Chapter 3 Observing Microorganisms Through a Microscope 2013
Chapter 3 Observing Microorganisms Through a Microscope © 2013 Pearson Education, Inc. Copyright © 2013 Pearson Education, Inc. Lectures prepared by Christine L. Case Lectures prepared by Christine L.
Figure 3. 2 Microscopes and Magnification. Unaided eye ≥ 200 m Light microscope 200 nm – 10 mm Tick Actual size Scanning electron microscope 10 nm – 1 mm Red blood cells Transmission electron microscope 10 pm – 100 m E. coli bacteria T-even bacteriophages (viruses) Atomic force microscope 0. 1 nm – 10 nm DNA double helix © 2013 Pearson Education, Inc.
Preparing Smears for Staining § Live or unstained cells have little contrast with the surrounding medium. Researchers do make discoveries about cell behavior by observing live specimens. ANIMATION Microscopy and Staining: Overview © 2013 Pearson Education, Inc.
Preparing Smears for Staining § Staining: coloring the microbe with a dye that emphasizes certain structures § Smear: a thin film of a solution of microbes on a slide § A smear is usually fixed to attach the microbes to the slide and to kill the microbes © 2013 Pearson Education, Inc.
Simple Stains § Simple stain: use of a single basic dye § A mordant may be used to hold the stain or coat the specimen to enlarge it ANIMATION Staining © 2013 Pearson Education, Inc.
Differential Stains § Used to distinguish between bacteria § Gram stain § Acid-fast stain © 2013 Pearson Education, Inc.
Gram Stain § Classifies bacteria into gram-positive or gram-negative § Gram-positive bacteria tend to be killed by penicillin and detergents § Gram-negative bacteria are more resistant to antibiotics © 2013 Pearson Education, Inc.
Gram Stain Color of Gram-Positive Cells Color of Gram-Negative Cells Primary Stain: Crystal Violet Purple Mordant: Iodine Purple Decolorizing Agent: Alcohol-Acetone Purple Colorless Counterstain: Safranin Purple Red © 2013 Pearson Education, Inc.
Figure 3. 12 Gram staining. Gram-positive Gram-negative Application of crystal violet (purple dye) Application of iodine (mordant) Rod (gram-negative) Cocci (gram-positive) © 2013 Pearson Education, Inc. Alcohol wash (decolorization) Application of safranin (counterstain)
Acid-Fast Stain § Stained waxy cell wall is not decolorized by acid-alcohol § Mycobacterium § Nocardia © 2013 Pearson Education, Inc.
Acid-Fast Stain Color of Acid-Fast Color of Non–Acid-Fast Primary Stain: Carbolfuchsin Red Decolorizing Agent: Acid-alcohol Red Colorless Counterstain: Methylene Blue Red Blue © 2013 Pearson Education, Inc.
Figure 3. 13 Acid-fast bacteria. M. bovis © 2013 Pearson Education, Inc.
Special Stains § Used to distinguish parts of cells § Capsule stain § Endospore stain § Flagella stain © 2013 Pearson Education, Inc.
Negative Staining for Capsules § Cells stained § Capsule is not stained--negative stain © 2013 Pearson Education, Inc.
Figure 3. 14 a Special staining. Capsules Negative staining © 2013 Pearson Education, Inc.
Figure 3. 14 b Special staining. Endospore staining © 2013 Pearson Education, Inc.
Figure 3. 14 c Special staining. Flagellum Flagella staining © 2013 Pearson Education, Inc.
Table 3. 3 A Summary of Various Stains and Their Uses © 2013 Pearson Education, Inc.
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