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sgRNA and Cas9 mRNA Characterization by Ion PairReversed Phase Liquid Chromatography Coupled to HRMS
With the rise of RNA vaccines and CRISPR-based therapies, the demand for robust analytical methods to characterize long RNAs like sgRNA and mRNA has grown significantly. In this application note, we demonstrate a comprehensive workflow combining Ion Pair-Reversed Phase (IP-RP) chromatography and high-resolution mass spectrometry (HRMS) using Biozen Oligo and WidePore C4 columns. Model systems (BRCA1-targeting sgRNA and Cas9-mRNA) were analyzed both intact and post-RNase 4 digestion. This approach enabled full sequence identification, impurity profiling, and resolution of phosphorothioate diastereomers. For Cas9-mRNA, the workflow also confirmed 5' cap integrity and poly-A tail heterogeneity. This streamlined strategy offers a powerful solution for RNA therapeutic characterization with high sensitivity and resolution.
sgRNA and Cas9 mRNA Characterization by Ion PairReversed Phase Liquid Chromatography Coupled to HRMS
With the rise of RNA vaccines and CRISPR-based therapies, the demand for robust analytical methods to characterize long RNAs like sgRNA and mRNA has grown significantly. In this application note, we demonstrate a comprehensive workflow combining Ion Pair-Reversed Phase (IP-RP) chromatography and high-resolution mass spectrometry (HRMS) using Biozen Oligo and WidePore C4 columns. Model systems (BRCA1-targeting sgRNA and Cas9-mRNA) were analyzed both intact and post-RNase 4 digestion. This approach enabled full sequence identification, impurity profiling, and resolution of phosphorothioate diastereomers. For Cas9-mRNA, the workflow also confirmed 5' cap integrity and poly-A tail heterogeneity. This streamlined strategy offers a powerful solution for RNA therapeutic characterization with high sensitivity and resolution.
Document Type:
Technical Notes
Target Industries:
Techniques:
Brands:
Separation Modes:
Reversed Phase
Phases:
sgRNA and Cas9 mRNA Characterization by Ion PairReversed Phase Liquid Chromatography Coupled to HRMS
With the rise of RNA vaccines and CRISPR-based therapies, the demand for robust analytical methods to characterize long RNAs like sgRNA and mRNA has grown significantly. In this application note, we demonstrate a comprehensive workflow combining Ion Pair-Reversed Phase (IP-RP) chromatography and high-resolution mass spectrometry (HRMS) using Biozen Oligo and WidePore C4 columns. Model systems (BRCA1-targeting sgRNA and Cas9-mRNA) were analyzed both intact and post-RNase 4 digestion. This approach enabled full sequence identification, impurity profiling, and resolution of phosphorothioate diastereomers. For Cas9-mRNA, the workflow also confirmed 5' cap integrity and poly-A tail heterogeneity. This streamlined strategy offers a powerful solution for RNA therapeutic characterization with high sensitivity and resolution.
| Target Industries: | Biopharmaceutical Analysis |
| Techniques: | HPLC Reversed Phase, Oligonucleotide Analysis |
| Brands: | Biozen LC |
| Separation Modes: | Reversed Phase |
| Phases: | Biozen-Oligo, Biozen WidePore C4 |
Phenomenex is a technology leader committed to developing novel analytical chemistry solutions that solve the separation and purification challenges of researchers worldwide
Phenomenex is a technology leader committed to developing novel analytical chemistry solutions that solve the separation and purification challenges of researchers worldwide