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Microbiology Lab Report - Experiment 2

Gram staining is a differential staining technique used to distinguish between gram-positive and gram-negative bacteria based on differences in their cell walls. Gram-positive cells have a thick peptidoglycan layer that retains the primary stain, while gram-negative cells have a thinner peptidoglycan layer surrounded by an outer membrane, causing them to take up the counterstain instead. Acid-fast staining uses the Ziehl-Neelsen technique to identify acid-fast organisms like mycobacteria by their waxy, impermeable cell walls resistant to decolorization. Endospore staining detects and identifies the presence of bacterial endospores, which are dormant, resistant structures produced by certain Firmic

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580 views2 pages

Microbiology Lab Report - Experiment 2

Gram staining is a differential staining technique used to distinguish between gram-positive and gram-negative bacteria based on differences in their cell walls. Gram-positive cells have a thick peptidoglycan layer that retains the primary stain, while gram-negative cells have a thinner peptidoglycan layer surrounded by an outer membrane, causing them to take up the counterstain instead. Acid-fast staining uses the Ziehl-Neelsen technique to identify acid-fast organisms like mycobacteria by their waxy, impermeable cell walls resistant to decolorization. Endospore staining detects and identifies the presence of bacterial endospores, which are dormant, resistant structures produced by certain Firmic

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Nida Ridzuan
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INTRODUCTION:

Gram staining is the most important stain in routine bacteriology. It is differential stain used for the
identification of bacteria. This is used to distinguish between gram-positive and gram-negative
bacteria which have consistent differences in their cell walls. Gram-positive cells have a thick
peptidoglycan layer, whereas the peptidoglycan layer in gram-negative cells is much thinner and
surrounded by outer lipid containing layers. Gram stain is a type of differential staining which
requires the use of at least three chemical reagent, crystal violet for primary stain, gram’s iodine as a
mordant, and 95% ethyl alcohol used as a decolorizing agent, that is applied sequentially to a heat
fixated smear. After the staining procedure, gram-positive cells turn into purple from the crystal
violet, while the gram-negative turns pink or red from the safranin.

Acid fast staining is the differential staining techniques which was first developed by Ziehl and then
modified by Neelsen. So, this method is also called Ziehl-Neelsen technique. Acid-fast stain is a
differential stain used to identify acid-fast organisms such as members of the genus Mycobacterium.
Acid-fast organisms are characterized by wax-like, nearly impermeable cell walls. They contain
mycolic acid and large amounts of fatty acids, waxes, and complex lipids. This type of cell wall is
resistant to most compounds. So, acid-fast organisms require a special staining technique.

An endospore is a non-vegetative structure produced by a group of bacteria belonging to the Firmicute


family. They have special characteristics that stabilize them to survive in adverse conditions for long
periods of time. This staining technique is known as the endospore stain, also known as the spore
stain. It is used majorly to detect and identify the presence of a bacterial endospore and vegetative
forms in a cell. Examples of these endospore-forming bacteria include Clostridium spp and Bacillus
spp. Spores are resistant to heat, chemicals, and radiation. Bacteria can form endospores in 6 to 8
hours after being exposed to adverse conditions. The normal growing cell that forms the endospore is
called a vegetative cell. Endospores can form within different areas of the vegetative cell. They can be
central, subterminal, or terminal. Central endospores are located within the middle of the vegetative
cell. Terminal endospores are located at the end of the vegetative cell. Subterminal endospores are
located between the middle and the end of the cell. Endospore staining techniques are classified based
on the types of reagents used which are Schaeffer Fulton Stain that used Malachite Green dye and
safranin and Dorner method of endospore staining that uses Carbolfuchsin stain, acid alcohol, and
Nigrosin solution.
OBJECTIVES:

1. To differentiate between the two major categories of bacteria: Gram positive and Gram
negative.
2. To understand how the Gram stain reaction affects Gram-positive and Gram-negative bacteria
based on the biochemical and structural differences of their cell walls.
3. To understanding the principle of Ziehl-Neelson stain.
4. To differentiate bacteria into acid-fast group and non-acid fast groups.
5. To properly perform an endospore stain.
6. To detect for the presence of an endospore.

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