Capsid testing kit sets new standard for AAV quality control

In gene therapy, ensuring the quality of viral vectors is crucial. As adeno-associated viruses (AAV) continue to gain footing as a robust delivery vehicle for genetic material, developers face the challenge of distinguishing between empty, partially filled, and fully filled viral capsids. Unwanted empty capsids – which lack therapeutic genetic material – can increase production costs and pose risks of immune reactions.

To address these issues, Bio-Rad Laboratories has developed the Vericheck ddPCR™ Empty-Full Capsid Kit, a streamlined, validated solution that is designed to provide rapid, specific, and compliant measurement of capsid ratios. With Bio-Rad’s Droplet Digital PCR (ddPCR) technology at its core, the Vericheck Kit stands out by promising a precise, fast, and reliable solution for AAV characterization – crucial for safe and effective gene therapy development.

The crucial role of AAV quality control

AAV vectors are favored in gene therapy for their ability to deliver genes with minimal side effects. However, not all AAV particles are identical. Post-production, some contain the therapeutic genetic material (full capsids), while others are only partially filled or even empty.

In regulatory terms, the empty-full capsid ratio is a critical quality attribute (CQA) for AAV production. High proportions of empty capsids can not only reduce therapeutic efficacy but also increase the risk of immunotoxicity in patients. Traditionally, analytical ultracentrifugation (AUC) and transmission electron microscopy (TEM) have been relied upon to characterize these ratios. However, these methods can be very labor-intensive, costly, and dependent on specialized equipment, limiting their accessibility and scalability.

Precision, simplicity, and speed

Bio-Rad’s Vericheck ddPCR Empty-Full Capsid Kits redefines AAV characterization by providing a robust solution that is adaptable to diverse sample types, compatible with both purified and crude lysate samples, and capable of measuring capsid contents swiftly. The ddPCR-based Vericheck Kit can achieve high-precision results in a single eight-hour day, providing lab managers and gene therapy developers a practical tool for more rapid turnaround times and more reproducible data.

“One of the key benefits of the Vericheck Kit is its streamlined workflow,” Dipika Gurnani, Global Product Manager for Bio-Rad’s Digital Biology Group, explained. “While AUC might take up to four days and TEM a full day or two, the Vericheck Kit offers an eight-hour, one-day turnaround. This faster turnaround enables better production efficiency and supports in-process testing.” This advantage is crucial in a fragmented capsid content testing market where speed and ease of use can impact both production timelines and cost.

Comprehensive coverage for a range of AAV samples

The Vericheck Kit also offers versatility in sample types. AAV testing often requires samples from various stages of production. Purified viral vectors, which are most common in quality control, provide clear data due to reduced impurities. However, early-stage production samples, such as crude cell lysates, contain high levels of impurities and may require additional steps in some testing methods. The Vericheck ddPCR Kit is validated across these types, ensuring that researchers can analyze crude, partially purified, or purified samples with confidence in data accuracy.

“For early-stage development, where rapid data on empty-full ratios helps in process optimization, the Vericheck Kit is particularly valuable,” Gurnani highlighted. “This kit has been validated to support early-stage samples, providing lab managers with crucial flexibility.”

Reliable and scalable alternative to traditional methods

Traditional methods such as AUC and TEM are accurate but come with notable limitations. AUC separates viral particles based on density, providing detailed results but requiring complex setups and expert interpretation. TEM offers direct imaging for identifying capsid contents, yet demands time-consuming manual analysis and specialized equipment. By contrast, ddPCR eliminates these limitations by using droplet digital technology to separate, quantify, and analyze capsid particles with minimal manual intervention.

Bio-Rad’s Vericheck ddPCR Kit not only provides rapid and reliable empty-full capsid analysis but also offers consistent performance across batches and across various sample types. Additionally, the Vericheck Kit measures both capsid titer (total particles) and genome titer, giving a complete view of viral vector quality with less sample input. As Gurnani noted, the kit’s low input requirements mean it can deliver precise measurements even with minimal sample volumes, a valuable benefit in high-throughput environments.

Vericheck ddPCR Empty-Full Capsid Kit requires only small volume of crude lysate or purified samples, which is ideal for any step in the AAV vector development or manufacturing process.

Recent study highlights Vericheck ddPCR robustness

For those who may be new to ddPCR, Bio-Rad provides clear guidance and validation documentation to facilitate its integration into existing workflows. In a recent study, Bio-Rad researchers demonstrated the kit’s versatility by extending its application to measure the fill rate of capsids containing diverse genomic targets. The study found that the Vericheck ddPCR Kit consistently delivered accurate capsid titers across various sample types and genetic targets, including partial genomes.

The study highlighted that ddPCR can discern partial capsids with greater clarity, thanks to the ability to selectively target and quantify specific AAV genomes. This capability offers new insights for characterizing AAV integrity, especially when comparing vectors from different sources. Results indicated that ddPCR provides a robust, flexible, and efficient method for capsid integrity analysis, allowing gene therapy labs to extend their assessments of AAV genome quality beyond standard measurements.

Simplified workflow with Bio-Rad’s support

For many laboratories, adopting new technology can cause challenges in terms of validation and integration. Bio-Rad has addressed these concerns by offering comprehensive on-site training for the Vericheck ddPCR Kit, along with ongoing technical support. “We provide our customers with a smooth transition by completing all necessary validations on our end and offering a detailed protocol and support for implementation,” Gurnani notes. “Additionally, our technical support teams are available to address questions throughout the process.”

This level of support is critical for labs that are implementing ddPCR for the first time, as it simplifies the learning curve and ensures consistent, reliable results. Moreover, Bio-Rad provides guidance on internal validation studies for labs that may require additional compliance documentation. This guidance ensures that gene therapy labs can quickly integrate the Vericheck Kit and maintain alignment with regulatory standards.

Broadening the potential of ddPCR in gene therapy

Looking ahead, the potential for ddPCR extends well beyond AAV capsid analysis. “We’ve worked with DNA and RNA quantification for over a decade, and now we’re thrilled to apply ddPCR for protein quantification,” Gurnani shares. This capability is especially exciting in the context of gene therapy, where the demand for versatile, scalable testing solutions is growing rapidly.

The Vericheck ddPCR Kit’s flexibility to accommodate various AAV genomes and capsid types also points to a future where labs can achieve more granular insights into vector integrity, packaging efficiency, and therapeutic viability. As Bio-Rad continues to innovate, the expanded capabilities of ddPCR could streamline other critical aspects of gene therapy production, such as measuring impurities, monitoring process development, and tracking viral vector stability across diverse platforms.

Bio-Rad’s Vericheck ddPCR Empty-Full Capsid Kit for serotypes 2,5,8 and 9 represents a new era in AAV characterization, delivering precise, consistent, and rapid data on capsid contents with minimal sample input and reliable performance across multiple sample types. As gene therapy developers navigate regulatory requirements and strive to bring safe, effective treatments to patients, the Vericheck Kit provides the tools needed to enhance quality control, optimize production, and drive the field of gene therapy forward.