Accell Red Cyclophilin B Control siRNA

siRNA delivery without a transfection reagent


Validated, fluorescent positive control siRNA targeting the PPIB housekeeping gene in human, mouse, or rat. Cytoplasmic fluorescence permits qualitative assessment of Accell siRNA passive delivery.
Accell Red Cyclophilin B Control siRNA is a fluorescent control reagent that provides highly reliable qualitative assessment of Accell siRNA uptake as determined by fluorescent microscopy to evaluate cytoplamic localization of the dye-labeled Accell siRNA or FACS analysis to determine general uptake. In addition, it acts as a positive control targeting Cyclophilin B. Also known as peptidylprolyl isomerase B (PPIB), cyclophilin B is abundantly expressed in most mammalian cells and because it is non-essential, knockdown of the corresponding mRNA does not affect cell viability.

Highlights

  • Targets accession numbers: NM_000942 (Human) or NM_011149 (Mouse) or NM_022536 (Rat)
  • Novel siRNA modifications facilitate uptake, stability, specificity and knockdown efficiency
  • Accell modifications also provide stability against nuclease-mediated degradation
  • Accell Red is labeled with  DY-547 (Cy3 analog). Absorbance/ Emission Max is 557/570 nm, a Cy™3, Rhodamine or PE filter can be used

 

Experimental considerations

 

  • Accell siRNA works at a higher concentration than conventional siRNA; recommended 1 µM working concentration
  • Delivery may be inhibited by the presence of BSA in serum. Optimization studies with serum-free media formulations (Accell Delivery Media) or < 2.5% serum in standard media is recommended
  • Full-serum media can be added back after 48 hours of incubation, optimal mRNA silencing is typically achieved by 72 hours, or up to 96 hours for protein knockdown
  
HazardousNo
Shipping ConditionAmbient
Stability at Recommended Storage ConditionsAt least 12 months
Storage Condition-20 C
Efficient delivery of Accell Red siRNA to SH-SY5Y

Efficient delivery of Accell Red siRNA to SH-SY5Y cells

Efficient delivery of Accell Red siRNA to SH-SY5Y

Uptake of Accell siRNA confirmed by fluorescence microscopy. SH-SY5Y cells were treated with 1 μM Accell Red Cyclophilin B Control siRNA in Accell delivery media. (Red fluorescence = Cytoplasmic localization of Accell siRNA; Blue = Nuclear Hoechst dye).


Cell types demonstrating effective silencing with Accell siRNA

Cell types demonstrating effective silencing with Accell siRNA

Cell types demonstrating effective silencing with Accell siRNA

Internal validation and peer-reviewed publications report numerous successes with difficult-to-transfect cell types. See the References tab for a list of publications.


The Accell siRNA application protocol simplifies targeted gene knockdown

The Accell siRNA application protocol simplifies targeted gene knockdown

The Accell siRNA application protocol simplifies targeted gene knockdown

The Accell siRNA application protocol simplifies targeted gene knockdown (A) Combine Accell siRNA with Accell delivery media (or other low- or no-serum media) (B) Add Accell delivery mix directly to cells and incubate for 72 hours.


References

  1. View the published references citing successful Accell siRNA application.

Citations

  1. Accell siRNA reagents in neuronal cells

    Vagnoni, A., et. al., Calsyntenin-1 mediates axonal transport of the amyloid precursor protein and regulates Aβ production., Human Molecular Genetics, 2012, 21;13 2845–2854 [rat primary cortical neurons (E18)]

    Suzuki, S., et. al., Differential Roles of Epac in Regulating Cell Death in Neuronal and Myocardial Cells J. Biol. Chem., Jul 2010; 285: 24248 – 24259 [primary mouse cortical neurons (E15-17)]

    Dolga, A.M. et al. TNF-alpha-mediates neuroprotection against glutamate-induced excitotoxicity via NF-kappaB-dependent up-regulation of K2.2 channels. J Neurochem 107, 1158-1167 (2008). [mouse primary cortical neurons]

    Dreses-Werringloer, U. et. al. A Polymorphism in CALHM1 Influences Ca2+ Homeostasis, Ab Levels, and Alzheimer’s Disease Risk. Cell, 27 June 2008; (133) 1149–1161 [SHSY-5Y; human neuroblastoma]

    Sebo, J. et al. Requirement for Protein Synthesis at Developing Synapses J. Neurosci. 2009; 29. 9778-9793. [rat primary hippocampal neurons]

    Mergenthaler, P., et. al., Mitochondrial hexokinase II (HKII) and phosphoprotein enriched in astrocytes (PEA15) form a molecular switch governing cellular fate depending on the metabolic state. PNAS Nov 2011 www.pnas.org/cgi/doi/10.1073/pnas.1108225109 [extended duration silencing in rat primary cortical neurons]

  2. Accell siRNA reagents in immunological cells

    Winning, S. et. al., Acute Hypoxia Induces HIF-Independent Monocyte Adhesion to Endothelial Cells through Increased Intercellular Adhesion Molecule-1 Expression: The Role of Hypoxic Inhibition of Prolyl Hydroxylase Activity for the Induction of NF-B J. Immunology, July 2010; 185; 1786 -1793 [THP-1 monocytes]

    Saini, V. et. al, CXC Chemokine Receptor 4 Is a Cell Surface Receptor for Extracellular Ubiquitin J. Biol. Chem., May 2010; 285: 15566 – 15576 [THP-1 monocytes]

    Perry, J.W. et. al., Endocytosis of Murine Norovirus 1 into Murine Macrophages Is Dependent on Dynamin II and Cholesterol J. Virol. 2010; 84:6163-6176 [murine macrophages]

    Steenport, M. et. al., Matrix Metalloproteinase (MMP)-1 and MMP-3 Induce Macrophage MMP-9: Evidence for the Role of TNF-a and Coclooxygenase-2. J. Immunology. doi:10.4049/jimmunol.0901925 Nov 2009 [RAW264.7 macrophages]

    Lai, C.B., Zhang, Y., Rogers, S.L. & Mager, D.L. Creation of the two isoforms of rodent NKG2D was driven by a B1 retrotransposon insertion. Nucleic Acids Res., gkp174 (2009). [mouse NK cell line]

    Mookherjee, N. et. al., Intracellular Receptor for Human Host Sefense Peptide LL-37 in Monocytes. J. Immunol. 2009; 183. 2688-2696 [THP-1; human monocytes]

    Smirnov, D. et. al. Genetic Analysis of Radiation-induced Changes in Human Gene Expression. Nature, 28 May 2009; 459 doi:10.1038/nature07940 [immortalized B cells]

    McElligott, A.M. et. al., The Novel Tubulin-Targeting Agent Pyrrolo-1,5-Benzoxazepine-15 Induces Apoptosis in Poor Prognostic Subgroups of Chronic Lymphocytic Leukemia. Cancer Research, Oct 2009; 10.1158/0008-5472.CAN-09-0131 [PGA-1; EBV-transformed chronic lymphocyctic leukemia (CLL) B cell line]

     

  3. Accell siRNA reagents in vivo

    Nakajima, H., et. al., A rapid, targeted, neuron-selective, in vivo knockdown following a single intracerebroventricular injection of a novel chemically modified siRNA in the adult rat brain. J. Biotechnology. Jan 2012 157(2): 326-333. [brain injection]

    Gonzalez-Gonzalez, E. et. al., Silencing of Reporter Gene Expression in Skin Using siRNAs and Expression of Plasmid DNA Delivered by a Soluble Protrusion Array Device (PAD) Molecular Therapy; 2010; doi:10.1038/mt.2010.126 [mouse intradermal injection]

    Bonifazi, P. et. al., Intranasally Delivered siRNA Targeting PI3K /Akt /mTOR Inflammatory Pathways Protects from Aspergillosis. Mucosal Immuno., Nov 2009 doi:10.1038/mi.2009.130 [in vivo intranasal delivery]

    DiFeo, A. et al. KLF6-SV1 Is a Novel Antiapoptotic Protein That Targets the BH3-Only Protein NOXA for Degradation and Whose Inhibition Extends Survival in an Ovarian Cancer Model Cancer Res. 69, 4733–41 (2009) [in vivo mouse model]

    Li, Q., et. al., Silencing MAP Kinase-activated Protein Kinase -2 Arrests Inflammatory Bone Loss. J. Pharmacol. Exp. Ther., Mar 2011; 336-642. [direct injection to rat gingival tissue; in vivo model of periodontal bone loss]

  4. Accell siRNA reagents in primary, stem, tumor and other cell types

    Liao, M., et. al., Inhibition of Hepatic Organic Anion-transporting Polypeptide by RNA Interference in Sandwich-cultured Human Hepatocytes: An in vitro Model to Assess Transporter-mediated Drug-drug Interactions Drug Metabolism and Deposition, Aug 2010; 38:9 1612-1622 [freshly isolated human hepatocytes]

    Byas, S. et. al., Human Embryonic Stem Cells Maintain Pluripotency after E-Cadherin Expression Knockdown FASEB J, Apr 2010; 24: lb172 [H9 stem cell lines]

    Desai, S. et. al., PRDM1 Is Required for Mantle Cell Lymphoma Response to Bortezomib Mol. Cancer Res., Jun 2010; 8: 907-918

    Barbieri, I., et. al., Constitutively Active Stat3 Enhances Neu-Mediated Migration and Metastasis in Mammary Tumors via Upregulation of Cten Cancer Research, Mar 2010; 70: 2558 - 2567.

    Bartholomeusz, C. et al. PEA-15 Induces Autophagy in Human Ovarian Cancer Cells and is Associated with Prolonged Overall Survival. Cancer Res. 68, 9302-9310 (2008). [OVCA 420; ovarian carcinoma]

    Tunquist, B. et. al., Mcl-1 Stability Determines Mitotic Cell Fate of Human Multiple Myeloma Tumor Cells Treated with the Kinesin Spindle Protein Inhibitor ARRY-520 Mol. Cancer Ther., Jul 2010; 9: 2046 – 2056 [multiple myeloma cell lines]

    Chetane, M. et. al., Interleukin-7 mediates glucose utilization in lymphocytes through transcriptional regulation of the hexokinase II gene Am J Physiol Cell Physiol, Jun 2010; 298: C1560 - C1571 [lymphocytes]

    Peters, GA. et. al. The double-stranded RNA-binding protein, PACT, is required for postnatal anterior pituitary proliferation. PNAS 106:26 10696-10701 (2009). [GH3; rat somatolactotrophs (pituitary cell line) and LßT2 gonadotrophs; basophilic cell of the anterior pituitary]