Lab 2A protocol

Lab 2A protocol - Chem 153L Lab 2A: Spectrophotometry &...

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Chem 153L Lab 2A: PROTOCOL : *Must dress in long pants, closed-toed shoes, lab coat, safety goggles, gloves A) Coomassie Blue (CB) Preparation 1. Obtain a test tube with the 0.100 mM, pH7 Coomassie Blue stock 2. Using the L1000 piston pipet, pipet 0.40 mL Coomassie Blue to a new test tube 3. Using a 5 mL serological pipet, transfer 1.6 mL distilled water to the test tube containing 0.40 mL Coomassie Blue; this is the 0.02 mM Coomassie Blue solution 4. Vortex this test tube briefly according to the guidelines learned in the previous lab B) Manual Spectrum at pH 7 1. Prepare blank with distilled water 2. Measure absorbance of the 0.02 mM Coomassie Blue solution at wavelengths from 450-750 nm, use a macrocuvet in doing this 3. Follow spectrophotometer instructions on page 10 of the lab manual 4. Print Results and label 5. Plot absorbance vs. wavelength, you should obtain a smooth curve, if not, repeat measurements 6. Use interpolation to estimate the wavelength of maximum absorbance, λ max C) Coomassie Blue Spectra at Neutral and Low pH *Use extreme caution with this portion of the lab since you will be using HCl, an acid. Discard the HCl waste in the appropriately labeled container in the lab. 1. Pipet 0.40 mL of 0.1 mM CB stock to a new test tube 2. Obtain a 0.02 mM CB solution by adding 1.6 mL of 2.5 M HL to this test tube 3. Vortex this solution carefully 4. Prepare blank with distilled water 5. For both neutral and low pH, use the automatic Wavelength Scan Mode and a macrocuvet, obtain spectra for both the acid CB solution just prepared and the
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This lab report was uploaded on 04/09/2008 for the course BIO CHEM 153L taught by Professor Kim during the Winter '08 term at UCLA.

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Lab 2A protocol - Chem 153L Lab 2A: Spectrophotometry &...

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