QIAcuity Cell and Gene Therapy (CGT) dPCR Assays

For vector genome titration with superior accuracy, reproducibility and speed on the QIAcuity Digital PCR System

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dPCR CGT Assay GFP (FAM)

Cat. No. / ID: 250236

For 500x12 µl reactions (20x): QIAGEN Cell and Gene Therapy assay for GFP to be used with the QIAcuity dPCR system.
$1,085.00
Assay
GFP
ITR2/5
bGH polyA
WPRE
SV40 promoter
SV40 polyA
CMV promoter
hGH polyA
CMV enhancer
AMP resistance
Dye
FAM
HEX
Cy5
The dPCR CGT Assay GFP (FAM) is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.

✓ 24/7 automatic processing of online orders

✓ Knowledgeable and professional Product & Technical Support

✓ Fast and reliable (re)-ordering

Features

  • A broad offering of ten different wet-lab validated dPCR CGT assays
  • Assays with different fluorophore choices allow multiplexing to sustain vector integrity
  • An easy and fast workflow comparable to qPCR

Product Details

Adeno-associated virus (AAV) is a widely used viral vector in gene therapy applications. However, the generation and purification of the viral vectors require rigorous quality control to enable safe and reliable dosing during clinical studies or patient care. The ability to accurately quantify vector titers and detect contamination is critical for safe and effective AAV-based gene therapies.

Performance

The QIAcuity Cell and Gene Therapy dPCR Assays are a broad offering of ten different wet-lab validated dPCR CGT assays that come in multiple fluorophores, enabling superior accuracy, reproducibility and a dynamic range of at least 4 orders of magnitude with speed in measuring viral titers in a multiplex setup. The assays work in conjunction with the QIAcuity Digital PCR System, QIAcuity Probe PCR Kit and QIAcuity Nanoplates, offering an end-to-end dPCR workflow comparable to qPCR, but delivering an absolute quantification of AAV vector genome copies in your sample. The assays have been designed with the requirements of biopharma manufacturing and QC in mind.

Principle

The principle of the dPCR reaction in the nanoplates is described here.

Dedicated CGT assays enable AAV quantification on the QIAcuity. These assays are validated and can be used in singleplex and multiplex reactions, and can be additionally combined with gene-of-interest assays, delivering:

  • Accurate quantification down to 0.3 copies/µl
  • High precision over a broad dynamic range
  • High accuracy across assays (independent of fluorophores) and operators
  • High precision (<10% deviation from the mean) independent of fluorophores and operators
  • Compatibility with both dPCR and qPCR readout

Procedure

The QIAcuity Cell and Gene Therapy dPCR Assays are provided in a 20x ready-to-use primer-probe mix, available in multiple fluorophore choices, and optimized for use with the QIAcuity Probe PCR Kit. These assays enable both singleplex and multiplex CGT applications, including viral vector titer and integrity measurements.

Applications

The QIAcuity Cell and Gene Therapy (CGT) dPCR Assays are used for various applications, including viral vector titer measurements, measuring vector genome integrity and determining potential plasmid contamination if using Amp plasmids for the production of AAVs. Incorporating these dPCR CGT Assays into the quality control process of gene therapy development means greater certainty in producing safe and potent treatments.

Supporting data and figures

FAQ

Do I need to use restriction enzymes when quantifying AAV genomes?

In some cases, it is recommended to perform a digest with compatible restriction enzymes. For example, when quantifying ITRs, the strong secondary structure of the terminal repeats might affect titration. Compatible restriction enzymes are indicated in the corresponding data sheets of the CGT dPCR assays.

FAQ ID - 3866
How can I resuspend the lyophilized assay?

Please resuspend the lyophilized assays in 330 µl TE buffer to obtain a 20× stock. Additional information can be found in the QSP and the data sheets.

FAQ ID - 3864
Any recommendations on how can I process my AAV samples?
Are the assays also compatible with qPCR?

Yes. The assays can also be used for vector genome titration in a qPCR.

FAQ ID - 3871
Can I use the CGT dPCR assays for the titer determination of RNA viruses?

Yes, you can. cDNA would be used in this case for quantification.

FAQ ID - 3869
In which channels can you detect the CGT dPCR assays?

Most of the assays are offered with a FAM, HEX, or Cy5 fluorophore, and can be detected in the Green, Yellow, or Crimson channel, respectively. SV40 poly A, hGH poly A, and Amp resistance assays are only available with a FAM or HEX fluorophore.

FAQ ID - 3863
How do I know if the assays are compatible with the sample DNA of interest?

An extended sequence context for each assay is provided in the corresponding data sheet. 

FAQ ID - 3870
Can they be used with lentiviral vectors?

Yes. The assays can also be used with lentiviral vectors. However, lentiviral vectors have an RNA genome. cDNA synthesis is crucial before performing the PCR for titer quantification.

FAQ ID - 3872
What quenchers are used?

The probes are double quenched with ZEN/TAO and IOWA black quenchers.

FAQ ID - 3865
Can I combine your CGT dPCR assays with custom designed assays?

Yes, the CGT dPCR assays can be used in singleplex and multiplex reactions. Custom designed assays can be added into a multiplex reaction according to customer needs.

FAQ ID - 3861
Can the CGT dPCR assays also be used for the analysis of AAV genome integrity?

Yes. When multiplexing different targets, it is possible to get additional information on genome integrity using the “Multiple occupancy” analysis in the QIAcuity Software Suite.

FAQ ID - 3867
Can the CGT dPCR assays also be used for the quantification of non-AAV samples?

Yes. They can be used as long as the assays match the region of interest.

FAQ ID - 3868
How many assays can be multiplexed?

Two to five assays can be combined into a multiplex reaction depending on the QIAcuity Digital PCR platform, hence the 2-plex or 5-plex instruments.

FAQ ID - 3862