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    Universiti Sains Malaysia Jib 221 Microbiology Practical 2. Simple Stain and Gram Stain Techniques

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    The document describes a laboratory experiment involving simple staining and Gram staining techniques to observe the morphology of different microorganisms. Students are instructed to prepare smears of various bacteria and yeast samples, including E. coli, B. subtilis, S. aureus, and C. albicans. The smears will then be stained using simple staining with methylene blue or Gram staining techniques to distinguish Gram positive from Gram negative bacteria based on their ability to retain or decolorize the crystal violet dye. The objectives are to observe microscopic and macroscopic morphology of microorganisms using different staining methods.

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    Universiti Sains Malaysia Jib 221 Microbiology Practical 2. Simple Stain and Gram Stain Techniques

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    The document describes a laboratory experiment involving simple staining and Gram staining techniques to observe the morphology of different microorganisms. Students are instructed to prepare smears of various bacteria and yeast samples, including E. coli, B. subtilis, S. aureus, and C. albicans. The smears will then be stained using simple staining with methylene blue or Gram staining techniques to distinguish Gram positive from Gram negative bacteria based on their ability to retain or decolorize the crystal violet dye. The objectives are to observe microscopic and macroscopic morphology of microorganisms using different staining methods.

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    The document describes a laboratory experiment involving simple staining and Gram staining techniques to observe the morphology of different microorganisms. Students are instructed to prepare smears of various bacteria and yeast samples, including E. coli, B. subtilis, S. aureus, and C. albicans. The smears will then be stained using simple staining with methylene blue or Gram staining techniques to distinguish Gram positive from Gram negative bacteria based on their ability to retain or decolorize the crystal violet dye. The objectives are to observe microscopic and macroscopic morphology of microorganisms using different staining methods.

    Copyright:

    © All Rights Reserved
    Download as PDF, TXT or read online from Scribd
    Download as pdf or txt
    25 views3 pages

    Universiti Sains Malaysia Jib 221 Microbiology Practical 2. Simple Stain and Gram Stain Techniques

    Uploaded by

    Baby girl
    The document describes a laboratory experiment involving simple staining and Gram staining techniques to observe the morphology of different microorganisms. Students are instructed to prepare smears of various bacteria and yeast samples, including E. coli, B. subtilis, S. aureus, and C. albicans. The smears will then be stained using simple staining with methylene blue or Gram staining techniques to distinguish Gram positive from Gram negative bacteria based on their ability to retain or decolorize the crystal violet dye. The objectives are to observe microscopic and macroscopic morphology of microorganisms using different staining methods.

    Copyright:

    © All Rights Reserved
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    Download as pdf or txt
    of 3

    UNIVERSITI SAINS MALAYSIA

    JIB 221 MICROBIOLOGY

    PRACTICAL 2. SIMPLE STAIN AND GRAM STAIN TECHNIQUES

    Name: .............................. Matric number: .............................. Group:

    OBJECTIVES:
    • Prepare a series of smears of different microorganisms and observe using simple
    staining techniques
    • Prepare a series of smears of different microorganisms and observe using differential
    staining techniques (Gram Stain)
    • Observe the morphology of various microorganisms both macroscopic (based on
    colony shape and texture) and microscopic (shape, size, arrangement and coloration
    features)

    INTRODUCTION:

    One of the main characteristics of bacteria is their morphology, size, shape, arrangement and
    structure. This feature can be observed by staining the cells in a certain way
    and observing under the microscope.
    Simple staining: Microorganisms under natural conditions are quite difficult to see even
    when observed using oil immersion. Therefore, a simple dye is placed on the microorganisms
    to see their morphology more clearly.
    Diffrential staining: Differential staining are used to differentiate or highlight a variety cell
    morphology. Gram stain is an example of a differential staining technique.
    This staining technique is derived from the name of Christian Gram who discovered it in the
    year l884. It is the most commonly used stain in bacteriology and it distinguishes bacteria
    into two groups, Gram positive and Gram negative. Gram-positive organisms maintain the
    crystal violet dye even when they are decoloured by alcohol. Gram negative organisms are
    decoloured by alcohol and reacted with other dyes such as safranin. However, like other
    staining techniques, Gram staining results can change. To get reliable results, the cultures

    11
    used must be between 12-18 hours. Gram staining is a fundamental technique in
    microbiology and should be mastered by all students enrolled in this course.

    2a. SIMPLE STAIN

    MATERIALS:

    Microscope
    4 clean slides
    1 inoculation wire
    sterile distilled water
    blue methylene solution
    plate cultures:
    Escherichia coli
    Bacillus subtilis
    Staphylococcus aureus
    Candida albicans

    PROCEDURE:
    1. Using a sterile inoculating loop, add 1 drop of sterile water to the slide. Prepare a smear of
    bacteria. Air dry and heat-fixed. (Prepare a heat-fixed smear for each of the above
    cultures).
    2. Place the slide on the staining rack and flood the smear with the
    1% blue methylene solution and let it react for 1 minute.
    3. Wash the dye with tap water and dry the slide with absorbent paper.
    4. Keep it dry and observe under the 40X objective.
    Record your observations.

    12
    2b GRAM STAIN

    MATERIALS:

    crystal violet solution


    iodine solution
    ethanol
    safranin solution
    sterile distilled water
    wire loop
    4 clean glass slides
    plate culture (24 h):
    Escherichia coli
    Bacillus subtilis
    Staphylococcus aureus
    Pseudomonas aeruginosa

    PROCEDURE:

    1. Using a sterile inoculating loop, add 1 drop of sterile water to the slide. Prepare a smear of
    bacteria. Air dry and heat-fixed. (Prepare a heat-fixed smear for each of the above cultures)
    2. Place the slide on the colouring rack and flood the smear with the crystal violet solution
    and let it sit for 1 minute. Wash the dye slowly with tap water.
    3. Stain the smear with iodine. Let it sit for 1 minute and then rinse with tap water.
    4. Decolourise the smear using 95% alcohol by tapping the slide on the edge of the sink and
    dripping alcohol onto the slide until the colour disappears (approximately 30 seconds). Rinse
    with tap water immediately.
    5. Restain the smear with safranin for 1 minute. Wash the slides, then dry with absorbent
    paper. Observe the slides using 40X objective lens with maximum illumination to distinguish
    the colours. Record your observations.

    13

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