What makes Accell™ 2.0 siRNA different from the previous generation?

Accell 2.0 siRNA incorporates next-generation design criteria and continuous transcriptome realignment to ensure each self-delivering siRNA reflects the latest gene annotations. This enhances on-target precision while maintaining reliable transfection-free performance in difficult-to-transfect cells.

How does Accell 2.0 siRNA reduce off-target effects?

Accell 2.0 siRNA uses updated transcriptome data and advanced ranking algorithms to prioritize highly specific sequences. Refined design parameters minimize seed-mediated off-target activity while preserving robust gene silencing.

Do Accell 2.0 siRNAs require transfection reagents?

No. Accell 2.0 siRNAs are engineered with optimized self-delivering chemistry, enabling effective cellular uptake without transfection reagents or electroporation.

In what cell types can Accell 2.0 siRNA be used?

Accell 2.0 is particularly well-suited for primary cells, neurons, immune cells, stem cells, and other difficult-to-transfect models where traditional siRNA delivery methods are inefficient or cytotoxic.

Are Accell 2.0 siRNAs guaranteed to knock down my target gene?

Yes. Accell 2.0 siRNAs are supported by a guaranteed gene knockdown standard when used under recommended experimental conditions (See Guarantee tab).

In what formats are Accell 2.0 siRNAs available?

Accell 2.0 siRNA is offered as individual siRNAs, set of 4 siRNAs and in siRNA SMARTpool format (a blend of four distinct siRNA designs targeting the same gene). Plate-based formats are available to support screening workflows through the Cherry pick custom plating tool.

Are Accell 2.0 siRNAs available for multiple species?

Yes. Pre-designed Accell 2.0 siRNAs are available for human, mouse and rat targets.

How does Accell 2.0 siRNA support screening workflows?

Accell 2.0 libraries are available for focused gene-family collections and custom cherry-pick collections, enabling targeted screening in models where transfection-based approaches are not feasible.

Our siRNA knockdown guarantee

Accell 2.0 siRNA reagents (SMARTpool and three of four individual siRNAs) are guaranteed to silence target gene expression by at least 75% at the mRNA level as measured by qPCR when demonstrated to have been used under optimal delivery conditions (confirmed using functionally validated positive control and measured at  72 hours after transfection and using 1µM siRNA).

If all the steps to assess and optimize conditions are carried out as described in our "Guaranteed silencing with Accell siRNA in 3 easy steps" article, you are guaranteed at least 75% target gene silencing by the SMARTpool and/or three of the Set of 4 individual siRNAs.

Please reach out to our scientific support team for any questions or for further guidance on optimizing the use of your siRNA products.

Effectively optimize Accell™ 2.0 siRNA delivery conditions and help control for variables in ongoing experiments using fluorescent or unlabeled positive and negative controls. Accell™ 2.0 Control siRNA Kits combine species-specific controls to support assessment of Accell technology in difficult-to-transfect cells. Control kits are available with validated components.

Experimental considerations

• Accell™ 2.0 siRNA is typically used at a higher concentration than conventional siRNA; a recommended starting concentration is 1 µM 
• Delivery may be affected by the presence of BSA in serum. Optimization using 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. mRNA silencing is typically observed around 72 hours, with protein knockdown potentially requiring up to 96 hours
• Choose from positive controls (targeting commonly expressed genes), fluorescent or unlabeled negative controls, or an Accell™ 2.0 Control siRNA Kit

Accell 2.0 Positive Control Reagents	Species	Catalog Number
Accell 2.0 Cyclophilin B Control siRNA	Human, Mouse, Rat	D2-001920-0Y
Accell 2.0 Cyclophilin B Control siRNA Pool	Human, Mouse, Rat	D2-001920-Y0
Accell 2.0 GAPD Control siRNA	Human, Mouse	D2-001930-0Y
Accell 2.0 GAPD Control siRNA Pool	Human, Mouse	D2-001930-Y0
Accell 2.0 Green Cyclophilin B Control siRNA	Human, Mouse, Rat	D2-001970-0Y
Accell 2.0 Red Cyclophilin B Control siRNA	Human, Mouse, Rat	D2-001975-0Y
Accel 2.0 eGFP Control siRNA	N/A	D2-001940-01
Accell 2.0 eGFP Control siRNA Pool	N/A	D2-001940-10
Accell 2.0 Negative Control Reagents	Species	Catalog Number
Accell 2.0 Non-targeting siRNAs	Human, Mouse, Rat	D2-001910-0X
Accell 2.0 Non-targeting siRNA Pool	Human, Mouse, Rat	D2-001910-10
Accell 2.0 Green Non-targeting siRNA	Human, Mouse, Rat	D2-001950-01
Accell 2.0 Red Non-targeting siRNA	Human, Mouse, Rat	D2-001960-01
Accell 2.0 Control siRNA Kits	Species	Catalog Number
Accell 2.0 Control siRNA Kits (Green)	Human, Mouse	K2-005000-G1-0Y
Accell 2.0 Control siRNA Kits (Red)	Human, Mouse	K2-005000-R1-0Y

Peer-reviewed publications demonstrate effective silencing in numerous cell types
Human Suspension Cell Lines	BDCM, IM-9, Jurkat, K662, THP-1, Jeko-1 and Mino MCL
Human Adherent Cell Lines	A-375, ARPE-19, CaCo-2, CG4, DLD-1, DU-145, EC52, GTM-3, MR32, LAN5, LNCaP, MIA PaCa-2, NCI/ADR-RES, OVCAR-3, OVCA-420, PGA1, PNT2, SH-SY5Y, SK-N-BE and SK-N-SH neuroblastoma, SK-BR3, SKOV3, TE15, U251, U87MG, U937
Mouse Adherent Cell Lines	ES-D3, H9c2, LbT2, LNK, mouse oviduct epithelial cells, mProx24, PC-12
Human Differentiated Stem Cells	Adipocytes and osteoblasts from hMSC
Human Primary Cells	HUVEC, HUASMC, hMSC, PBMC, astrocytes, hypothalamic neurons, T cells, NK cells, macrophages, immortalized B cells, esthesioneuroblastoma cells, hepatocytes, bronchial smooth muscle cells
Mouse and Rat Primary Cells	Cortical neurons, hippocampal neurons, striatal neurons, Naïve CD4+ T cells, LN T cells, D1 T cells, RAW264.7 macrophages, hepatocytes, neonatal ventricular cardiomyocytes, fibroblasts derived from genetically-modified mice, bone marrow-derived DC, NOD CD4+ CD25- splenic cells, NOD BMMs, VSMCs, somatolactotroph GH3, McArdle 7777 cells (hepatoma cell line), osteoclast precursors, calvarial (cranial) organ, pituitary cell types derived from primary tumors
In vivo Animal Applications	Please contact Scientific Support for guidance

Please see our list of recommended reading that features Dharmacon Accell siRNA.

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Application of Accell 2.0 siRNA simplifies targeted gene knockdown

1. Combine Accell 2.0 siRNA with Accell siRNA delivery media (or other reduced serum media).
2. Add Accell 2.0 delivery mix directly to cells and incubate for 72 hours.
   Figure created in with BioRender.com

 

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Accell 2.0 siRNA maintains performance of gene target knockdown set by Accell siRNA in neurons derived from induced WTC-11 iPSC cells

 

Characterization of target mRNA knockdown by Accell and Accell 2.0 siRNAs in cultured neurons. Data are shown relative to neurons treated with non-targeting control (NTC) siRNA. Experimental schematic (A) and relative gene expression in WTC-11-derived neurons (B).  Neurons derived from WTC-11 iPSCs were cultured in differentiation medium for 14 days, with daily medium replenishment. On day 14, 50 µL of medium was removed and replaced with 50 µL fresh medium containing 2 µM Accell or Accell 2.0 pooled siRNA designs (4 siRNA designs at 0.5 µM each) or 2 µM Accell PPIB control (Cat# D-001920-01), resulting in a final siRNA concentration of 1 µM in all conditions. An identical concentration of Accell Non-targeting Control siRNA #1 (D-001910-01) was used for each target NTC condition. siRNA mixtures were replenished by half-medium change on differentiation days 16 and 18, for a total of three siRNA additions. Total RNA was extracted on differentiation day 20 for downstream processing. Figure A is created with BioRender.com.

 

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Performance of target mRNA knockdown in matrix-free 3D spheroids is maintained with Accell 2.0 siRNA

Characterization of Accell and Accell 2.0 siRNA-mediated mRNA knockdown in ECM-free HCT-116 spheroids. Control, PTEN (A), and PPP2R1B (B) mRNA expression were quantified in HCT-116 spheroids after 144 hours of siRNA treatment. (C) Dosing timeline for Accell 2.0 siRNA delivery. Spheroids were generated by seeding 3,000 HCT-116 cells into ECM-free, 96-well ultralow attachment round-bottom plates in HCT-116 medium (McCoy’s 5A medium supplemented with 10% FBS and 2 mM L-glutamine). After two days, 50% of the medium was removed and replaced with 2 µM Accell or Accell 2.0 single or pooled siRNA designs (4 siRNAs at 0.5 µM each) for a final concentration of 1 µM siRNA. Matching concentrations of Accell Non-targeting Control siRNA #1 (D-001910-01) or Accell PPIB control siRNA (D-001920-01) were added on the same schedule. After three rounds of siRNA replenishment, spheroids were lysed, total nucleic acid was collected, and RT-qPCR was used to measure target mRNA abundance. Data are shown relative to target expression in spheroids treated with non-targeting control siRNA. Figure C is created with BioRender.com.

 

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Target protein knockdown in matrix-free 3D spheroids using Accell 2.0 siRNA

Characterization of Accell siRNA-mediated cyclophilin B (PPIB) target protein knockdown in ECM-free HCT-116 spheroids. Data are presented relative to expression in spheroids treated with non-targeting control (NTC) siRNA. (A) Western blot and (B) quantification of Cyclophilin B protein expression in HCT-116 spheroids determined from (A) after 144-h siRNA treatment. Each lane is representative of an independent pool of spheroids (N=3). (C) Immunocytochemistry images of serially sectioned HCT-116 spheroids after 144-h siRNA treatment with an Accell PPIB Control Pool or NTC Pool. Spheroids were then collected, fixed in 2% paraformaldehyde, embedded in optimal cutting (O.C.T.) temperature compound, flash frozen, and serially sectioned. Material was incubated with 1:200 dilution of primary antibody against PPIB (anti-cyclophilin B, Abcam ab16045) and secondary incubation with 1:500 dilution of Alexa 488-conjugated Goat-anti-rabbit antibody (Invitrogen A11034) and 1:1000 dilution of Hoechst 33342 (Molecular Probes H-3570). Scale bar = 200 µM.

 

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Robust gene target knockdown in primary lung fibroblasts using Accell 2.0 siRNA

 

IMR-90 human lung fibroblasts (5,000 cells / well) were plated in 96-well plates in IMR-90 media (EMEM supplemented with 10% FBS, L-glutamine and 0.1 mM non-essential amino acids). After adhesion and outgrowth for 16-18 hours, IMR-90 cells were at 80% confluency and experiments were initiated. After media removal, cells were washed with D-PBS and immersed in 100 mL Accell media with 1 µM Accell 2.0 siRNA. After 5 hours of incubation, 100 mL of IMR-90 media with 4% FBS was added to each well to result in 2% FBS. After culturing for 72 hours, total RNA was extracted and used for downstream RT-qPCR analysis of target mRNA abundance from PTEN, PPP2R1B, AKT1, and AKT2 genes relative to Non-targeting Control sample (NTC).

 

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Accell control siRNA demonstrates effective uptake and silencing in organotypic brain slices

Accell siRNA shows increased uptake with extended incubation. 250 µM cerebellar sections were prepared, cultured, and incubated for 3 hours (A) and 72 hours (B) with Accell Red Non-targeting control (NTC) siRNA (Cat# D-001960-01-05) before inspection by microscopy.