Team:Calgary/Notebook/Protocols/BenzylpenicillianAssay2

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Benzylpenicillin Assay 2

Reagents and Materials

  • 96-well plate
  • Bromothymol Blue pH Indicator
  • 60 mg/mL Benzylpenicillin
  • Purified β-lactamase 1µg/µL
  • Commercial β-lactamase 1µg/µL
  • pH 8 Water

Protocol

  1. Dilute β-lactamase to the appropriate concentrations in 100 µL of pH 8 water. Add this to the wells.
  2. Add an additional 50 µL of pH 8 water to the wells.
  3. Add 10 µL of bromothymol blue pH indicator to these wells.
  4. Add 100 µL of 60 mg/mL benzylpenicillin to the wells.
  5. Measure the change from blue to yellow using a spectrophotometer at an absorbance wavelength of 616 nm.