Questions and Answers
What is the main purpose of phase contrast microscopy?
Phase contrast microscopy does not use fixed smears.
True
What does fluorescent microscopy utilize for illumination?
Fluorophores or fluorochromes
What is total magnification?
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The resolving power is the shortest distance between two points that can be determined to be separate _____ .
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Which method is used to enhance contrast in microscopic specimens?
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Bright-field microscopy uses visible light that appears dark against a brighter background.
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What is the use of oil immersion in microscopy?
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In dark-field microscopy, how does the light reach the specimen?
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Study Notes
Phase Contrast Microscopy
- Most widely used method for detecting microorganisms directly in clinical specimens.
- Characterizes microorganisms grown in culture.
- Helps determine the presence of infections and identify organisms not visible to the naked eye.
- Involves direct observation of unstained specimens through wet mounts, enhancing visualization without using fixed smears.
- Screens specimen adequacy utilizing Bartlett's classification.
Fluorescent Microscopy
- Utilizes fluorophores that absorb UV light and emit visible light when returning to a lower energy state.
- Involves direct chemical interaction with components of bacterial cells via fluorochromes (e.g., Acridine orange, Auramine, Calcofluor white).
- Immunofluorescence uses antibodies conjugated to fluorescent dyes (e.g., Fluorescein isothiocyanate, FITC).
- Highlights fluorescing objects against a dark background for improved visibility.
- Utilizes excitation/barrier filters to protect the observer’s eyes and select transmitted light wavelengths.
- Digital photography frequently employed for permanent record-keeping.
Principal Factors Influencing Microscopic Image Quality
- Magnification: Size of the magnified image; total magnification is calculated by multiplying eyepiece magnification by objective magnification (e.g., 10x eyepiece and 100x oil immersion equals 1000x total magnification).
- Resolution: Ability to maintain detail in a magnified object; resolving power defines the shortest distance between distinguishable points, reliant on the numerical aperture of the objective lens.
- Contrast: Ability to visualize specimen details distinctly; can be enhanced by staining or adjusting the diaphragm.
Types of Microscopy
- Bright-Field Microscopy: Visible light passes through specimens; images appear dark against a brighter background. Magnification can reach up to 1000x, enabling visualization of fungi, parasites, and bacteria, often requiring oil immersion for clarity.
- Dark-Field Microscopy: Alters the microscopic technique to achieve contrast without dyes or stains; light is directed at an oblique angle, causing specimens to appear luminous against a dark background.
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Description
Test your knowledge on phase contrast microscopy, a vital technique in microbiology for examining specimens. This quiz covers key concepts related to microscopy, specimen staining, and direct examination. Enhance your understanding of how light interacts with different densities in microorganisms.
