Dojindo,HilyMax/1/H357,A549 细胞的信号转导通过 TNF-α 刺激得到证实
2022年4月30日Principle
HilyMax 很容易与 DNA 相互作用，因为阳离子脂质体 (+) 和阴离子 DNA(-) 自发形成 DNA 脂质体复合物。 DNA-HilyMax 复合物的总电荷为正电荷，因此 DNA-HilyMax 复合物静电结合在阴离子细胞表面，并通过内吞作用将 DNA 引入细胞。
Procedure
The procedure is extremely simple. No exchange of the media is required during the whole process,because serum in the medium does not interfere with the transfection.
来自 A549 细胞的信号转导通过 TNF-α 刺激得到证实。 为了检测细胞反应，用 HilyMax 或 L2000 转染 IL-8 依赖性荧光素酶表达载体。 信号转导反应在刺激和抑制后检测为荧光素酶活性。 使用 HilyMax 的信号响应与受刺激细胞中表达的 IL-8 的量有关。如果您对此问题感兴趣，请单击此处获取更多信息。
Comparison Data in Insect Cell<New Data>
Data was kindly provided by Dr. Takashi Suzuki at Max Planck Institute of Neurobiology.
Culture ConditionCell: S2(Schneider 2) cell, 200,000 cells/wellMedia: Schneider’s Drosophila medium with 10% FCSAntibiotics: 50 units Penicillin/ml, 20 µg Streptomycin/mlMicroplate: 24-well plate
Transfection ConditionVector: 1 µg/well <pAct Gal4(6 kb), pUAS-mCD8::GFP(10 kb)>Reagent: 5 µl/well <HilyMax or Cfectin>*Medium was changed in 4 hours after transfection.*Schneider’s Drosophila medium(without seum and antibiotics) was used for the complex preparation.
GFP Transfected Cells with HilyMax
Transfection Efficiency in Various Cell Lines
Comparison Data of Transfection Efficiency with Commercially Available Reagents
The tranfection efficiency of HilyMax is higher than commercially available reagents in widely used cells. GFP expressed DNA was transfected using HilyMax and the other transfection reagents in serum containing medium. The amount of expressed protein indicates the transfection efficiency.
1. The volume of HilyMax is not enough. Increase the HilyMax volume.2. Cell density is too high. Reduce the cell density. The appropriate cell density for transfection is about 40-90%.3. HilyMax reagent is not be dissolved completely. Please check that the HilyMax solution is homogeneous.4. Incubation time for the preparation of HilyMax and DNA complex is too long.5. Your culture medium for DNA-HilyMax complex formation contains serum and/or antibiotics. Please use serum-free and antibiotics-free medium for the complex formation.
1. Reduce the amount of DNA and/or HilyMax. Prepare the complex.2. Cell density is too low. Reduce the cell density. The appropriate cell density for transfection is about 40-90%.