prior to kirby-baur test and selective in differential make sure you suspend 1 mL of sterile broth in a test tube. then take a sample of a pure culture and place in sterile broth.
**Kirby-Baur Test* (used for antibiotic susceptibility)
Steps:__
- 1.make a "lawn" of bacteria with broth and bacteria on mueller-hinton agar (lawn: back and forth swabbing covering entire surface of plate)
- 2.use flame and alcohol to sterilize tweezers. Use tweezers to place antibiotic disks on plate
- make sure you place one negative disk on plate
- placement of disks should be about 1/2 and inch apart
Selective In Differential Agar (used for metabolism of bacteria which helps identify)
Steps:
- 1. streak for isolated colonies on EMB (eosin-methylene blue agar)- the dark red one! and MSA (mannitol salt agar) -the light red one!
Serial Dilution Technique
take .1 mL out of original sample and place in test tube #1. then take out .1 mL from test tube #1 and place in test tube #2. continue this process through all diluted test tubes. Then plate 0.1 mL out of each test tube onto its own separate plate and use hockey sticking technique. (HINT: if you start from last dilute and work backwards you only need to use one pipette)
Hockey-Sticking Technique
Steps:
- 1. label plates 1:10^n (n being the number of dilutes) for example 2 dilutes would be 1:10^2, 1:10, 1:1 (the last one being original sample)
- 2. start from last dilute. take .1 mL out using pipette and squirt onto middle of plate. do this for all dilutes
- 3. dip hockey stick in alcohol and set on fire (do not keep in fire too long it will shatter!)
- 4. run hockey stick back and forth while rotating plate (make sure you re sterilize hockey stick every time)
