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AllStars Hs Cell Death Control siRNA

For siRNA transfection optimization and positive control experiments in human cells

  • Optimization and positive control experiments made easy
  • Valuable control for high-throughput RNAi
  • Simple evaluation by light microscopy
  • Highly effective for a wide range of cell types

AllStars Hs Cell Death Control siRNA is a blend of highly potent siRNAs targeting ubiquitously expressed human genes that are essential for cell survival. Knockdown of these genes induces a high degree of cell death, which is visible by light microscopy. Transfection efficiency can be quickly estimated by simply observing cells by straightforward light microscopy 48–96 hours after transfection of AllStars Hs Cell Death Control siRNA, avoiding the need for any complex or laborious downstream assay. AllStars Hs Cell Death Control siRNA is a useful tool for siRNA transfection optimization and can be used routinely as a positive control.

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AllStars Hs Cell Death siRNA
Positive cell death phenotype control,
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Positive Controls
Negative Controls
Reporter Controls
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The AllStars Hs Cell Death Control siRNA is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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High level of cell death.
[A]  SW620 cells or  [B] SW480 cells (5 x 103 cells per well in 96-well plates) were transfected with 20 nM of AllStars Hs Cell Death Control siRNA or AllStars Negative Control siRNA or an siRNA targeting PLK (a well-known target for induction of cell death). Untransfected cells and cells treated with transfection reagent only were also analyzed. After 72 hours, cell death was quantified using a CellTiter-Blue assay (Promega). AllStars Hs Cell Death Control siRNA resulted in high levels of cell death in both cell types. (Data kindly provided by Dr. Amanda Hummon and Dr. Thomas Reid, National Cancer Institute, Maryland, U.S.A.)
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Cell death in primary cells.
Primary human NHBE cells (2 x 104) were transfected with 10 nM, 20 nM, or 100 nM AllStars Hs Cell Death Control siRNA or 100 nM nonsilencing siRNA (AllStars Negative Control siRNA) using HiPerFect Transfection Reagent. After 48 hours, cell death was observed by light microscopy.
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Easy siRNA transfection optimization.
MCF-7 cells (5 x 103) in 96-well plates were transfected with various amounts of AllStars Hs Cell Death Control siRNA or 50 nM nonsilencing siRNA (AllStars Negative Control siRNA) using HiPerFect Transfection Reagent. After 72 hours, cell death was observed by light microscopy. Optimal conditions were 5 nM siRNA and 1.5/2 μl HiPerFect Reagent or 2 nM siRNA and 2 μl HiPerFect Reagent.
Performance
Fast and easy detection in a wide range of cell types

The performance of AllStars Hs Cell Death Control siRNA is validated for a wide range of cell types (see table).

AllStars Hs Cell Death Control siRNA Validated Cell Types
Cell line Cell type
293 Human embryonic kidney cells
A549 Human alveolar basal epithelial cells
CaCo Human colonic carcinoma cells
HeLa Human cervical cancer cells
HeLa S3 Human cervical cancer cells
HepG2 Human hepatoma cells
Huh7 Human hepatoma cells
MCF-7 Human breast adenocarcinoma cells
MDA-MB-231 Human breast adenocarcinoma cells
ME180 Human cervical squamous carcinoma cells
SW 480 Human colon adenocarcinoma cells
Primary cells Cell type
HUVEC Primary human umbilical vein endothelial cells
NHBE Primary normal human bronchial epithelial cells
NHLF Primary normal human lung fibroblast cells
Highly efficient knockdown and cell death

Knockdown efficiency using AllStars Hs Cell Death Control siRNA has been quantified by real-time RT-PCR and is >95% for each target mRNA.

The AllStars Hs Cell Death Control significantly outperforms other commercially available toxic control siRNAs in both the level of cell death induced and in the wide range of cell types in which it has been validated. Consistently high levels of cell death are achieved using AllStars Hs Cell Death Control, ensuring effective control experiments (see figure High level of cell death).

Principle
Easy transfection optimization

When establishing RNAi in start-up experiments or in a new cell line, multiple transfections should be performed under different conditions to determine the optimal conditions for maximum transfection efficiency. These experiments can be performed using AllStars Hs Cell Death Control siRNA (see figures Easy siRNA transfection optimization and Cell death in primary cells). Transfection conditions that result in the greatest degree of cell death compared with transfection with a nonsilencing control siRNA can be maintained in future experiments.

Reliable high-throughput RNAi

In high-throughput RNAi screens, the use of internal positive controls is essential to ensure that optimal transfection conditions are maintained on every plate of the screen. Positive controls that can be analyzed phenotypically have the advantage of allowing transfection efficiency to be observed immediately. Using AllStars Hs Cell Death Control siRNA on every plate allows rapid and inexpensive estimation of the transfection rate on individual plates.

Procedure

Using AllStars Hs Cell Death Control siRNA, transfection efficiency can be quickly estimated by simply observing cells by straightforward light microscopy 48–96 hours after transfection, avoiding the need for any complex or laborious downstream assay.

AllStars Hs Cell Death Control siRNA can be ordered in tube format in 1 nmol, 5 nmol, or 20 nmol amounts. This control is also available in 96-well plates and 384-well plates (FlexiPlate siRNA) in 1 nmol, 0.25 nmol, or 0.1 nmol amounts.

Applications
Start-up RNAi experiments
Transfection optimization
High-throughput RNAi
Sample to Insight
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