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U937

Multiporator Transfection Protocol Protocol No. 4308 915.037 11/1999 Cell line U937, human leukemic monoblasts Transfection with Plasmid pEGFP-N1 (in bidistilled H2O) Electroporation buffer Eppendorf Hypoosmolar Electroporation Buffer (PH) Culture medium RPMI / 10% FCS Cuvette Eppendorf, 4 mm gap width, 800 l Temperature RT (20-25 C) Reference Dr. Steven Grant Box 980230 Medical College of Virginia Richmond VA 23 298 USA
Phone +1 804 828 5168 Fax +1 804 225 3788 e-mail: stgrant@hsc.vcu.edu
  1. Harvest the cells in the exponential growth phase and centrifuge them (for 5-10 minutes, 200 x g, at room temperature).
  2. Resuspend the cells in RPMI / 0.5% FCS, determine the number of cells and centrifuge them (for 5-10 minutes, 200 x g, at room temperature). Remove supernatant.

    Note: The overall incubation time in the Eppendorf Electroporation Buffer must not exceed 30 minutes to guarantee a successful electroporation!

  3. Resuspend the cells in Hypoosmolar Electroporation Buffer. When doing so, set the cell concentration to 2 x 106 cells/ml.
  4. Add and m
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