Lentivirus titer and structure
Being able to quickly and comprehensively monitor the quality of your lentiviral prep from crude sample all the way through your purification process is key for rapid process development. Current methods are limited in their ability to handle crude viral sample, making it impossible to get an accurate lentiviral titer, monitor viral stability, and measure non-viral contaminants in early stage samples.
The most common method for measuring physical lentivirus titer is a p24 ELISA assay. Since p24 protein is relatively abundant in free solution, this assay is seriously limited because it can't tell the difference between p24 used in lentivirus capsids and soluble p24 not associated with a virus. This results in inaccurate lentiviral titers when the concentration of p24 protein doesn't scale along with the quantity of the virus - especially a problem in crude samples with high concentrations of soluble p24.
When it comes to detecting contaminants, ELISAs also can't give any info on lentivirus aggregation and the high specificity of ELISAs leave you in the dark about any empty viral particles that don't contain p24. All this means the p24 ELISA falls short of delivering comprehensive answers on the composition and integrity of your viral prep, and the effectiveness of your lentivirus production and purification process.
The right tool for the job
Then Leprechaun uses interferometry to size up captured particles one-by-one, between 35 and 200 nm diameter. Finally, immunofluorescence microscopy shows off exactly which particles are the lentiviruses you’re after – the ones that are the right size and have VSVG and p24 present. Anything else that doesn't have the right combo of size and detected protein is a damaged, incorrectly assembled lentivirus, an EV, or contaminants like soluble p24 and EVs.
Any time you check on your lentivirus with Leprechaun, from crude samples at early stages of production to your purified virus ready to use for CAR-T manufacture, contaminants will stick out as the wrong size or by having the wrong structural components. Tracking lentivirus titer and all kinds of contaminants throughout your process paints the full picture of lentivirus stability and gives you the readout you need to optimize your lentivirus production and purification.
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