Advantages of Sequential or Bracketed Injection Methods to Improve the Chromatographic Analysis of Biotherapeutics

High-performance liquid chromatography (HPLC) analysis of biotherapeutics introduces new challenges because large molecules may require distinct injection modes or sample handling protocols to improve the analysis. Some challenges can be addressed by using autosampler methods that allow an analyst to program specific injection sequences. This article explores three case studies where autosampler injection programs help to (i) reduce the carry-over observed for large nucleic acids during anion-exchange chromatography (AEX), (ii) reduce peak distortion and breakthrough for monoclonal antibodies (mAbs) during hydrophilic-interaction chromatography (HILIC), and (iii) facilitate dissolution of lipid nanoparticles (LNPs) so that a size-exclusion chromatography (SEC) method can be applied for ribonucleic acid (RNA) payload analysis. The appropriate use of autosampler capabilities offers unique benefits to analyze biotherapeutics.