Cell Fractionation of U937 Cells in the Absence of High-speed Centrifugation

This protocol allows researchers to obtain distinct fractions of a cell without the use of specialized equipment, expensive reagents or complex techniques.The primary advantage of this protocol is the simplicity of the techniques which make it modifiable and easy to perform.Demonstrating the procedure will be Matt Deragon, Rob Haluska and Alexa Hodges.Three graduate students from my laboratory.To begin centrifuge the prepared cell suspension to create a pellet.Remove the supernatant and re-suspend the pellet in room temperature PBS.Centrifuge the cell suspension at 400 xg for ten minutes.Then remove the supernatant and re-suspend the cell pellet in ice cold buffer.Prepare buffer A solution according to the text protocol.Centrifuge the cell suspension at 400 xg for ten minutes and remove the supernatant.Re-suspend the pellet in buffer A solution to a final concentration of 200 million cells per ml, and pipette gently to break up any clumps.After this incubate the cell suspension on an end over end rotator at 4