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Gently flood the smear with crystal violet and leave for 1 minute. Gram stain procedure - Gram staining a sampleġ. This kills the microbes in the smear and fixes the sample to the slide, be careful not to overheat the sample however as this can distort cellular morphology.

Once the smear has air dried, pass the smeared slide through a flame two or three times. Resuspend a loop of colony material in sterile phosphate buffered saline (PBS) and then proceed as for a liquid culture.ģ. If the source material is from a bacterial plate: For very dense cultures it may be necessary to pre-dilute your culture to ensure individual bacterial cells can be seen under a microscope following staining. Gently smear the droplet in a circular motion into an area of approximately 1 cm diameter. If preparing your slide from a liquid bacterial culture:ĭab a small drop culture onto the slide using a sterile loop. Ensure you use a pencil as many inks are removed by the reagents used in the staining procedure.Ģ. Label a clean glass microscope slide with your sample identification. Gram stain procedure - Preparing a sampleġ. One such useful classification – if a bacterium is Gram positive or Gram negative - is based on the structure of bacterial cell walls. Depending on the characteristic being studied, bacterial species may be broken down into broad groups, but taken together this information can narrow the possible identities greatly. This includes characteristics like their shape (bacilli vs cocci for example), growth in particular nutrients and preference for high or low oxygen environments. But even without getting into the molecular nitty gritty, there are phenotypic differences between groups of bacteria that can be used to differentiate them. Bacterial species, and even specific strains can be differentiated using a number of molecular techniques such as PCR, quantitative PCR, genome sequencing and mass spectrometry. Being able to differentiate bacterial species is important for a host of reasons, from diagnosing infection or checking food safety, to identifying which species it is that gives a cheese it’s fantastic character.