September 8 Lab Notes

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. 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. 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. 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. 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. 2. start from last dilute. take .1 mL out using pipette and squirt onto middle of plate. do this for all dilutes
  3. 3. dip hockey stick in alcohol and set on fire (do not keep in fire too long it will shatter!)
  4. 4. run hockey stick back and forth while rotating plate (make sure you re sterilize hockey stick every time)
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