Review_CH2

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Chemistry

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Feb 20, 2024

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Review CH2 1) Fill in the following blanks. a) 1 μm = 10 26 m b) 1 nm = 10 -9 m c) 1 μm = 10 3 nm 2) Which type of microscope would be best to use to observe each of the following? a) a stained bacterial smear i) compound light microscope b) unstained bacterial cells: the cells are small, and no detail is needed. i) Darkfield microscope c) unstained live tissue when it is desirable to see some intracellular detail. i) Phase-contrast microscope d) a sample that emits light when illuminated with ultraviolet light. i) Fluorescence microscope e) intracellular detail of a cell that is 1 μm long. i) Electron microscope f) unstained live cells in which intracellular structures are shown in color. i) Differential interference contrast microscope 3) DRAW IT Label the parts of the compound light microscope in the figure below, and then draw the path of light from the illuminator to your eye.

4. Calculate the total magnification of the nucleus of a cell being observed through a compound light microscope with a 10× ocular lens and an oil immersion lens. 5. The maximum magnification of a compound microscope is (a) 1500x ; that of an electron microscope, (b) 10,000,000x . The maximum resolution of a compound microscope is (c) 0.2μm ; that of an electron microscope, (d) 10 pm . One advantage of a scanning electron microscope over a transmission electron microscope is (e) Seeing three-dimensional detail . 6. Why is a mordant used in the Gram stain? In the flagella stain?  In a Gram stain, the mordant combines with the basic dye to form a complex that will not wash out of gram-positive cells. In a flagella stain, the mordant accumulates on the flagella so that they can be seen with a light microscope. 7. What is the purpose of a counterstain in the acid-fast stain?  A counterstain stains the colorless non–acid-fast cells so that they are easily seen through a microscope. 8. What is the purpose of a decolorizer in the Gram stain? In the acid-fast stain?

 In the Gram stain, the decolorizer removes the color from gram-negative cells. In the acid-fast stain, the decolorizer removes the color from non–acid-fast cells. 9. Fill in the following table regarding the Gram stain: 10. DRAW IT A sputum sample from Calle, a 30-year-old Asian elephant, was smeared onto a slide and air dried. The smear was fixed, covered with carbolfuchsin, and heated for 5 minutes. After washing with water, acid-alcohol was placed on the smear for 30 seconds. Finally, the smear was stained with methylene blue for 30 seconds, washed with water, and dried. On examination at 1000×, the zoo veterinarian saw red rods on the slide. What microbe do these results suggest? Multiple Choice 1) Assume you stain Bacillus by applying malachite green with heat and then counterstain with safranin. Through the microscope, the green structures are. a) cell walls. b) capsules. c) endospores. d) flagella . e) impossible to identify . 2) Three-dimensional images of live cells can be produced with a) darkfield microscopy. b) fluorescence microscopy.

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c) transmission electron microscopy. d) confocal microscopy. e) phase-contrast microscopy. 3) Carbolfuchsin can be used as a simple stain and a negative stain. As a simple stain, the pH is a) 2. b) higher than the negative stain. c) lower than the negative stain. d) the same as the negative stain. 4. Looking at the cell of a photosynthetic microorganism, you observe the chloroplasts are green in brightfield microscopy and red in fluorescence microscopy. You conclude: a. chlorophyll is fluorescent. b. The magnification has distorted the image. c. you’re not looking at the same structure in both microscopes. d. The stain masked the green color. e. none of the above 5. Which of the following is not a functionally analogous pair of stains? a. nigrosin and malachite green b. crystal violet and carbolfuchsin c. safranin and methylene blue d. ethanol-acetone and acid-alcohol e. All of the above pairs are functionally analogous. 6. Which of the following pairs is mismatched ? a. capsule—negative stain b. cell arrangement—simple stain c. cell size—negative stain d. Gram stain—bacterial identification e. none of the above 7. Assume you stain Clostridium by applying a basic stain, carbolfuchsin, with heat, decolorizing with acid-alcohol, and counterstaining with an acidic stain, nigrosin. Through the microscope, the endospores are 1 , and the cells are stained 2 . a. 1—red; 2—black b. 1—black; 2—colorless c. 1—colorless; 2—black d. 1—red; 2—colorless e. 1—black; 2—red 8. Assume that you are viewing a Gram-stained field of red cocci and blue rods through the microscope. You can safely conclude that you have. a. made a mistake in staining. b. two different species. c. old bacterial cells. d. young bacterial cells. e. None of the above

9. In 1996, scientists described a new tapeworm parasite that had killed at least one person. The initial examination of the patient’s abdominal mass was most likely made using. a. brightfield microscopy. b. Darkfield microscopy. c. electron microscopy. d. phase-contrast microscopy. e. fluorescence microscopy. 10. Which of the following is not a modification of a compound light microscope? a. brightfield microscopy b. darkfield microscopy c. electron microscopy d. phase-contrast microscopy e. fluorescence microscopy Analysis 1. In a Gram stain, a step could be omitted and still allow differentiation between gram- positive and gram-negative cells. What is it?  Step 1 can be omitted. Safranin can be omitted. when it is omitted, gram-negative cells will be unstained instead of pink and gram-positive cells will be purple in color. 2. Using a good compound light microscope with a resolving power of 0.3 μm, a 10× ocular lens, and a 100× oil immersion lens, would you be able to discern two objects separated by 3 μm? 0.3 μm? 300 nm?  All of them can discern objects. 3. Why isn’t the Gram stain used on acid-fast bacteria? If you did Gram stain acid-fast bacteria, what would their Gram reaction be? What is the Gram reaction of non–acid-fast bacteria?  The high lipid content of acid-fast bacteria makes it impermeable to most stains, so the gram stain decolorizer won't do anything to the cell.  1. these bacteria have mycolic acid in their cell walls which prevents the stain from entering the cells

2. the bacteria appear colorless because the cell walls are resistant to Gram stain 3. there would be a typical reaction, either gram-positive or gram-negative 4. Endospores can be seen as refractile structures in unstained cells and as colorless areas in Gram-stained cells. Why is it necessary to do an endospore stain to verify the presence of endospores?  because they can't be differentiated from inclusion of stored material without a special stain endospores can't be stained by ordinary methods (such as gram staining) because the dyes don't penetrate the wall of the endospore Clinical Applications and Evaluation 1. In 1882, German bacteriologist Paul Erhlich described a method for staining Mycobacterium and noted, “It may be that all disinfecting agents which are acidic will be without effect on this [tubercle] bacillus, and one will have to be limited to alkaline agents.” How did he reach this conclusion without testing disinfectants?  Mycobacterium being an acidophilic bacterium, it seems logical that it will be more resistant to an acidic chemical, therefor the use of alkaline agents is advised. 2. Laboratory diagnosis of Neisseria gonorrhoeae infection is based on microscopic examination of Gram-stained pus. Locate the bacteria in this light micrograph. What is the disease?

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 Gonorrhea caused by Gram-negative cocci. 3. Assume that you are viewing a Gram-stained sample of vaginal discharge. Large (10 μm) nucleated red cells are coated with small (0.5 μm wide by 1.5 μm long) blue cells on their surfaces. What is the most likely explanation for the red and blue cells?  Red cells = damaged yeast cells. Blue cells = lactic acid bacteria.  The red cells are the erythrocytes or the RBCs. Whereas, the blue cells are the Gram- positive bacteria, they are appearing as blue because they have given positive test with the Gram staining process.