BCH4160/2011:Notebook/Jigeshs Lab Notebook 2011/2011/10/25

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Binding Fluorescence: Preliminary Preparations

The purpose was to prepare solutions of concentrations 2-30μL in preparation for future binding fluorescence lab.

Materials

Materials used in this experiment:

  • Lipid films made using protocol on October 20th
  • Peptide (MW = 642.79g)
  • Sodium phosphate buffer (made using [[]] protocol)
  • Fluorescence spectrophotometer

Procedure

Preliminary Preparations of Peptide Solutions

The purpose of protocol is to prepare peptide solutions of concentrations 2μM, 10μM, and 30μM for future binding fluorescence lab.

  1. Add 0.001g peptide to 1mL sodium phosophate buffer. The pH should be ≈ 7.0.
  2. Use a UV spectrophotometer to measure the absorbance of the peptide solution at 280nm.
  3. Calculate the concentration of the peptide solution using the measured absorbance and Beer's Law.
    • Beer's Law: A=εlc → c = (A÷(εl))
    • ε=5500 M-1 cm-1
    • Wavelength = 280 nm
    • l = 0.2cm
  4. Once the concentration of the stock peptide solution is known, make 1mL dilute peptide solution in sodium phosphate buffer (2μM, 10μM, and 30μM peptide).

Results & Data

Contact

  • Who has experience with this protocol?

or instead, discuss this protocol.




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