Gene therapy is a medical technology that aims to produce a therapeutic effect by manipulating gene replacement, gene suppression, gene supplementation, or gene editing. One of the keys to the development of gene therapy is delivering genetic constructs to target cells. Various non-viral vectors such as plasmids and viral vectors are developed for the delivery of oligonucleotides (such as ASO, siRNA, miRNA, and aptamers), mRNA, and plasmid DNA (pDNA). Plasmids are common non-viral vectors with topologies that include supercoiled, open-circle, and linear forms. Compared to open-circular or linear plasmids, supercoiled plasmids have more compact structure and are smaller in size, which enables them more easily penetrate the cell membrane into the cytoplasm with reduced susceptibility to degradation and less likely to induce immune responses. Given the impact of plasmid topology on transfection efficiency, stability, immunogenicity, production process, and safety, it is necessary to establish accurate analytical methods for plasmid purification and characterization to comply with regulatory requirements.
High performance liquid chromatography (HPLC) is a common method for the analysis of key quality attributes of oligonucleotides, nucleic acids, plasmids, and viral vectors. This method has the characteristics of selectivity, sensitivity, throughput, and repeatability to ensure the efficacy and safety of gene therapeutics. In the HPLC techniques, anion-exchange chromatography is commonly used to analyze plasmid topological morphology and adeno-associated virus (AAV) capsid content. Size exclusion chromatography can be used to characterize the aggregates in nucleic acids, plasmids, and AAVs. Ion-pair reversed phase chromatography is widely used to determine the purity of oligonucleotides.
Amerigo Scientific offers a series of DNACore liquid chromatography columns specially designed for the high-efficiency separation of oligonucleotides, nucleic acids, plasmids, and AAVs. These columns are manufactured based on the advanced monodispersed particle technology and column chemistry, and feature high resolution, ideal selectivity, good stability, and excellent consistency.
Each DNACore column is manufactured by reliable packing methods and undergoes well-designed chromatography tests to ensure quality and consistency. Each batch of separation media is produced in accordance with a strict quality management system and tested with relevant biological molecules to ensure separation performance and batch-to-batch consistency. A certificate of assurance for the separation media and a quality assurance report for the column will be shipped with each DNACore column.
IEX Columns
DNACore anion-exchange columns are designed for analysis of oligonucleotides, DNAs, mRNAs and plasmid topology. These columns have ideal selectivity for plasmids of different topologies, high column efficiency and mechanical strength, excellent tolerance to acids, bases and organic solvents, and good column-to-column consistency.
Figure 1. Analysis of Plasmids Using DNACore NP-Q and NP-T Columns
SEC Columns
DNACore size exclusion chromatography columns are designed for separating plasmids, circle RNAs, retroviruses, adenoviruses, lentiviruses, and related aggregates and fragments. Based on advanced column chemistry, the undesired interactions between the stationary phase and analytes are minimized, resulting in good peak shape and high recovery.
Monodispersed, high pore-volume, spherical silica particles
3μm
1000Å/2000Å
3000 psi
40°C
2-8
Applications
Figure 2. Analysis of circRNA Using DNACore SEC-1000 Columns
Figure 3. Analysis of Plasmids Using DNACore SEC-1000/2000 Columns
RP Columns
DNACore reversed-phase columns are designed for analysis of oligonucleotides, DNA, mRNA, plasmids, etc. These columns have ideal selectivity for oligonucleotides of different sizes, high column efficiency and mechanical strength, and low column bleed for MS compatibility.
Figure 4. Analysis of Oligonucleotides Using DNACore 1000 C18 Columns
Figure 5. Analysis of RNAs Using DNACore 1000 C18 Columns
AAV SEC Columns
Adeno-associated virus (AAV) is widely used vector for gene therapy because of its non-pathogenicity, low immunogenicity and long-term gene expression. In the process of AAV production, viral titer, capsid content, and aggregation are key factors affecting the purity, efficacy, and safety of AAV-based gene therapy products.
Amerigo Scientific provides DNACore size exclusion chromatography columns for the aggregate analysis of AAVs in different serotypes. Innovative particle technology enables columns to achieve high monodisperse efficiency and long mechanical strength, high resolution.
Figure 1. Analysis of Plasmids Using DNACore NP-Q and NP-T Columns
Figure 2. Analysis of circRNA Using DNACore SEC-1000 Columns
Figure 3. Analysis of Plasmids Using DNACore SEC-1000/2000 Columns
Figure 4. Analysis of Oligonucleotides Using DNACore 1000 C18 Columns
Figure 5. Analysis of RNAs Using DNACore 1000 C18 Columns
Figure 6. Analysis of AAV Aggregates in Different Serotypes Using DNACore AAV-SEC Column
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