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The Impact of Digital PCR on Quantification in Biopharma Research and Manufacturing

The Impact of Digital PCR on Quantification in Biopharma Research and Manufacturing
Courtesy of Niba Labs

During an online conference held by Stilla, Assis. Prof. Dr. David Dobnik discussed the history of PCR technologies and the proprietary multiplex dPCR assay platform NIBAplex.

Niba Labs, founded in 2021, is a spin-out of The National Institute of Biology (NIB) in Slovenia which built a reputation as a leading laboratory for cutting-edge expertise in viral characterization and absolute quantification. Since 2016, experts from the institute have been engaged in digital PCR gene therapy analytics. 

In a conference held by Stilla on Tuesday, April 23, entitled “The Impact of Digital PCR on Quantification in Biopharma Research and Manufacturing”; Assis. Prof. Dr. David Dobnik offered insights into the history of PCR, advances in digital PCR, and details about Niba Labs as experts for digital PCR (dPCR, including ddPCR and ndPCR) assay development and testing.

“We have a mission with NIBA Labs to improve the speed and efficiency of process development in the segment of cell and gene therapies by providing expert support in development and analytics. We have labs that are located at Novartis Industrial Campus in Slovenia,” Dobnik stated at the beginning of the conference.

Having earned a PhD in Biotechnology at the Biotechnical Faculty, University of Ljubljana, Slovenia, in 2012; David Dobnik has followed the evolution of PCR technology closely, notably regarding the development of digital PCR since its conception in 1992—much has changed.

PCR Technologies: Evolution and Performance Today

PCR technologies have evolved significantly, from the groundbreaking classical PCR technique of the 1980s to the introduction of real-time PCR and, eventually, digital PCR (dPCR) in the early 1990s. Real-time PCR allowed monitoring of amplification in real-time through fluorescence, laying the groundwork for more precise methodologies. However, it was digital PCR that truly revolutionized molecular quantification by enabling absolute quantification of targets through microfluidics-based partitioning of reaction mixtures. Despite initial skepticism, dPCR’s superior accuracy, precision, and tolerance to inhibitors made it indispensable, particularly in fields like oncology and gene therapy.

“A paper we’ve written shows why people choose digital PCR. One example is due to the lower variability in the results received via qPCR. Now, projections foresee a 10 times increase in the digital PCR market size in the next decade.”

Courtesy of Niba Labs

Understanding Digital PCR and the Role of Multiplex dPCR in Gene Therapy

Digital PCR (dPCR) offers precise and absolute quantification of nucleic acid targets by partitioning the reaction mixture into discrete compartments, each containing a single or few target molecules. Unlike traditional PCR methods, dPCR’s end-point approach enables digitalized signal detection, making it highly versatile in multiplexing for the simultaneous quantification of multiple targets within a single reaction. This capability, coupled with dPCR’s superior performance in multiplex quantification compared to traditional qPCR methods, has made it increasingly attractive for various applications.

Multiplex dPCR plays a crucial role in gene therapy, particularly in characterizing viral vectors. By employing multiplex dPCR assays designed to assess vector genome integrity, researchers gain insights into viral capsid composition, enabling informed decisions in process optimization. This leads to better-characterized products and potentially lower production costs per dose. Accurate quantification of different fragments of vector genomes provides a comprehensive understanding of viral populations, contributing to safer and more effective therapies.

Courtesy of Niba Labs

READ more about Nio+ by Stilla in our article here.

The NIBAplex Platform and Stilla’s Nio+ Digital PCR System

The new NIBAplex platform, developed by Niba Labs, is a multiplex dPCR assay system for precise nucleic acid target quantification. Leveraging dPCR capabilities, NIBAplex offers absolute quantification of multiple targets within a single reaction, facilitating comprehensive nucleic acid sample analysis. The platform, customizable to specific research needs, integrates seamlessly with Stilla’s Nio+ digital PCR system, known for its high throughput and advanced multiplexing capabilities.

The workflow involves preparing the dPCR reaction mixture with the nucleic acid sample and specific NIBAplex assays and loading it onto the Stilla Nio+ system for partitioning into individual compartments through microfluidic technology. After PCR cycling and endpoint analysis, researchers obtain absolute quantification of multiple targets simultaneously, enhancing viral vector and nucleic acid sample characterization for informed decision-making.

As dPCR continues to evolve and find applications across various fields, its role as a gold standard for nucleic acid quantification solidifies, driving advancements in research, diagnostics, and therapeutics.

Courtesy of Niba Labs
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