Difference between revisions of "UW-Stout/Hydroxychloroquine SP21"
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==Precautions== | ==Precautions== | ||
Use gloves and safety glasses for this experiment. Hydroxychloroquine is a potent chemical used to treat malaria and some autoimmune conditions. However, side effects include but are not limited to nausea, headache, dizziness, rash. | Use gloves and safety glasses for this experiment. Hydroxychloroquine is a potent chemical used to treat malaria and some autoimmune conditions. However, side effects include but are not limited to nausea, headache, dizziness, rash. | ||
| + | ==Calibration Protocol== | ||
| + | #Vortex the yeast culture briefly to resuspend the yeast cells. | ||
| + | #Prepare a stock solution of hydroxychloroquine 3000 ug/mL. | ||
| + | #Set up 8 wells according to the table: | ||
| + | #Aliquot 50 uL of each stock and add 50 uL of yeast culture to each sample. | ||
| + | #To measure cell growth over a period of 24 hours, set up the plate reader as follows: | ||
| + | *Temperature: 30°C | ||
| + | *Mode: Kinetic | ||
| + | *Wavelength: 600 nm | ||
| + | *Interval: 5 minutes | ||
| + | *Total run time: 24 hours | ||
| + | *Shake before reading: 30 seconds. | ||
| + | #Transfer the assay plate to the reader and read for 24 hours. | ||
==Protocol== | ==Protocol== | ||
Revision as of 09:44, 27 April 2021
Materials
- Corning COSTAR 96-well clear flat-bottom assay plate
- Wild-type yeast in 2x synthetic complete media at an OD600 of 0.1-0.2
- Hydroxychloroquine, 300 ug/uL solution in sterile water (as calculated in the calibration protocol)
- P20, P200 micropipettors
Equipment
Precautions
Use gloves and safety glasses for this experiment. Hydroxychloroquine is a potent chemical used to treat malaria and some autoimmune conditions. However, side effects include but are not limited to nausea, headache, dizziness, rash.
Calibration Protocol
- Vortex the yeast culture briefly to resuspend the yeast cells.
- Prepare a stock solution of hydroxychloroquine 3000 ug/mL.
- Set up 8 wells according to the table:
- Aliquot 50 uL of each stock and add 50 uL of yeast culture to each sample.
- To measure cell growth over a period of 24 hours, set up the plate reader as follows:
- Temperature: 30°C
- Mode: Kinetic
- Wavelength: 600 nm
- Interval: 5 minutes
- Total run time: 24 hours
- Shake before reading: 30 seconds.
- Transfer the assay plate to the reader and read for 24 hours.
Protocol
- Vortex the yeast culture briefly to resuspend the yeast cells.
- Set up seven wells aliquoting 50uL of the stock hydroxychloroquine and 50uL of yeast culture. Use the seventh well for the wild-type yeast strain.
- The final concentration of hydroxychloroquine is 300ug/mL.
- To measure cell growth over a period of 24 hours, set up the plate reader as follows:
- Temperature: 30°C
- Mode: Kinetic
- Wavelength: 600 nm
- Interval: 5 minutes
- Total run time: 24 hours
- Shake before reading: 30 seconds
- Transfer the assay plate to the reader and read for 24 hours.
- Note: Repeat the experiment a few times to account for natural variation.
