Purpose: Use procedure to identify if bacteria is gram positive or gram negative. This would help someone know how to treat an infection. Prelab: 1. Why do you run the loop in the fire? 2. Why do you have to wait for the water to evaporate before heat- fixing? To kill anything that is on the loop…so you do not contaminate your experiment. If you don’t and you “fix it”, the water will boil and the bacteria will boil away with the water leaving you with no bacteria.

3. Fixation: Waving the slide through a flame several times The purpose is to make the bacterial cells stick to the slide 4. Primary Stain: Called “crystal violet” Stains bacteria purple Gram + will soak this in 5. Mordant: Called “iodine” Forms a complex with the crystal violet in gram + bacteria, making it harder to wash off throughout the remainder of the procedure 6. Ethanol: Does nothing to gram + bacteria Washes stain off of gram - bacteria

8. Counter Stain: Called “safranin” Stains gram – bacteria red What color is the bacteria? Gram Positive is Purple Gram Negative is Red 11. What is a “Smear” Bacteria rubbed into a drop of water

 The smaller the drop of water you use for your smear, the faster it will evaporate.  You DO NOT NEED a big “chunk” of bacteria…this will be too much to see.  Do not rush the evaporation process. Be PATIENT!  Read the directions and follow them!  If it says, “drop” then only use a drop. If it says, “apply” then cover the slide (squirt). o Record which tube you took your bacteria from and record the number in the appropriate space. o Record whether or not it is gram + or gram - o Draw what you see in your slide o Record the shape and arrangement by using the Latin terms in your notes o Write the genus name of the bacteria based on the above.