume across which to exchange substances. -The maximum size to which a cell can grow is determined by the rate of diffusion of nutrients and gases across the cell surface. A cell can increase the maximum size to which it can grow by developing an intracellular transport system (membrane systems, e.g. ER), compartmentalise processes (organelles) -Heat loss per unit surface area is equal in the two animals, but the smaller animal will lose heat at a faster rate relative to its volume because the surface area to volume ratio (SA:V) is increasing with decreasing size. Likewise, larger cells have a lower SA:V ratio so their overall heat loss relative to surface area will be lower than that of smaller cells. -The principle also applies to substances that are transported in organism, or across a cell membrane (e.g. water, oxygen, wastes). Small cells have a large SA:V ratio compared to large cells. As cell size increases, the rate of movement of substances across the surface becomes insufficient to serve all the volume of the cell. Cell metabolism: the sum of all the chemical reactions in the cell -The energy released from the breakdown of food molecules is used to produce ATP (Adenosine triphosphate) -Aerobic respiration occurs when oxygen is available. The aerobic respiration of a glucose molecule produces 36 to 38 ATP molecules. When oxygen is not available anaerobic respiration occurs, producing (in animals) lactic acid. Structure of DNA Deoxyribonucleic acid (DNA) is the genetic material that organisms inherit from their parents; DNA determines the structural and functional characteristics of the cell by specifying the inherited structure of a cell s proteins. A long molecule of DNA (with associated proteins) makes up a chromosome, with sections of that DNA molecule forming genes. Each chromosome usually consists of between several hundred to more than 1000 genes. Gene = A sequence of nucleotides in a DNA molecule that specifies the structure of a protein or RNA molecule DNA exists in the nucleus of a cell and consists of a double strand (double helix structure) of building blocks called nucleotides

5