BLOCK-iT Fluorescent Oligo as RNAi Transfection Control
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实验试剂
实验步骤
1. Handling the BLOCK-iT™ Fluorescent Oligo
4) Take precautions to ensure that the stock solution does not become contaminated with RNase
- Use RNase-free sterile pipette tips and supplies for all manipulations.
- Wear gloves when handling reagents and solutions.
2. Using the BLOCK-iT™ Fluorescent Oligo for Cationic Lipid-Mediated Transfection
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The amount of BLOCK-iT™ Fluorescent Oligo to use depends on the growth rate and transfection efficiency of the mammalian cells. If you are transfecting your mammalian cell line for the first time, we recommend evaluating several concentrations of lipid and varying the final concentration of the BLOCK-iT™ Fluorescent Oligo from 10 to 200 nM to determine the optimal amount of BLOCK-iT™ Fluorescent Oligo to use to obtain a strong fluorescence signal.
Note: For most cell lines tested (e.g. HEK293, A549, HeLa), we obtain a readily detectable fluorescence signal when using 100 nM BLOCK-iT™ Fluorescent Oligo for transfection. - Prepare and seed mammalian cells at a density recommended by the manufacturer of the transfection reagent you are using.
- Prepare lipid-BLOCK-iT™ Fluorescent Oligo complexes as directed by the manufacturer of the transfection reagent you are using. Always dilute the BLOCK-iT™ Fluorescent Oligo immediately before transfection (i.e. do not store diluted Oligo) and into an appropriate medium. We recommend diluting the BLOCK-iT™ Fluorescent Oligo into Opti-MEM® I Reduced Serum Medium (Catalog no. 31985-062) available from Invitrogen.
- Assess fluorescent uptake at 6 to 24 hours post-transfection. The fluorescence signal may be detected at longer time points depending on the transfection efficiency and growth rate of the cells.
3. Using the BLOCK-iT™ Fluorescent Oligo for Electroporation
