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| Problem with old cultures |
As cultures age, some cells begin to die and their cell walls break down, and are no longer able to prevent the alcohol from leaching out the crystal violet....creates false reading of gram pos bacteria to look like gram neg |
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bhoward86 Sun, 27 Sep 2009 23:15:03 GMT |
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| Gram neg cell wall |
only a single layer of peptidoglycan surrounded by layers of various types of lipids.
when ethyl alcohol tries to decolorize, lipids are dissolved and crystal violet is leached out of the cell |
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bhoward86 Sun, 27 Sep 2009 23:13:40 GMT |
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| Gram pos cell wall |
thick and composed of many layers of peptidoglycan
when ethyl alcohol tries to decolorize, peptidoglycan becomes dehydrated, shrinks, and becomes impermeable, thus locking in purple stain |
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bhoward86 Sun, 27 Sep 2009 23:13:40 GMT |
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| permeability theory |
explanation for the mechanism of gram stain;
bacteria stain differently because of chemical and structural differences in their cell walls |
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bhoward86 Sun, 27 Sep 2009 23:13:40 GMT |
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| differentiating step |
decolorizer; removes primary stain from some bacteria (gram neg), which now become colorless, but remains in others (gram pos) |
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bhoward86 Sun, 27 Sep 2009 23:13:40 GMT |
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| counterstain |
safranin |
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bhoward86 Sun, 27 Sep 2009 23:08:06 GMT |
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| decolorizer |
ethyl alcohol |
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bhoward86 Sun, 27 Sep 2009 23:08:06 GMT |
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| mordant |
iodine |
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bhoward86 Sun, 27 Sep 2009 23:08:06 GMT |
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| primary stain |
crystal violet |
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bhoward86 Sun, 27 Sep 2009 23:08:06 GMT |
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| function of mordants |
causes a stain to become more tightly bound to the cell; iodine does this by intensifying the ionic chemical bond btw crystal violet and the bacteria |
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bhoward86 Sun, 27 Sep 2009 23:07:30 GMT |
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| 4 steps |
1. primary stain 60 sec, cover slide
rinse briefly w/ water
2. mordant 60 sec, droppers full
3. decolorizer dropper full to rinse quickly
rinse immediately w/ water
4. counterstain 30 sec, cover slide
blot slide dry |
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bhoward86 Sun, 27 Sep 2009 23:18:09 GMT |
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| Separates bacteria in 2 groups: |
Gram positive
Gram negative |
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bhoward86 Sun, 27 Sep 2009 23:07:30 GMT |
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Invented by _____
Year____
Was trying to differentiate ______ from ______ |
Hans Christian Gram
1884
bacteria; human tissue |
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bhoward86 Sun, 27 Sep 2009 23:07:30 GMT |
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