Talk:Knight:Beta-galactosidase assay/96 well format

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#Repeat until the well's solution looks totally clear.
#Repeat until the well's solution looks totally clear.
#Measure A<sub>420</sub> of each well in the plate reader.
#Measure A<sub>420</sub> of each well in the plate reader.
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 +
Concentration series:
 +
 +
4mg/mL -> 2mg/mL -> 1mg/mL -> 0.5 mg/mL -> 0.25 mg/mL -> 0.125 mg/mL -> 62.5 &mu;g/mL -> 31.25 &mu;g/mL -> 15.625 &mu;g/mL
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 +
-> 7.8125 &mu;g/mL -> 3.90625 &mu;g/mL -> 1.953125 &mu;g/mL -> 0.9765625 &mu;g/mL

Revision as of 14:01, 24 October 2007

A420 versus o-nitrophenol concentration

  1. Make up a solution of 7 mL to control for background absorbance in β-galactosidase assays
    • 800 μL permeabilization solution
    • 6 mL substrate solution without ONPG
    • 200 μL EZ rich media supplemented with kanamycin and AHL
  2. Make up 1mL of 4 mg/mL ONP.
  3. Add 350 μL of 4mg/mL solution to the first well.
  4. Move 175 μL of previous well to next well.
  5. Add 175 μL of background solution to that well.
  6. Repeat until the well's solution looks totally clear.
  7. Measure A420 of each well in the plate reader.

Concentration series:

4mg/mL -> 2mg/mL -> 1mg/mL -> 0.5 mg/mL -> 0.25 mg/mL -> 0.125 mg/mL -> 62.5 μg/mL -> 31.25 μg/mL -> 15.625 μg/mL

-> 7.8125 μg/mL -> 3.90625 μg/mL -> 1.953125 μg/mL -> 0.9765625 μg/mL

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