Merhan, Kristine Mae M. (Lab Activity 5)

NAME Kristine Mae M.
Merhan DATE April 19, 2022

Year/Section/Set: BSN-1B (Set B)
Activity No. 5
STAINING OF MICROORGANISMS FOR MICROSCOPIC EXAMINATION
Questions:
1. Label the parts of your drawings.
2. How can you differentiate the bacteria in the cheek cells from cell components?
After introducing the simple staining using a methylene blue dye to the specimen, we can
easily distinguish and differentiate the bacteria in the cheek cells from its cell components. We
all know that bacteria cells are much smaller than eukaryotic cells (cheek cells) so by
comparing the sizes we can easily locate the bacteria cells and it is found inside the cheek
cell. By comparing also the shapes, we can differentiate the bacteria cells from the cell
components. The large dark blue oval shaped structures are nuclei while smaller pill shaped
structures are bacteria on the cell surface.
3. Explain why the microorganisms in the negative staining are not stained?
Based on the principle of negative staining wherein it use nigrosin and india ink, being an
acidic dye and not really a bacterial stain, it readily gives up a hydrogen ion (proton) and the
chromophore of the dye becomes negatively charged. Since the surface of most bacterial
cells is negatively charged, the cell surface repels the stain. The glass of the slide will stain,
but the bacterial cells will not. The bacteria will show up as clear spots against a dark
background due to the fact that the two pigments can’t penetrate the microorganisms.
4. Of what value is Gram staining?
Gram staining has great diagnostic value because of its differential staining properties for
specific bacteria thus revealing little internal structure.It is also considered as one of the most
crucial staining techniques, it is perhaps the most important differential staining procedure in
microbiology because the great majority of bacteria are either gram-positive or gram-
negative. . For its clinical significance, Gram stain is often the initial diagnostic test for the
evaluation of infections.
5. Why are the steps in Gram staining so carefully standardized?
The steps in Gram staining must be carefully standardized so that we can reduce errors and
make Gram stain a precisely controllable and meritorious test. The application of the
decolorizer is the most critical stage of the Gram procedure. There is danger of incorrect
results from both over and under decolorization; therefore, the decolorizer and the technique
should be as carefully standardized as possible.The strict standardization of the method is
more important than the choice of method.
6. Why is it necessary to use young culture (24-hr old culture) in Gram staining?
First, older bacterial cells may have cell wall damage, making them seem gram-negative even
if the species is gram-positive. As a result, for Gram staining, it is ideal to utilize fresh
bacterial cultures. Gram staining should be performed on young, actively developing cultures.
For an accurate gram stain, the cell wall must be intact. Older cultures may have cell wall
breaks and often produce gram-variable results with a mix of pink/red cells among blue/purple
cells.
7. How will differentiate acid fast from non-acid fast bacteria? Explain the
differences between acid fast and non-acid fast bacteria.
The major distinction between acid fast and non acid fast bacteria is that acid fast bacteria
resist decolorization by acid after taking a stain, whereas non acid fast bacteria are easily
decolored by acid after staining. Acid fast bacteria have a thin layer of peptidoglycans on
their surface. Mycolic acid is bound to carbol-fuchsin after the primary stain, carbol-fuchsin, is
added to the slide containing bacteria. As a result, even after decolorization, the acid fast
bacteria stain pink. Because the carbol-fuchsin stain is incapable of penetrating the cell wall
of non acid fast bacteria, the stain is not retained by the cell wall of non acid fast bacteria.
The non acid fast bacteria absorb the counterstain, which appears blue under the
microscope.
8. Why is spore staining important?
The spore staining is a differential stain which selectively stains bacterial endospores.
Endospore staining is used to distinguish bacterial spores from other vegetative cells and
spore formers from non-spore formers. As such, it enables the differentiation of structures
and, as a result, the characterization of a cell based on its physical and chemical properties

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NAME Kristine Mae M.

Merhan DATE April 19, 2022

4. Of what value is Gram staining?

5. Why are the steps in Gram staining so carefully standardized?

8. Why is spore staining important?

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