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Young bacterial cultures should be used for the Gram stain method to accurately identify Gram-positive and Gram-negative bacteria. The amount of alcohol applied during differential staining must be carefully controlled so as not to over-decolorize the sample and incorrectly identify bacteria. An even, thin smear is also important to avoid uneven decolorization from the alcohol treatment.
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this documents will help you to have better understanding about precaution in gram stain method.
Young bacterial cultures should be used for the Gram stain method to accurately identify Gram-positive and Gram-negative bacteria. The amount of alcohol applied during differential staining must be carefully controlled so as not to over-decolorize the sample and incorrectly identify bacteria. An even, thin smear is also important to avoid uneven decolorization from the alcohol treatment.
Young bacterial cultures should be used for the Gram stain method to accurately identify Gram-positive and Gram-negative bacteria. The amount of alcohol applied during differential staining must be carefully controlled so as not to over-decolorize the sample and incorrectly identify bacteria. An even, thin smear is also important to avoid uneven decolorization from the alcohol treatment.
Always use a young culture because older cultures of Gram-positive
bacteria tend to lose the ability to retain the crystal violet-iodine complex and appear to be Gram-negative; but some bacteria are naturally only weakly Gram-positive. The amount of alcohol treatment (the differential stage) must be judged carefully because over-treatment washes the crystal violetiodine complex from Gram-positive bacteria and they will appear to be Gram-negative. Take care to make an even smear otherwise alcohol will continue to wash the violet/purple colour from thick parts of the smear while thin parts are being over-decolorised. At the end of the procedure, check that the labeling has not been washed off by the alcohol. Dont despair if the stained smear is not visible to the naked eye; this may happen with a Gram-negative reaction. Specimen stained slides are available from suppliers.
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