Cells and Microscopes

Transcription

1 Cells and Microscopes Introduction...2 Living and non-living things...2 Types of cells...2 The microscope...3 Types of microscopes...3 Parts of the microscope...3 TASK 1: IDENTIFYING THE DIFFERENT PARTS OF A MICROSCOPE...4 Total magnification power...4 Focusing Specimens...4 TASK 2: FOCUSING SPECIMENS...4 Drawing Specimens...5 Measuring microscopic structures...5 TASK 3: ESTIMATING THE SIZE OF THE FIELD OF VIEW...5 Prokaryotic Cells: Bacteria...6 TASK 4: PREPARED SLIDE OF BACTERIAL TYPES...6 Eukaryotic Cells: Animal Cells...6 TASK 5: WET MOUNT OF CHEEK CELLS...6 TASK 6: PREPARED SLIDE OF AMOEBA...7 Eukaryotic Cells: Plant Cells...7 TASK 7: WET MOUNT OF ELODEA CELLS...7 TASK 8: WET MOUNT OF ONION EPIDERMAL CELLS...7 TASK 9: WET MOUNT OF POND WATER...8 Animal and Plant Cellular Structures...9 TASK 10: ANIMAL AND PLANT CELLS ORGANELLES...9 General Biology 1 Instructor: Jose Bava, Ph.D

2 Introduction Living and non-living things If we are asked to define whether something is alive or not, the question seems nonsense, we all realize when we see something if that thing is a living or non-living thing. But if we are asked to define more precisely what attributes characterize living things, all of them without exception from a bacterium to an elephant and a redwood tree, what do we use? All living things have in common one main thing: they are able to self-maintain their body organization and grow to a given extent, and they are able to add more individuals to the population by means of reproduction (the act of producing offspring in whatever way they can). Viruses cannot do this and they use the resources of the host in order to replicate, a reason why they are not considered living things. The smallest units of life able to accomplish the previous are cells; some living things like bacteria are composed of only one cell, they are unicellular (uni=one), while a human being is composed of about 100 trillion cells (100,000,000,000,000) with hundred of different types that make organs and organs systems, all originating from the fertilized egg. In 1838, the cell theory was proposed, and this theory states that, Types of cells 1. All organisms are composed of one or more cells, and the life processes of metabolism and heredity occur within these cells. 2. Cells are the smallest living things, the basic units of organization of all organisms. 3. Cells arise only by division of a previously existing cell. Imagine bacteria, they are very small, have only one cell. This cell has a very simple design when compared to organisms like animals and plants. However, all living things need cells with the same basic components in order to survive, a generalized cell would look more or less like the one below. Bacterial cells lack a nucleus and the DNA is then free in the aqueous matrix of the cell, called cytosol or cytoplasm. This fact gives the name to this first type of cell in the living world: prokaryotic (pro=before, karyon=core). Bacteria and Archaea and the only two groups of living things having a prokaryotic cell, hence their name: prokaryotes. Prokaryotes have very small cells compared to other living things because they lack not only a nucleus but also most organelles (organ-like structures) present in other types of cells. Bacteria have a cell wall that surrounds the cell membrane giving strength and protection to the cell and some other things particular of them, but for the scope of this lab we do not need to know more than this about these simple organisms. All other organisms of on earth have cells with a nucleus, hence their name: eukaryotes (eu=true, karyon=core). Eukaryotes also have many organ-like structures called organelles in the cells responsible for different activities inside the cells. Now, consider a human being; we have different cell designs for the heart, the stomach, the brain, and all other organs and parts of our body. However, the general structure of all these cells is the same; they have the same internal organization even if the way they look or work in the body is different. This general type of cell is named animal cell and is present, as the name indicates, in animals but also in a type of animal-like unicellular organisms called protozoans. Plants, on the other hand, have a different cell design; plant cells have thick, rigid General Biology 2 Instructor: Jose Bava, Ph.D

3 walls (different from those of bacteria and fungi) that consist of a material called cellulose. Animal cells do not have this material. The cellulose enables plants to stand upright without the aid of an internal or external skeleton. Also, plants are able to make photosynthesis and the chlorophyll responsible for trapping the sunlight resides in a specific organelle called chloroplast. During this lab, we will observe these two main differences between animal and plant cells, and we will be able to infer the third one: plant cells have a compartment called central vacuole, a large, water-filled organelle in plant cells that stores toxins, degrades enzymes, disposes of wastes, and is involved in cell growth. The microscope As the name indicates, microscopes (micron = small and scopos = aim) is an instrument for viewing objects that are too small to be seen by the naked or unaided eye, these instruments first appeared around the sixteenth century. Plant and animal cells have different characteristic sizes, but they are always really big when compared to bacteria, the microscope we will use in today s lab is called light or compound microscope and allows observing eukaryotic (plant, animal) cells but not prokaryotic cells (bacteria). Check the picture below in order to compare the relative size of the different types of cells. Types of microscopes The name light microscope refers to the use of compound (two) lenses and light to magnify objects. The lenses bend or refract the light, which makes the object beneath them appear closer. The light has to go through the object in order to magnify it, so whatever you are observing has to be very, very thin! You may notice in the picture above that we may be able to observe a bacterium with the compound microscope, but we cannot see viruses; in order to do this we need to use electron microscopes (EM). These microscopes work in a different way: instead of using light waves; they use electrons (negatively charged electrical particles) to magnify objects up to two million times. Parts of the microscope 1. Ocular lens: Can have one or two pieces and the magnification power is normally 10X 2. Objective lenses: The lens in a microscope closest to the specimen, can have up to four lenses with, normally, 4X, 10X, 40X, and 100X. 3. Stage: The platform on the microscope where the specimen is mounted for examination 4. Mechanical stage: The mechanical stage is mounted on the microscope stage 5. Mechanical stage adjustment knobs: Two knobs allows precise movement and positioning of the slide in the X and Y direction 6. Brightness control knob, ON/OFF: Controls luminosity of the microscope and also has the on/off switch 7. Lamp: Located at the base of the microscope, provides the light to illuminate the specimen 8. Iris diaphragm: Controls the size of the lens opening and therefore the amount of light that reaches the specimen 9. Condenser lens: A lens system which concentrates the light from coming from the lamp into a beam 10. Arm: Always carry the microscope by holding it from the arm and the base 11. Coarse adjustment knob: Is the first device for focusing. With the scanner objective (4X) in place, this knob is used to determine a sharp focus 12. Fine adjustment knob: Allows fine adjustment for a sharp image General Biology 3 Instructor: Jose Bava, Ph.D

4 TASK 1: IDENTIFYING THE DIFFERENT PARTS OF A MICROSCOPE Total magnification power In this lab we will use only three magnifications: Scanning (4x), Low (10x) and High (40x). Each objective will have written the magnification. In addition to this, the ocular lens (eyepiece) has a magnification. The total magnification is the ocular x objective (i.e. 10 X 40 = 400X). Focusing Specimens 1. Always start with the scanning objective (4x). Odds are, you will be able to see something on this setting. Use the Coarse Knob to focus, image may be small at this magnification, but you won't be able to find it on the higher powers without this first step. Do not use stage clips, try moving the slide around until you find something. 2. Once you've focused on Scanning, switch to Low Power (10x). Use the Coarse Knob to refocus. Again, if you haven't focused on this level, you will not be able to move to the next level. 3. Now switch to High Power (40x). (If you have a thick slide, or a slide without a cover, do NOT use the high power objective). At this point, ONLY use the Fine Adjustment Knob to focus specimens. TASK 2: FOCUSING SPECIMENS Procedure: Obtain a slide of a letter e and place it on the microscope stage in the orientation in which you normally read the letter. Observe it with the scanning objective (4x). 1) How does the orientation change when you observe the letter under the microscope?... 2) While observing the letter through the microscope, move the letter to the right, the letter appears to move to the?... General Biology 4 Instructor: Jose Bava, Ph.D

5 Procedure: Obtain a slide of crossed fibers (threads) and observe them with the low power objective (10x). 3) Which thread appears to be on top?. 4) Which thread appears to be at the bottom?... 5) Can you observe a sharp image of the three fibers at the same time where they cross over? YES / NO Drawing Specimens All drawings should include clear and proper labels of whatever you can distinguish in the specimen (i.e. cell membrane, cell wall, nucleus, etc.). Drawings should be labeled with the specimen name and magnification used. You do not have to draw the entire field of view of cells; if that is happening, then you should move to a better magnification power and sketch just one cell with more details, like in the picture on the side. Measuring microscopic structures To estimate the size of a specimen when viewed under the microscope, an instrument called ocular micrometer is used. This device is a glass disk that fits in a microscope eyepiece that has a ruled scale, which is used to measure the size of magnified objects. The physical lengths of the marks on the scale depend on the degree of magnification. An indirect way to estimate the size of microscopic structures is knowing in advance the diameter of the field of view for a given magnification power (objective lens). TASK 3: ESTIMATING THE SIZE OF THE FIELD OF VIEW Procedure: A. Place the 4x objective lens in position (total magnification 40X) B. Slide a transparent ruler onto the stage while observing it under the microscope C. Align the ruler in such a way that a line corresponding a millimeter touches the left side of the field of view (see picture on the side) D. Count the number of millimeters you see, that is the size (width) of your field of view 6) Record in the table below the size of the field of view for the 40x magnification power in millimeters. Multiply the number in mm *1000 and obtain the size in micrometers (µm), the unit really used to estimate the size of microscopic objects like cells FOV 40X Field of View (millimeters, mm). Field of View (micrometers, µm).. E. Repeat the procedure for the 10x objective lens (total magnification 100X) 7) Record in the table below the size of the field of view for the 100x magnification power in millimeters. Multiply the number in mm *1000 and obtain the size in micrometers (µm) FOV 100X Field of View (millimeters, mm). Field of View (micrometers, µm).. F. We cannot repeat the procedure for the 40x objective lens because the size of the field of view (FOV) is smaller than one millimeter, but we can apply the following formula to calculate its FOV if we know the size of the field of view for one of the other two objective lenses. 8) Apply the following formula and record in the table below the size of the field of view for the 400x magnification power in millimeters. Multiply the number in mm *1000 and obtain the size in micrometers (µm) General Biology 5 Instructor: Jose Bava, Ph.D

6 Field of View 400x = (40 X FOV 40X ) / 400 FOV 400X Field of View (millimeters, mm). Field of View (micrometers, µm).. 9) Analyze the previous results, what is the relationship between the magnification power used and the size of the field of view (FOV) or area observed under the microscope? Prokaryotic Cells: Bacteria TASK 4: PREPARED SLIDE OF BACTERIAL TYPES Bacterial cells lack a nucleus and the DNA is then free in the aqueous matrix of the cell, called cytoplasm. This fact gives the name to this first type of cell in the living world: prokaryotic (pro=before, karyon=core). Bacteria and Archaea and the only two groups of living things having a prokaryotic cell, hence their name: prokaryotes. 10) Observe a prepared slide of different bacterial types and notice the small size prokaryotic cells have 11) Why are bacterial cells (prokaryotic) so small? Explain (see introduction) Eukaryotic Cells: Animal Cells TASK 5: WET MOUNT OF CHEEK CELLS Procedure: 1. Gently scrape the inside of your cheek with a clean toothpick 2. Make the tip of the toothpick touch the center of a clear microslide to leave a sample of cells there 3. Add a drop of stain (iodine or methylene blue) 4. Top with a cover slip (that protects the microscope from the sample while prevents the sample from drying out) 12) Observe the cheek cells and LABEL what you see in the space provided below Cheek cell ( X) Label: Cell membrane, cytoplasm, nucleus Amoeba cell ( X) Label: Cell membrane, cytoplasm, nucleus, pseudopods General Biology 6 Instructor: Jose Bava, Ph.D

7 TASK 6: PREPARED SLIDE OF AMOEBA Amoebas are unicellular animal-like protists or protozoans, they move around extending the cell membrane and the cytoplasm in one direction. These projections are called pseudopods (pseudo=false, pod=leg). The amoeba you will observe belongs to a group of free-living organisms, but there are others that live as parasites, infecting all types of vertebrates and some invertebrates. One type of amoeba, Entamoeba histolytica, is spread by means of contaminated water or food and causes amebic dysentery. 100,000 people die worldwide every year, and this disease is the third most important disease caused by parasites after malaria (caused by another protozoan) and schistosomiasis (caused by a fluke, a type of flatworm) 13) Obtain a prepared slide of amoeba, sketch and LABEL what you see in the space provided in the previous page 14) Which type of cell is bigger? The prokaryotic (bacteria) or the eukaryotic (amoeba, cheek cell)? Eukaryotic Cells: Plant Cells TASK 7: WET MOUNT OF ELODEA CELLS Procedure: 1. Place a drop of water on the center of a clean dry slide 2. Using the tweezers, place a leaf of elodea in the middle of the drop. 3. Top with a cover slip (DO NOT STAIN!) 15) Observe the elodea cells under the microscope, sketch one cell in the space provided below with the best magnification you can and label what you see. Elodea cell ( X) Label: Cell wall + cell membrane, cytoplasm, chloroplasts Onion cell ( X) Label: Cell wall + cell membrane, cytoplasm, nucleus TASK 8: WET MOUNT OF ONION EPIDERMAL CELLS Procedure: 1. Place a drop of water on the center of a clean dry slide 2. Using the tweezers, peel the thin layer of sin of an onion 3. Add a small drop of stain if (iodine or methylene blue) 4. Top with a cover slip General Biology 7 Instructor: Jose Bava, Ph.D

8 16) Observe the onion cells under the microscope, sketch one cell in the space provided above with the best magnification you can and label what you see 17) Onion cells are plant cells but they lack chloroplasts, so no photosynthesis happens there, why is that? 18) Consider the size of both cheek and elodea cells, what type of cell was bigger? (ANIMAL= cheek cell / PLANT= elodea cell) 19) What cellular structures you observed differentiate both types of cells (animal versus plant)? TASK 9: WET MOUNT OF POND WATER Procedure: 1. Make a wet mount of pond water (DO NOT STAIN) 2. Observe under the microscope, you should find several unicellular creatures swimming in the water. 3. Sketch and label two different organisms you can identify in the sample. Ask your lab instructor if you are not sure about what you see!.. cell ( X) Label whatever you can differentiate.. cell ( X) Label whatever you can differentiate General Biology 8 Instructor: Jose Bava, Ph.D

9 Animal and Plant Cellular Structures 20) Complete the following information regarding the cellular structures observed under the microscope ONLY present in plants cells Present in BOTH cellular types... TASK 10: ANIMAL AND PLANT CELLS ORGANELLES 21) The following figure shows a generalized view of a cell that combines both plant and animal characteristics. Name the cellular structures present and specify if these are present in plant cells, in animal cells, or in both cellular types. Cell or Plasma membrane: ALL living things have cells where the cell membrane makes the outer boundary of the cytoplasm, a bi layer of phospholipids that controls de movement of materials in and out of the cell General Biology 9 Instructor: Jose Bava, Ph.D

10 Cell wall: a rigid layer of polysaccharides enclosing the membrane of plant, algae, fungi and prokaryotic (bacteria) cells; maintains the shape of the cell and serves as a protective barrier Centrioles: composed of proteins, are involved in the process of cell division, centrioles are exclusive of animal cells (not present in plant cells) Chloroplasts: (phospholipids) membrane organelles that are the site for photosynthesis: a process in plants, algae and some bacteria that uses sunlight to produce glucose and, as a waste product, molecular oxygen Cytoplasm: the watery solution that bathes and suspends all the organelles Cytoskeleton: The cytoskeleton is a cellular "scaffolding" or "skeleton" contained within the cytoplasm. The cytoskeleton is present in all cells; it was once thought this structure was unique to eukaryotes, but recent research has identified the prokaryotic cytoskeleton. It is a dynamic structure that maintains cell shape, protects the cell, enables cellular motion and plays important roles in both intracellular transport and cellular division Golgi apparatus: Stack of (phospholipids) membranes where substances are further modified, stored, and packaged in vesicles for transport in and out of the cell. The golgi is also the site where lysosomes are produced in animal cells Lysosomes: Are (phospholipids) membrane vesicles containing digestive enzymes that break down molecules, normally present only in animal cells Mitochondria: (phospholipids) membrane organelles involved in aerobic cellular respiration, a process that burns glucose using oxygen (O 2 ) and produces ATP, the energy currency for metabolic processes Nuclear membrane: Made also of phospholipids. Separates the inside of the nucleus, the DNA, from the rest of the cell. It has pores that allow things to move in and out of the nucleus Nucleus: Is the information storage place of the cell, a large spherical organelle that holds the genetic material (DNA) safe inside the cell Nucleolus: Present inside the nucleus, the nucleolus is the place where ribosomes are produced Ribosomes: very small organelles composed of RNA and proteins, are the site or factory for protein synthesis. Ribosomes may be free in the cytoplasm, where they are mostly involved in the synthesis of proteins that will be used in the cell, or attached to the Rough Endoplasmatic Reticulum (RER), where they are used to produce for export proteins, proteins that will be used in other cells or will fulfill their function in the blood stream Rough endoplasmic reticulum (RER): consists of folded (phospholipids) membranes confluent with the nuclear membrane. Its surface is studded with ribosomes, is involved in synthesis and modification of proteins. Smooth endoplasmic reticulum (SER): also consists of folded (phospholipids) membranes, and is so named because it lacks attached. It extend outward the RER and is connected with synthesis of non-protein molecules such as lipids. Vacuoles: (phospholipids) membrane vesicles used to store substances. Plants cells have a very big vacuole, called central vacuole, which is not present in animal cells. In both plant and animal cells vacuoles are used to remove waste products and store ingested food 22) Which cellular structure or organelles are made of proteins? 23) Which cellular structure or organelles are made of phospholipids membranes?... General Biology 10 Instructor: Jose Bava, Ph.D