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  • TransPass D2 Protocol 1: Transfection in the presence of serum

    Introduction

    The amounts below are given for a 12-well plate format. Use Table 1 to adjust the reagent volumes for other plate sizes.

    Protocol

    1. Plate cells (in complete growth medium containing 5-10% serum and no antibiotics/antimycotics) at an appropriate density so that they will reach 70-80% confluence at the time of transfection.
    2. Mix 1.5 µg plasmid DNA in 100 µl serum-free medium.
    3. Gently mix TransPass D2 Transfection Reagent prior to pipetting. Do not vortex. Add 1.5-4 µl to the DNA/medium mix from step 2. Mix gently by flicking the tube.
    4. Incubate at room temperature for 20-30 minutes to form the transfection complex.
    5. Add the transfection complex mixture to cells. Rock the plate gently in order to evenly disperse the complex mixture.
    6. Return the plate to the incubator and incubate 24-72 hours before assaying.
    7. Replace medium as needed to maintain healthy cells.