🔬 𝐀𝐝𝐯𝐚𝐧𝐜𝐢𝐧𝐠 𝐎𝐥𝐢𝐠𝐨𝐧𝐮𝐜𝐥𝐞𝐨𝐭𝐢𝐝𝐞 𝐒𝐞𝐪𝐮𝐞𝐧𝐜𝐢𝐧𝐠 𝐰𝐢𝐭𝐡 𝐈𝐨𝐧 𝐌𝐨𝐛𝐢𝐥𝐢𝐭𝐲–𝐌𝐚𝐬𝐬 𝐒𝐩𝐞𝐜𝐭𝐫𝐨𝐦𝐞𝐭𝐫𝐲 🧬
As therapeutic oligonucleotides continue to gain traction in the treatment of rare and chronic diseases, the need for accurate, high-throughput sequence confirmation has never been greater. Traditional LC-MS/MS approaches often require multiple injections and are limited by charge-state-specific fragmentation and low throughput.
📈 In this study, a 𝐦𝐮𝐥𝐭𝐢𝐝𝐢𝐦𝐞𝐧𝐬𝐢𝐨𝐧𝐚𝐥 𝐋𝐂-𝐈𝐌𝐒-𝐂𝐈𝐃-𝐌𝐒 platform was developed to tackle these challenges. By integrating 𝐢𝐨𝐧 𝐦𝐨𝐛𝐢𝐥𝐢𝐭𝐲 𝐬𝐩𝐞𝐜𝐭𝐫𝐨𝐦𝐞𝐭𝐫𝐲, 𝐝𝐫𝐢𝐟𝐭 𝐭𝐢𝐦𝐞–𝐝𝐞𝐩𝐞𝐧𝐝𝐞𝐧𝐭 𝐜𝐨𝐥𝐥𝐢𝐬𝐢𝐨𝐧-𝐢𝐧𝐝𝐮𝐜𝐞𝐝 𝐝𝐢𝐬𝐬𝐨𝐜𝐢𝐚𝐭𝐢𝐨𝐧, and 𝐦𝐚𝐬𝐬 𝐬𝐩𝐞𝐜𝐭𝐫𝐨𝐦𝐞𝐭𝐫𝐲, the authors enabled:
✅ Separation and analysis of multiple charge states in a single injection
✅ Application of optimized, charge-specific collision energies using drift time-dependent voltage ramps
✅ Sequence coverage up to 85% for 21-mer DNA/RNA sequences
✅ A novel data preprocessing pipeline coupling the PNNL PreProcessor with Agilent BioConfirm, enabling streamlined all-ion fragmentation data interpretation
🧪 The method was validated across diverse oligonucleotide types—including phosphorothioate-modified strands and sequences up to 40-mers—demonstrating adaptability and robustness. While sequence coverage decreased with increasing strand length, the workflow proved particularly well-suited to the therapeutic range (15–30 bases).
📊 This represents a promising direction toward 𝐞𝐟𝐟𝐢𝐜𝐢𝐞𝐧𝐭 𝐐𝐂 𝐰𝐨𝐫𝐤𝐟𝐥𝐨𝐰𝐬 for synthetic oligonucleotides, including those with chemically modified backbones.
🎯 𝐊𝐞𝐲 𝐓𝐚𝐤𝐞-𝐀𝐰𝐚𝐲𝐬:
• LC-IMS-CID-MS enables single-injection, multi-charge-state oligonucleotide analysis
• Drift time–dependent CE ramps optimize fragmentation across species
• New data workflow supports high-throughput sequence confirmation
• Effective for therapeutic-length oligos, with potential to expand via other dissociation methods
🔗 𝐅𝐮𝐥𝐥 𝐩𝐚𝐩𝐞𝐫 available here (behind paywall 💰): https://lnkd.in/emMg7Wix
#Oligonucleotides #MassSpectrometry #IonMobility #AnalyticalChemistry #Biopharma #NucleicAcidTherapeutics #DrugDevelopment #LCMS #Bioanalysis #PharmaceuticalScience
Jack Ryan, PhD, Gordon Slysz, Peter Rye, Sarah Stow, James Dodds, John Sausen & Erin Baker