Lumit® dsRNA Detection Assay

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Assay to Detect and Quantify Double-Stranded RNA Contamination in IVT mRNA Samples

  • Specific: No cross reactivity with ssRNA or DNA
  • Quantitative: Not dependent on sequence or size (≥30bp)
  • Simple Workflow: No time-consuming wash steps or immobilization
  • Available through our Early Access program

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Lumit® dsRNA Detection Assay
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Quantify dsRNA with Confidence

High-quality in vitro transcription (IVT) products are crucial for research and RNA therapeutic development. However, IVT can generate double-stranded RNA (dsRNA) byproducts. This contaminant can have significant negative downstream effects on drug substances and products, such as high immunogenicity. Sensitive and quantitative assessment of dsRNA is critical to control and optimize mRNA therapeutic and vaccine manufacturing processes.

The Lumit® dsRNA Detection Assay is a novel technique to quantify dsRNA in a sample. Lumit® Assay technology is based on NanoBiT® luciferase complementation. NanoBiT® subunits, SmBiT and LgBiT, are genetically fused to dsRNA binding proteins. In the presence of dsRNA, the SmBiT and LgBiT subunits are brought together to form an active NanoBiT® Luciferase, generating a luminescent signal proportional to the amount of dsRNA in a sample. The result is a concentration-dependent luminescent signal for quantitative and specific measurement of dsRNA.

Simple Protocol Streamlines dsRNA Quantification

The assay is solution-based and homogeneous:

  • No wash or sample transfer steps needed
  • Amenable to high-throughput applications
Depiction of Lumit® dsRNA Detection Assay protocol.

Size and Sequence-Independent dsRNA Assay

The Lumit® dsRNA Detection Assay can sensitively quantify dsRNA independent of sequence and size.

Better Performance than Commercial ELISA Kits

The Lumit® dsRNA Detection Assay provides an accurate linear range of 0.08–5ng/ml. Commercial dsRNA ELISA kits do not accurately quantitate dsRNA at these concentrations.

Detect and Quantify dsRNA in Complex IVT Samples

The Lumit® dsRNA Detection Assay can accurately quantify dsRNA in IVT samples with mixed RNA species.

Detecting dsRNA

The Lumit® dsRNA Detection Assay can sensitively quantify dsRNA independent of sequence and size.

Line graph showing titrations of dsRNA analyzed with the Lumit® dsRNA Detection Assay.

Sample

Slope

R2

LMW Poly(I:C) Batch 1 1.16 0.997
LMW Poly(I:C) Batch 2 1.14 0.999
HMW Poly(I:C) 1.15 0.997
60bp dsRNA 1.11 0.999
142bp dsRNA 1.09 0.998

Titrations of dsRNA were analyzed with the Lumit® dsRNA Detection Assay.

Samples:

  • LMW Poly(I:C): Synthetic dsRNA analog (0.2–1kb).
  • HMW Poly(I:C): Synthetic dsRNA analog (1.5–8kb).
  • 60bp dsRNA: Synthesized dsRNA comprised of the first 60bp from the SARS-CoV-2 spike ORF.
  • 142bp dsRNA: Commercially available “standard”.

Benchmarking Lumit® Assay Versus dsRNA ELISA Kit

The Lumit® dsRNA Detection Assay provides an accurate linear range of 0.08–5ng/ml. Commercial dsRNA ELISA kits do not accurately quantitate dsRNA.

Line graph showing Lumit® dsRNA Detection Assay range of luminescence detection compared to ELISA.

Sample

Expected (ng/ml)

Observed (ng/ml)

60bp dsRNA 0.16 0.15
60bp dsRNA 1.3 1.3
142bp dsRNA "standard" 1.6 0.91
Poly(I:C) 2.5 1.9
IVT Product 1.8
Line graph showing Lumit® dsRNA Detection Assay range of absorbance detection compared to ELISA.

Sample

Expected (ng/ml)

Observed (ng/ml)

60bp dsRNA 5 not detected
60bp dsRNA 75 not detected
142bp dsRNA "standard" 20 2.2
Poly(I:C) 5 6.1
IVT Product 0.3

Lumit® dsRNA Detection Assay range of detection compared to ELISA. Both assays were tested for their ability to detect different species of dsRNA contaminants:

  • 60bp dsRNA: Synthesized dsRNA comprised of the first 60bp of the SARS-CoV-2 spike ORF.
  • 142bp dsRNA “standard”: Commercially available, chemically defined 142bp long dsRNA sequence intended to be used as a positive control in dsRNA detection.
  • Poly(I:C): Synthetic dsRNA analog (0.2–1kb).
  • IVT Product: In-house-generated 429bp IVT product using T7 RiboMAX™ Express Large Scale RNA Production System, purified with LiCl2.

Specific Detection of dsRNA in IVT Products

The Lumit® dsRNA Detection Assay can accurately quantify dsRNA in IVT samples with mixed RNA species.

Depiction of Lumit® dsRNA Detection Assay quantification of dsRNA in IVT samples with mixed RNA species. IVT samples were treated ± RNase I(f) (ssRNA-specific); total RNA (QuantiFluor® RNA) was quantified.
Depiction of Lumit® dsRNA Detection Assay quantification of dsRNA in IVT samples with mixed RNA species. IVT samples were treated ± RNase III (dsRNA-specific); dsRNA (Lumit®) was quantified.
Bar graphs showing that digestion of ssRNA with RNase I(f) significantly reduced total RNA but did not impact the amount of dsRNA detected.
Bar graphs showing that digestion of dsRNA with RNase III into fragments ≤20bp results in a complete loss of activity in the Lumit® dsRNA Detection Assay.

Lumit® dsRNA Detection Assay quantification of dsRNA in IVT samples with mixed RNA species. IVT samples were treated ± RNase If (ssRNA-specific) or RNase III (dsRNA-specific); total RNA (QuantiFluor® RNA) and dsRNA (Lumit®) were quantified.

  • Left: Digestion of ssRNA with RNase If significantly reduced total RNA but did not impact the amount of dsRNA detected.
  • Right: Digestion of dsRNA with RNase III into fragments ≤20bp results in a complete loss of activity in the Lumit® dsRNA Detection Assay.

Especificaciones

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Contenido

Item Part # Presentación

dsRNA Standard

CS355701 1 × 15μl

Lumit® dsRNA Sensor SmBiT

CS355702 1 × 15μl

Lumit® dsRNA Sensor LgBiT

CS355703 1 × 15μl

dsRNA Assay Buffer (5X)

CS355704 1 × 8ml

Lumit® Detection Substrate B

VB405A 1 × 160μl

Certificado de Análisis

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Condiciones de Almacenaje

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Especificaciones

You are viewing: CS355706 Cambiar Configuración

Contenido

Item Part # Presentación

dsRNA Standard

CS355701 5 × 15μl

Lumit® dsRNA Sensor SmBiT

CS355702 5 × 15μl

Lumit® dsRNA Sensor LgBiT

CS355703 5 × 15μl

dsRNA Assay Buffer (5X)

CS355704 5 × 8ml

Lumit® Detection Substrate B

VB405A 5 × 160μl

Certificado de Análisis

Search by lot number

Condiciones de Almacenaje

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