Anisha Patel COSMOS UC Davis Cluster 1: Biotechnology
Anisha Patel COSMOS UC Davis Cluster 1: Biotechnology
Anisha Patel COSMOS UC Davis Cluster 1: Biotechnology
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PART 114. Add 10-15 µl of Laemmli buffer to a microtube.15. Add beta mercaptoethanol. (5% of total volume).16. Add a protein sample volume that will contain 100 – 150 µg of protein to the tube withthe other reagents.17. Heat the microtubes at 70 degrees Celsius for 2 minutes, then place on ice18. Pour SDS Buffer into the Western Blot tank.19. Position the acrylamide gels in the gel holder assembly and immerse into the tank.20. Fill the inner compartment (between the two gels) with SDS Buffer.21. Carefully load the samples in the wells.22. Place the lid on the tank and plug it into the power source (100V).23. Run the apparatus at 100V until the samples have passed the stacking gel.24. Turn the voltage up to 160V and allow the samples time to separate; use a pre-stainedmolecular weight marker to determine the end-point of the electrophoresis.25. Cut filter paper in approximately7 x 20 cm pieces.26. Cut PVDF membrane to 7 x 20 cm.27. Pre wet the PVDF membrane using 100% methanol for 10 seconds and immerse indH2O.