Biology subjects

Dawson, E. D.

Publications and source records attributed to Dawson, E. D..

6 recordsLinked to original sources

bioRxiv2026

Single Platform Solution for Identity and Quantification of Influenza Hemagglutinin (HA) mRNA Constructs and Resulting Expressed Proteins for Application to Influenza mRNA Vaccines

The rapid response to the Coronavirus Disease (COVID-19) pandemic highlighted mRNA as an attractive modality for vaccines and therapeutics to address infectious diseases, cancer, and other diseases. The current analytical methods for identification and quantification of both the mRNA construct as well as the resulting expressed protein(s) relevant to mRNA vaccines are inherently singleplex and usually labor and time intensive, presenting a bottleneck as these vaccines become increasing multivalent. Here, we first present performance data from an approximately 2-hour DNA microarray assay developed specifically to provide identity and quantity measurements for the mRNA constructs present in an influenza virus hemagglutinin (HA)-encoding multivalent mRNA vaccine regardless of the specific strain or codon optimization strategy utilized. The assay functions on both naked and lipid nanoparticle (LNP)-encapsulated mRNAs without the requirement of an upfront mRNA extraction or amplification. Second, we show that a separate [~]2-hour multiplexed immunoassay executed on the same VaxArray Platform can be utilized to measure the expressed proteins produced from these influenza HA mRNAs post-transfection as an in vitro potency assay readout. Both assays provide the necessary specificity for each component present in a multivalent mixture and represent new tools in the analytical toolbox for multiplexed mRNA vaccine analytics.

molecular biology↗

bioRxiv2025

Assay for Rapid Quantification of Capped and Tailed Intact mRNA

The rapid development of mRNA vaccines since the COVID-19 pandemic has highlighted the potential of mRNA technology for vaccine and therapeutic applications. However, evaluating mRNA quality, including its integrity, 5' capping efficiency, and 3' poly(A) tailing, remains complex and challenging. Current analytical methods provide detailed insights, but are often limited by long analysis times, complex sample preparation, and the need for specialized expertise. The 5 CapQ assay addresses these challenges and provides a novel, rapid, and straightforward method that offers a single, unique measurement of intact mRNA with both a 5'cap and 3'poly(A) tail. This 2-hour benchtop-based assay provides quantitative or relative analysis, with high accuracy and precision without extensive sample processing. The 5 CapQ assay offers a significant advancement in mRNA analytics to alleviate bottlenecks in applications including in vitro transcription (IVT) reaction optimization, post-transcriptional capping optimization, and assessment of batch-to-batch IVT bioprocess consistency.

molecular biology↗

bioRxiv2022

30-Minute Highly Multiplexed VaxArray Immunoassay for Pneumococcal Vaccine Antigen Characterization

Pneumonia accounts for over 20% of deaths worldwide in children aged 1 to 5 years, disproportionately affecting lower- and middle-income countries. Effective, highly multivalent pneumococcal vaccines are available to decrease disease burden, with numerous new vaccines currently under development to serve a variety of worldwide markets. However, pneumococcal conjugate vaccines are among the hardest biologics to manufacture and characterize due to their complexity and heterogeneity. Current characterization methods are often inherently singleplex, requiring separate tests for each serotype present. In addition, identity and quantity are often determined with separate methods. We have developed the VaxArray Pneumococcal Assay for applications in identity, quantity, and stability testing of pneumococcal polysaccharide and pneumococcal conjugate vaccines. The VaxArray Pneumococcal Assay has a time to result of less than 30 minutes and is an off-the-shelf multiplexed, microarray-based immunoassay kit that can identify and simultaneously quantify 23 pneumococcal polysaccharide serotypes common to many on-market and in-development vaccines. Here, we highlight the potential of the assay for identity testing by showing high reactivity and serotype-specificity to a wide variety of native polysaccharides, CRM197-conjugated polysaccharides, and drug product. The assay also has vaccine-relevant lower limits of quantification in the low to mid ng/mL range and can be used for accurate quantification even in adjuvanted vaccines. Excellent correlation to the anthrone assay is demonstrated, with VaxArray resulting in significantly improved precision over this antiquated chemical method.

immunology↗

bioRxiv2022

Rapid Identity and Quantity CQA Test for Multivalent mRNA Drug Product Formulations

The COVID-19 pandemic highlighted mRNA as a promising platform for vaccines and therapeutics. Many of the analytical tools used to characterize the critical quality attributes of mRNA are inherently singleplex and are not necessarily optimal from a labor and cost perspective. Here we demonstrate feasibility of a multiplexed platform (VaxArray) for efficient identity verification and concentration determination for both monovalent and multivalent mRNA formulations. A model system comprised of mRNA constructs for influenza hemagglutinin and neuraminidase was used to characterize the analytical performance metrics for a VaxArray mRNA assay. The assay presented herein had a time to result of less than 2 hours, required no PCR-based amplification nor extraction of mRNA from lipid nanoparticles, and exhibited high construct specificity that enabled application to the bivalent mixture. The sensitivity for influenza hemagglutinin and neuraminidase mRNA was sub-{micro}g/mL, which is vaccine-relevant, and the average accuracy (%recovery) and precision were 104%{+/-}2%and 9%{+/-}2%, respectively.

bioengineering↗

bioRxiv2021

VaxArray Immunoassay for the Multiplexed Quantification of Poliovirus D-Antigen

Next generation poliovirus vaccines are critical to reaching global poliovirus eradication goals. Recent efforts have focused on creating inactivated vaccines using attenuated Sabin strains that maintain patient safety benefits and immunogenicity of conventional inactivated vaccines while increasing manufacturing safety and lowering production costs, and on developing novel oral vaccines using modified Sabin strains that provide critical mucosal immunity but are further attenuated to minimize risk of reversion to neurovirulence. In addition, there is a push to improve the analytical tools for poliovirus vaccine characterization. Conventional and Sabin inactivated poliovirus vaccines typically rely on standard plate-based ELISA as in vitro D-antigen potency assays in combination with WHO international standards as calibrants. While widely utilized, the current D-antigen ELISA assays have a long time to result (up to 72 hours), can suffer from lab-to-lab inconsistency due to non-standardized protocols and reagents, and are inherently singleplex. For D-antigen quantitation, we have developed the VaxArray Polio Assay Kit, a multiplexed, microarray-based immunoassay that uses poliovirus-specific human monoclonal antibodies currently under consideration as standardized reagents for characterizing inactivated Sabin and Salk vaccines. The VaxArray assay can simultaneously quantify all 3 poliovirus serotypes with a time to result of less than 3 hours. Here we demonstrate that the assay has limits of quantification suitable for both bioprocess samples and final vaccines, excellent reproducibility and precision, and improved accuracy over an analogous plate-based ELISA. The assay is suitable for adjuvanted combination vaccines, as common vaccine additives and crude matrices do not interfere with quantification, and is intended as a high throughput, standardized quantitation tool to aid inactivated poliovirus vaccine manufacturers in streamlining vaccine development and manufacturing, aiding the global polio eradication effort.

immunology↗

bioRxiv2021

Multiplexed VaxArray Immunoassay for Rapid Antigen Quantification in Measles and Rubella Vaccine Manufacturing

Measles-containing vaccines (MCV), specifically vaccines against measles and rubella (MR), are extremely effective and critical for the eradication of measles and rubella diseases. In developed countries, vaccination rates are high and vaccines are readily available, but continued high prevalence of both diseases in developing countries and surges in measles deaths in recent years have highlighted the need to expand vaccination efforts. To meet demand for additional vaccines at a globally affordable price, it is highly desirable to streamline vaccine production thereby reducing cost and speeding up time to delivery. MR vaccine characterization currently relies on the 50% cell culture infectious dose (CCID50) assay, an endpoint assay with low reproducibility that requires 10-14 days to complete. For streamlining bioprocess analysis and improving measurement precision relative to CCID50, we developed the VaxArray Measles and Rubella assay kit, which is based on a multiplexed microarray immunoassay with a 5-hour time to result. Here we demonstrate vaccine-relevant sensitivity ranging from 345 - 800 IFU/mL up to 100,000 IFU/mL and specificity that allows simultaneous analysis in bivalent vaccine samples. The assay is sensitive to antigen stability and has minimal interference from common vaccine additives. The assay exhibits high reproducibility and repeatability, with 15% CV, much lower than the typical 0.3 log10 error (~65%) observed for the CCID50 assay. The intact protein concentration measured by VaxArray is reasonably correlated to, but not equivalent to, CCID50 infectivity measurements for harvest samples. However, the measured protein concentration exhibits equivalency to CCID50 for more purified samples, including concentrated virus pools and monovalent bulks, making the assay a useful new tool for same-day analysis of vaccine samples for bioprocess development, optimization, and monitoring.

immunology↗