Viral Vector Lysis Kit

For lysis of viral vectors, such as different adeno-associated virus (AAV) serotypes and adenoviruses

Products

The Viral Vector Lysis Kit is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
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Cell and Gene Therapy Viral Vector Lysis Kit (100 rxn)

Cat. No. / ID:  250272

The kit includes: CGT Sample Stabilizer, DNase I, Nuclease-Free Water, CGT Lysis buffer, CGT DNase I buffer and CGT Dilution Buffer enough to process 100 AAV samples 
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₩294,000.00
Recommended restriction enzyme HpaII is not included in the kit.
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Cell and Gene Therapy Viral Vector Lysis Kit (1000 rxn)

Cat. No. / ID:  250273

The kit includes: CGT Sample Stabilizer, DNase I, Nuclease-Free Water, CGT Lysis buffer, CGT DNase I buffer and CGT Dilution Buffer enough to process 1000 AAV samples 
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₩1,826,000.00
Recommended restriction enzyme HpaII is not included in the kit.

Features

  • One kit and one protocol solution for AAV lysis enables standardization and quality control (QC) of current workflows
  • Consistent and reproducible measurement of viral titers for multiple serotypes (e.g., AAV serotypes)
  • Complete workflow combined with QIAcuity Cell and Gene Therapy (CGT) dPCR Assays and accurate measurement of AAV titers on the QIAcuity Digital PCR System

Product Details

Adeno-associated virus is a widely used viral vector in gene therapy applications. However, the process of generation and purification of the viral vectors requires precise quality control to enable safe and reliable dosing during clinical studies or patient care. The ability to accurately quantify vector titers as well as determine contaminations is key to safe and effective AAV-based gene therapies. 


With the introduction of the CGT Viral Vector Lysis Kit that includes reagents for either 100 or 1000 reactions, we now have an optimized and standardized workflow for viral vector lysis through to an accurate and precise way of determining the viral titer. The kit is also compatible with adenoviruses.

Performance

We offer a complete workflow from performing AAV lysis to quantification of the viral titer from cell lysates. The CGT Viral Vector Lysis Kit with its improved formula gives a consistent, robust, and accurate determination of the final titer. It works in conjunction with the QIAcuity Cell and Gene Therapy dPCR Assays, QIAcuity Digital PCR System, QIAcuity Probe PCR Kit and QIAcuity Nanoplate 8.5K, offering an end-to-end dPCR workflow comparable to qPCR but delivering absolute quantification of AAV vector genome copies in your sample.

Principle

The all-in-one solution for AAV lysis provides:

  • Standardization of AAV lysis with much easier standard operating procedure (SOP) implementation and QC
  • Consistent quantification of titers
  • Broad range of detection between 2.5 copies/µl to 15,000 copies/µl in QIAcuity Nanoplate 8.5k
  • Robustness of <10% CV between operators and assays

The CGT Viral Vector Lysis Kit in conjunction with the QIAcuity Cell and Gene Therapy dPCR Assays and the QIAcuity Probe PCR Kit running either singleplex or multiplex reactions on the QIAcuity instruments, now delivers a complete viral titer workflow.

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

 

Procedure

The CGT Viral Vector Lysis Kit is provided with reagents for 100 or 1000 reactions in two boxes for AAV vectors. The kit is suitable for the lysis of AAV2, AAV5, AAV6, AAV8 and AAV9. The lysates are optimized for QIAcuity Cell and Gene Therapy dPCR Assays in combination with the QIAcuity Probe PCR Kit. These assays enable singleplex as well as multiplex cell and gene therapy applications, including viral titer and vector copy number measurements. The kit does not include restriction enzymes (for e.g., Hpall) that are stated in the protocol (included in the Resources section below).

Applications

The CGT Viral Vector Lysis Kit is used for the lysis of AAV and adenoviruses for a range of applications including viral vector genome titer and vector copy number measurements.

Supporting data and figures

Resources

Brochures & Guides (2)
In this booklet, discover why and how to adapt dPCR to your area of research. Benefit from experimental data and discover complete workflows for optimal dPCR performance in cell & gene therapy, mutation detection, gene expression analysis, microbial detection and many more.
Quick-Start Protocols (2)
This protocol is optimized for the processing of viral vectors for genome titer quantification on the QIAcuity® digital PCR instrument using the dPCR CGT Viral Vector Lysis Kits (cat. nos. 250272, 250273). The kits are optimized for usage with the QIAcuity Probe PCR Kit (cat. nos. 250101, 250102, 250103) and the QIAcuity Cell and Gene Therapy (CGT) dPCR Assays.
This protocol is optimized for the processing of viral vectors for genome titer quantification on the QIAcuity® digital PCR instrument using the dPCR CGT Viral Vector Lysis Kits (cat. no. 250272, 250273). The kits are optimized for usage with the QIAcuity Probe PCR Kit (cat. no. 250101, 250102, 250103) and the QIAcuity Cell and Gene Therapy (CGT) dPCR Assays
Safety Data Sheets (1)
Certificates of Analysis (1)

FAQ

Do you have to use a restriction enzyme to relieve ITR secondary structures and increase target accessibility?
It is highly recommended and an important step in the protocol. ITR secondary structures reduce target accessibility and negatively affect quantification of ITR targets and non-ITR targets.
We recommend using the following HpaII restriction enzymes: Thermo Scientific™ HpaII (10 U/µL, cat. nos. ER0511, ER0512) or Invitrogen™ Anza™ 93 HpaII (cat. no. IVGN0936).
FAQ ID - 3974
Do you need to dilute the lysates before setting up the PCR mix?
The lysates must be diluted in total of at least 1:100 taking together the dilution after capsid lysis and the dilution in the PCR reaction. A total of 200x dilution is recommended.
FAQ ID - 3978

Do I need to use Proteinase K in the CGT Viral Vector Lysis Kit? What is the recommended concentration?
No, Proteinase K use is optional and does not interfere with the kit performance. It is recommended to use 2 µL of Proteinase K (RP107B) in a 50 µL reaction. The Proteinase K has a concentration of 20mg/mL with 800 U/mL.
FAQ ID - 3971
Is the kit compatible with unpurified in-process samples?
Yes, it is.
FAQ ID - 3965
Can I use the lysate from the CGT Viral Vector Lysis kit and combine it with another PCR system?
Yes, however, the complete workflow has been optimized with the QIAcuity Cell and Gene Therapy (CGT) dPCR Assays on the QIAcuity Digital PCR System to ensure standardization and reproducibility of viral vector titer determinations.
FAQ ID - 3979

How do I know if the CGT dPCR assays are compatible with the sample DNA of interest?
An extended sequence context for each assay is provided in the corresponding data sheets. 
FAQ ID - 3980

Are the CGT dPCR assays also compatible with qPCR?
Yes, the assays can also be used for vector genome titration in a qPCR setup.
FAQ ID - 3981

Is it possible to store the lysates? How can the lysates be stored?
Long-term storage of lysates is not recommended. If lysates must be stored, it is recommended to store them at 4°C. Do not freeze.
FAQ ID - 3973
Can I use MspI or HpaII restriction enzymes from other suppliers?
The HpaII enzymes from Thermo Fisher Scientific ER0511, ER0512 and IVGN0936 have been extensively tested and show high compatibility with the QIAcuity Probe PCR chemistry. Use of MspI enzymes is not recommended. Compatibility of alternative enzymes must be individually tested and may need additional optimization.
FAQ ID - 3975
How many AAV genome copies can I load into the QIAcuity?
Recommended detection range in the dPCR using a 8.5k nanoplate is between 2.5 cop/μL to 15,000 copies/μL.
FAQ ID - 3977
Can the CGT Viral Vector Lysis kit be used with assays of other suppliers or custom designed assays?
Yes. Optimization of the cycling and imaging conditions might be needed.
FAQ ID - 3982

What is the recommended HpaII concentration?
Enzyme concentration depends on sample input and incubation time. When following our protocol described in the handbook HpaII enzymes work in the following range:
  • IVGN0936: 0.025 – 0.125 U/µL
  • ER0511/ER0512: 0.25 – 0.5 U/µL
FAQ ID - 3976
Do you recommend storage of highly diluted AAV samples?
No, it is not recommended to store AAV samples in a highly diluted state.
FAQ ID - 3972