Exp_2_KEY_S11

Exp_2_KEY_S11 - Experiment 2 KEY MCB120L, S11 Overlay...

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1 Experiment 2 KEY MCB120L, S11 Overlay Spectra 1. The molar absorption coefficients, ε , for tyrosine and tryptophan at 280 nm molar extinction coefficient, ε (M -1 cm -1 ) at 280 nm Tyr (Y) ε = (0.35 / 2.5 x 10 -4 M · 1 cm) = 1400 M -1 cm -1 Trp (W) ε = ( 1.4 / 2.5 x 10 -4 M · 1 cm) = 5600 M -1 cm -1 2. The absorption coefficients , a , for BSA, lysozyme and gelatin at 280 nm absorption coefficient, a (mg/ml) -1 · (cm) -1 at 280 nm (sample values) [BSA] = 0.4 mg/ml a = (0.265 / 0.4 mg/ml · 1cm) = 0.66 (mg/ml) -1 (cm) -1 [lysozyme] = 0.4 mg/ml a = (1.010 / 0.4 mg/ml · 1cm) = 2.53 (mg/ml) -1 (cm) -1 [gelatin] = 0.4 mg/ml a = (0.044 / 0.4 mg/ml · 1cm) = 0.11 (mg/ml) -1 (cm) -1 3. A brief explanation for performing the Bradford protein assay at 595 nm. A protein assay can be developed anywhere there is a measurable change in absorbance between the reagent and the reagent plus protein. Examining the overlay spectra for the Bradford reagent and the Bradford reagent plus protein, there are many wave lengths that can be chosen to measure the difference, including a negative change in absorbance (on the left side of the overlay spectra; it is a measurable change!). The 595 nm wave length is chosen since it gives the greatest change in absorbance ( ie it is the most sensitive change).
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This note was uploaded on 11/10/2011 for the course MCB 120L taught by Professor Fairclough during the Spring '08 term at UC Davis.

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Exp_2_KEY_S11 - Experiment 2 KEY MCB120L, S11 Overlay...

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