BIO Lab 3
BIO 101 Lab 03: Microscopy and Cells NOTIFICATION: If you have a technology issue or disability that makes it difficult to complete this lab, please contact your instructor at least 2 days before the due date. For disabilities, please provide your instructor a copy of the Memorandum of Accommodation (MOA) from NVCC Disability Support Services. Objectives: • Identify the parts of compound and dissecting microscopes and explain their functions. • Focus the microscope using the scanning, low-, and high-power lenses. • Accurately describe prepped and wet mount specimens viewed using the compound light microscope. • Identify and explain the functions of the major cellular organelles • Explain similarities and differences between prokaryotic and eukaryotic cells (plant and animal cells) Microscope Care and Storage Microscopes are essential in many laboratories and should always be handled with care. A microscope should always be carried upright with one hand under the base and the other hand holding the arm. Care should be taken when placing a microscope down onto a table or into the storage cabinet. When the lenses become dirty, only special lens cleaner and lens paper should be used to prevent scratches. When done using a microscope, turn the light down and switch the power button to “off” before unplugging it. To store microscopes, all slides should be removed from the stage, the stage should be lowered, and the lowest powered lens should be in place. Taking these steps will ensure the usefulness of the microscope for a long time. If immersion oil is used for viewing using the 100X lens, then the lens should be wiped clean with special lens cleaner solution and lens paper before storage. Both large and small organisms make important contributions to our ecosystem. Our naked eye can easily see many plants and animals, but without the assistance of magnifying lenses, we cannot see the smaller organisms. Microscopes are the instruments we use to assist our eyes when viewing these minute specimens. There are different types of microscopes that serve different purposes. During this exercise you’ll learn to use a dissecting microscope and a compound microscope. Dissecting microscopes (Fig 1A) can be used to view larger objects that are not easily mounted on flat slides and have a maximum magnification of 40X. Compound microscopes (Fig 1B) are used to view objects that can be mounted on flat slides (not bulky), and have a maximum magnification of 1000X. Cells under the microscope The cell is the basic unit of life and living organisms are categorized as either prokaryotic or eukaryotic depending on the structure of their cells. Prokaryotic and eukaryotic cells can be quite different in size, shape, structure, and function. Prokaryotic cells do not have a nucleus, but instead have a nucleoid region in the cytoplasm where chromosomes (genetic material) are located. Prokaryotic cells are very small and are not visible under a dissecting microscope; most cellular structures are difficult to see even using 100x magnification with a compound light microscope. Eukaryotic cells have a collection of organelles that are responsible for the proper functioning of our bodies, including a nucleus which houses the chromosomes. Eukaryotes can be either single-celled or multi-cellular (made of many cells). Individual eukaryotic cells are often large enough to be visible under a light microscopy. Some organelles are also visible within the cytoplasm of these cells at higher magnification. 1 BIO 101 Lab 03: Microscopy and Cells Bacteria (Figure 2A) are an example of prokaryotic cells. Like other prokaryotes, bacteria do not have organelles, but can still contain various internal and external structures. Many eukaryotes are single-celled organisms and include protists (Figure 2B) such as amoebas (Figure 2B) and fungi such as yeasts (Figure 2C). The shape, size, and internal structure of single-celled eukaryotes is varied. Multi-cellular eukaryotes are composed of aggregation of cells, and include protists such as algae, fungi such as molds, all plants, and all animals. Table 1 below lists cellular structures and organelles of prokaryotic and eukaryotic cells that are visible under light microscopy. While cells contain many more structures than the ones listed, many of the structures are too small to be seen without the use of certain stains to highlight them or require much higher magnification to be visible. TABLE 1: Cellular structures visible under light microscopy and their functions Structure or Organelle Prokaryote or Eukaryote Cell membrane Both Cell wall Both Central vacuole Cilium (pl: cilia) Chloroplast Eukaryote Contractile vacuole Cytoplasm Eukaryote Flagellum (pl: flagella) Both Nucleus (pl: nuclei) Eukaryote Eukaryote Eukaryote Both Function Surrounds the cytoplasm; barrier between the environment and the cytoplasm; controls the movement of materials in and out of the cell. External support and protection; located outside of the cell membrane, when present; not found in animal cells. Water storage organelle in a mature plant cell; not found in animal cells. Hair-like projections from the cell membrane; assist cell in locomotion or movement of materials across the cell surface. Not found in plant cells. Organelle in which photosynthesis takes place. Contains chlorophyll and is green in color. Not found in most animal cells. Organelle that regulates fluid and waste inside cell; only present in some protists and lower animals. Viscous substance inside the cell membrane; all internal organelles are suspended in the cytoplasm. Long projections from the cell membrane; assist cell in locomotion and sensing. Different structure in prokaryotes and eukaryotes. Only present on sperm cells in plants, fungi, and animals. Organelle that stores genetic information (DNA and RNA) needed to control cell functions. 2 BIO 101 Lab 03: Microscopy and Cells THIS PROTOCOL SHOULD BE PRINTED OUT SO YOU CAN FOLLOW IT DURING LAB. For this lab, you will be working independently so everyone should have their own microscope. Make your own preps and do not share drawings or you will receive no credit. Materials: • • • • • Microscope Slides and cover slips Pond water Methylene blue dye Cotton swab • • • • Distilled water bottle 0.9% NaCl Toothpick Transfer pipette Safety: Follow all standard laboratory safety procedures. You will be working with glassware, chemicals, and microbiological agents. Use caution. Procedures: Activity 1: Operating a compound microscope Your instructor will go over the parts of the microscope before you begin. • • • Listen carefully as your instructor will walk you step-by-step for Activity 1. Be sure to label microscope parts (#1 on the answer worksheet) as you instructor describes each structure. Everyone must complete Activity 1 before moving on to activity 2. 1. Grab a microscope from the cabinet. Carry it properly by grabbing it by the neck with one hand and holding it under the base with the other hand. 2. Select a slide titled letter “e”. 3. Turn on the microscope and place the slide into the clip on the microscope stage. 4. Using the stage knobs, move the stage so the center of the slide (glass circle) is directly under the light. 5. Using the nosepiece, rotate the objectives and place the 4X objective over the slide. DO NOT rotate objectives by grabbing them directly. 6. Using the course adjustment knob, make sure the stage is all the way down. 7. Adjust both eye pieces so that your field of view merges into one field of view (one circle instead of two). 8. SLOWLY rotate the course adjustment knob to move up the stage, bringing the stage E closer to the objective. 9. Watch for the “e” to appear in your field of view. You will miss it if you bring the stage up too fast. 10. Center the “e” in your field of view and use the fine adjustment knob to focus it. 11. Switch objectives so you can view the “e” using the 10X objective. 12. Show your instructor the “e” under 10X. Have your instructor check off for completion in #3 on the answer worksheet. 13. Draw the “e” as accurately as possible in color, shape, and size, using the 10X objective in #4 on the answer worksheet. 3 BIO 101 Lab 03: Microscopy and Cells Activity 2: Familiarize yourself with single-celled bacteria, protist, and fungi 1. Take a prepped slide of ONE bacteria, ONE protist, and ONE fungi (only do one at a time) for observation. 2. Place each slide under the microscope and find your specimen using the scanning objective (4x). 3. Once you find the sample, then increase magnification to the 10X and then 40X objective. 4. Sketch your observations at 40X in your answer worksheet. 5. Be as detailed as possible in your drawings. 6. Follow the instructions in your worksheet for labeling and drawing the cells. 7. Return all slides to their proper tray. Activity 3: Prep Wet Mount of Plant cells for observations in dissecting and compound microscope 1. Take a single leaf from the Elodia plant and place it on a slide (Elodea are in beakers with water in the back of the room) 2. Place the slide (no coverslip) in the stage of the dissecting microscope. 3. Using the focus knob, properly focus on the leaf until it is in focus. 4. Use the zoom knob and zoom in (approx. 40x total mag) as much as possible. Re-focus if needed. 5. Sketch your observations in your answer worksheet at the highest zoom. Be as detailed as possible in your drawings and label any cellular structures that you observe in the individual plant cells. 6. After you are done, place a large drop of water over your Elodea leaf. 3. Carefully place a coverslip on the leaf. Make sure the sample is as flat as possible. 4. Using the light microscope, find your specimen as you did in activity 1 and 2. 5. Sketch your observations using the 40X objective in your answer worksheet. Be as detailed as possible in your drawings and label any cellular structures that you observe in the plant cells. 6. Throw away the coverslips and Elodea in the trash. Do not dispose of coverslips and plant leaves in the sink. Rinse off the slides and place them on the paper towels in the back of the room. Activity 4: Staining plant cells (onion leaves) 1. Very carefully cut a thin slice of onion. Thinner sections are easier to see under the microscope. 2. Place it on a slide and add 1 small drop of Methylene Blue onto the thin onion slice (be careful not to get the stain on you because it will stain your lab coat and hands) 3. Place a coverslip on the sample with stain and look at it under the microscope. 4. Methylene blue is a simple stain that is absorbed by the cell and can you help visualize cell structures. 5. Draw your cells using 40X objective on your answer worksheet. Label the different structures that are visible in the animal cells. 6. Throw away the coverslips and onion bits in the trash. Do not dispose of coverslips and onion in the sink. Rinse off the slides and place them on the paper towels in the back of the room. Activity 5: Wet Mount of pond water sample 1. Stir the pond water so that debris is suspended in the water, then use a pipette to transfer a drop of pond water onto a center of a clean slide. 2. Place a coverslip on the drop of water, avoiding making bubbles. 3. Using the 4x objective, systematically scan the slide in search of microorganisms. Remember, if it is green or it moves is a living organism! 4. Sketch at least 2 different microorganisms in your answer worksheet with as much detail as possible. 5. If one slide doesn’t yield visible specimen, make more slides. 6. If you need help finding specimen, call your instructor over to help you. 7. For each organism, try to identify it using the posters that are hanging on the dry-erase wall. 8. When you finish, throw away the cover slips but rinse off the slides and place them on the paper towels in the back of the room. 4 BIO 101 Lab 03: Microscopy and Cells Name: _________________________________________ Date: ____________ THE FOLLOWING PAGES SHOULD BE PRINTED OUT TO COMPLETE AND SUBMIT IN LAB. Data Analysis and Synthesis Questions: 1. Write the name and purpose of each structure of the microscope. (3 pts) 2. You found the letter “e” under using 10X and showed it to your professor. ________________ (1 pt) 4. Center the letter “e” under the objective and draw it using the 10x objective. Your drawings should be as close as possible to what you are seeing under the microscope (size, color, etc.). (2 pts) Total magnification: ___________ 5 BIO 101 Lab 03: Microscopy and Cells 5. Choose a bacterial slide from any of the following: “Bacteria Types Gram Stained”, “Bacteria Types w.m.”, Staphylococcus epidermidis, or Anabeana. Draw what you observe using 40X objective. (2 pts) • Use Table 1 in the introduction to properly label all cellular structures that are visible. • In the space below, write other structures that are found on bacteria. Name of slide: __________________________ 6. Choose a protist slide from any of the following: “Mixed protozoa”, “Dinolagellates”, “Flagellates from white ant” (Turtox No. P1.247), Volox (Turtox No. B1. 221), or Amoeba proteus. Some slides contain a variety of specimen. Draw only ONE of the single-celled protists you can see using the 10X or 40X objective. (2 pts) • Use Table 1 in the introduction to properly label all cellular structures that are visible. • In the space below, write other structures that are found in protists. Name of slide: ___________________________ 7. Choose a fungi slide from any on the tray labeled fungi. Draw ONE of the single-celled yeast you can see using the 40X objective. (2 pts) • Use Table 1 in the introduction to properly label all cellular structures that are visible. • In the space below, write other structures that are found in protists. Name of slide: ___________________________ 6 BIO 101 Lab 03: Microscopy and Cells 8. Draw the Elodea leaf (and cells if visible) at 40X total magnification using the dissecting microscope. Then draw the leaf cells at 400x total magnification using the light microscope (2 pts). • Use Table 1 from the introduction to properly label all cellular structures that are visible. Dissecting microscope (40x): Compound microscope (400x): 9. Draw your onion (plant) cell using 40x objective. (2 pts) • Use Table 1 from the introduction to properly label all cellular structures that are visible. • In the space below, write what structures the methylene blue stained for visualization in onion cells. 10.Draw two pond water organisms using the 10x objective or higher. Be as detailed as possible in your drawings and label any cellular structures that you observe. Try to identify the organisms using the posters on the board. (2 pts) Total magnification: Total magnification: ________ ________ ID: _________________________ ID: _________________________ 7 BIO 101 Lab 03: Microscopy and Cells 12. You observe cells under 400X total magnification. You see various organelles in the cytoplasm, including food vacuoles and cilia. What type of organism are you likely looking at? (0.5 pts) 13. You observe a very small cell at 1000X magnification. You can make out various external appendages, including a flagellum. What organism might you be looking at? (0.5 pts) 13. Name three organelles that are found in plant cells but are not found in animal cells. (1 pts) 14. Name three structures that are found in all living cells. (1 pts) Clean up and return microscopes to proper location • When you are done with all the experiments, clean the ocular and objective lenses using lens paper with lens cleaning solution. DO NOT use paper towels and water because you will ruin the objectives. • Wrap the cord in place on the microscope and return it to its proper number spot in the cabinet. • Be sure there is no plant or slide debris in sinks. 8
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