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First, put on a lab coat, safety glasses, and nitrile gloves. Some solutions use hexane or dimethylformamide, so prepare those solutions in a fume hood.
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Prepare a 1 mM stock solution of fluorescein in water. Measure 0.0166 g of fluorescein and place it in a 50-mL volumetric flask.
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Add about 30 mL of deionized water to the flask, stopper it, and swirl it until as much solid has dissolved as possible. Then, fill the flask to the line with deionized water.
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Stopper the flask, wrap the top in plastic paraffin film, and invert it several times while firmly holding the stopper in place to thoroughly mix the solution. Label the flask with the solution's name and concentration and wrap it in aluminum foil to protect the dye from light. Label the outside of the foil as well.
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Prepare a 3 µM fluorescein test solution from the stock solution. Use a micropipette to transfer 150 µL of the stock solution to a clean 50-mL volumetric flask. Add about 30 mL of deionized water and swirl the flask to mix them.
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Fill the flask to the line with deionized water, stopper and seal it, and invert the flask several times until the solution is well mixed. Label the flask with the solution's name and concentration, wrap it in foil, and label the outside of the foil as well.
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Prepare 1 mM stock solutions and 3 µM test solutions of indigo in dimethylformamide and β-carotene in hexane in the same way. The masses of solid β-carotene and indigo that you will need to make the solutions are listed in the tables.
All stock solutions in 50 mL solvent |
Dye | Stock concentration | Mass | Solvent |
β-carotene | 1 x 10-3 (1 mM) | 0.027 g | Hexane |
Indigo | 1 x 10-3 (1 mM) | 0.013 g | DMF |
All test solutions diluted to 50 mL |
Dye | Test concentration | Stock volume | Solvent |
β-carotene | 3 x 10-6 (3 µM) | 150 µL | Hexane |
Indigo | 3 x 10-6 (3 µM) | 150 µL | DMF |
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Use the stock solution of β-carotene to make five more solutions of the concentrations listed in this table. Wrap the flasks in foil afterward.
All solutions diluted to 50 mL with hexane |
Dye | Concentration | Stock volume |
β-carotene | 1.9 x 10-6 (1.9 µM) | 95 µL |
β-carotene | 3.7 x 10-6 (3.7 µM) | 185 µL |
β-carotene | 7.5 x 10-6 (7.5 µM) | 375 µL |
β-carotene | 11 x 10-6 (11 µM) | 550 µL |
β-carotene | 15 x 10-6 (15 µM) | 750 µL |
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When you're finished, you'll have three micromolar solutions of fluorescein, β-carotene, and indigo dye for the first part of the lab and five β-carotene solutions with different concentrations for the second part of the lab. Store the solutions in a refrigerator for now if you have one. If not, store them at room temperature for up to 24 hours.