Real-time monitoring of attenuated cytomegalovirus using Raman spectroscopy allows non-destructive characterization...

Real-time monitoring of viral particles can have a crucial impact on vaccine manufacturing and can alleviate public health challenges by supporting continuous supply. Spectroscopic methods such as Raman spectroscopy can provide rapid and non-invasive measurements. Here, we have developed a Raman spectroscopy-based tool to monitor the quality and quantity of viral particles in a continuous flow setup. We characterized the attenuated human cytomegalovirus (CMV) across a wide range of concentrations (1.45 × 10¹⁰ to 2.90 × 10¹¹ particles/mL) and flow rates (100 μm/s to 1000 μm/s) within a square quartz capillary. This process analytical technology (PAT) tool enables the detection of viral particles even at high flow rates such as 1000 μm/s. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and dynamic light scattering (DLS) demonstrated that the samples maintain their integrity even after laser exposure, reiterating the non-invasive nature of Raman spectroscopy. To the best of our knowledge, this is the first report on characterizing CMV particles using Raman spectroscopy, especially under flow conditions. We have also demonstrated the limit of detection (LOD_min) (2.01 × 10¹⁰ particles/mL) for CMV particles in continuous flow (1000 μm/s) (via the Raman spectroscopy method), addressing the effects of flow rate, concentration, and sample integrity. This technology could enable process control in the bio-manufacturing of vaccines.