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L6 Check it out IV - Coggle Diagram
L6 Check it out IV
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Smear preparation
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quality of smear
Its necessary to heat fix the smear so as to cause denaturion of the proteins on the surface of the bacteria, causing their adhesion to the slide, hence preventing washing off of the bacteria during staining
If smear is not completely air dried prior to heat fixation, brief heat fixing will not result In the adhesion of bacterial cells to the slides, while longer heat fixing to dry up the smear will cause the alteration of the shape and size of bacteria
bacteria staining
Concept of staining
With the lack of contrast of unstained bacteria, it is hard to visualize under even a good light microscope
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Most commonly used dyes are basic in nature; carrying positively-charged chromophore (color-bearing ions)
Basic dyes are attracted to negatively charged components within the bacteria cells including proteins and nucleic acid
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mechanism of gram stain
Due to differences in the bacterial cell wall structure, Gram-positive and Gram-negative bacteria are differentiated during the decolorization with 95% alcohol (decolorizer):
Gram-negative bacteria – the lipids in the outer membrane dissolve, leading to the removal of crystal violet-iodine complex; the colorless cell wall will pick up the red/ pink color from the counterstain safranin
Gram-positive bacteria – the thick peptidoglycan layer traps the crystal violet-iodine complex, hence retaining the purple/ blue color
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