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Questions and Answers
What is the primary purpose of gram staining in a laboratory setting?
What is the primary purpose of gram staining in a laboratory setting?
- To differentiate between types of bacteria (correct)
- To count the number of bacteria present
- To determine the size of bacterial cells
- To stain all bacteria uniformly
Which item acts as a mordant in the gram staining procedure?
Which item acts as a mordant in the gram staining procedure?
- Safranin
- 95% Ethyl alcohol
- Iodine (correct)
- Crystal violet
What is the purpose of using 95% Ethyl alcohol in gram staining?
What is the purpose of using 95% Ethyl alcohol in gram staining?
- To act as a primary stain
- To enhance cell visibility
- To fix the smear on the slide
- To decolorize gram-negative bacteria (correct)
If a slide shows gram-positive bacteria appearing pink under the microscope, what does it indicate?
If a slide shows gram-positive bacteria appearing pink under the microscope, what does it indicate?
Why is it crucial to wash the slide with distilled water immediately after adding 95% Ethyl alcohol?
Why is it crucial to wash the slide with distilled water immediately after adding 95% Ethyl alcohol?
What color would a gram-negative bacterium appear after complete gram staining?
What color would a gram-negative bacterium appear after complete gram staining?
What is indicated by the absence of bacteria on a stained slide observed under a microscope?
What is indicated by the absence of bacteria on a stained slide observed under a microscope?
What role does the counterstain play in the gram-staining process?
What role does the counterstain play in the gram-staining process?
What happens to gram-positive bacteria during the decolorization process?
What happens to gram-positive bacteria during the decolorization process?
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Study Notes
Gram Staining Overview
- Gram staining is a technique used to differentiate between bacteria based on their cell wall structure.
- The technique involves staining bacteria with a specific dye and then treating them with a decolorizing agent and a counterstain.
Gram Staining Procedure
- Primary Stain: Crystal violet dye is applied to the bacterial smear, staining all bacteria initially purple.
- Mordant: Gram's iodine solution is added to form a complex with the crystal violet, enhancing its retention by the cell wall.
- Decolorizing Agent: 95% Ethyl alcohol is used to remove the crystal violet-iodine complex from the cell wall. Gram-positive bacteria retain the stain due to their thicker peptidoglycan layer.
- Counterstain: Safranin dye is applied to stain the decolorized bacteria pink. This allows for visualization of gram-negative bacteria which have lost the primary stain.
Gram Staining Results
- Gram-positive bacteria retain the primary stain (crystal violet) and appear purple.
- Gram-negative bacteria lose the primary stain and are stained by the counterstain (safranin) and appear pink.
Gram Staining Issues
- Incorrect application of reagents: Failure to properly apply reagents can result in inaccurate staining results.
- Over-decolorization: Using the decolorizing agent for too long will remove the primary stain from both gram-positive and gram-negative bacteria.
- Under-decolorization: Insufficient decolorization may cause gram-negative bacteria to retain the crystal violet resulting in a false-positive result.
- Aging bacterial cultures: Older bacterial cultures may lose their cell wall integrity and become more susceptible to decolorization, potentially leading to inaccurate results.
- Overheating the sample: Excessive heating may cause cell wall damage and lead to inaccurate results.
Gram Staining Significance
- Gram staining is a commonly used diagnostic tool in microbiology.
- It helps identify bacteria, providing crucial information for their classification and the selection of appropriate antibiotic treatment.
- It helps to understand the differences in cell wall structure between bacterial types.
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