An AAV ELISA measures assembled capsid, full or empty, by binding an antibody raised against the intact particle. It is quick, it needs no nucleic acid extraction, and it answers a question no genome assay answers: how much protein shell is in the preparation. It also does not tell you how much of that shell contains a genome, or how much of it will deliver one, and most of the confusion around AAV titres comes from reading one of those numbers as though it were another.
Capsid titre and genome titre are different numbers
A capsid ELISA counts particles by their shell. A genome assay, by quantitative PCR or droplet digital PCR, counts copies of the packaged sequence. The ratio between them is a property of the preparation, not a constant, and it is one of the more informative things you can know about a process, because it says how much of what you made is empty. An infectivity or transduction assay measures a third thing again. A preparation described by a single titre is a preparation whose empty fraction somebody has not measured, and the honest specification quotes at least the first two numbers with the method for each.
Serotype specificity decides which kit works
The antibodies in these kits are raised against conformational features of a particular capsid, so a kit for one serotype may read a related one weakly, differently, or not at all, and an engineered or chimeric capsid may fall outside every catalogue kit. That is the first question to ask, before sensitivity or price: has this kit been shown to bind this capsid, and with what standard. A kit used outside its validated serotype produces a number, which is worse than producing nothing.
Where the empty capsid problem shows up
Because the ELISA cannot distinguish a full particle from an empty one, a process change that improves packaging can look like it did nothing, and one that produces more empties can look like it improved yield. Analytical ultracentrifugation, charge-based separation or mass photometry are the methods that resolve the population; the ELISA's contribution is the denominator. Use it that way, and pair every capsid reading with the genome reading from the same sample rather than from a neighbouring run.
Validating the kit against your own process
Run the kit against your own material across the range you actually see, including a deliberately diluted and a deliberately concentrated sample, and check linearity and recovery rather than trusting the kit's own curve. Ask what the reference standard is and whether it is the same serotype. Watch for matrix effects from your formulation buffer, surfactant or excipients, which are a common cause of a titre that moves when the process did not. For material heading into a regulated study, the expectations for analytical methods sit with the therapy's own manufacturing and control requirements.
Questions people ask about aav elisa
Does an AAV ELISA measure infectious virus?
No. It measures assembled capsid whether or not it contains a genome and whether or not it can transduce a cell. Infectivity needs a cell-based assay and correlates only loosely with either titre.
Which AAV ELISA kit fits my serotype?
Only one shown to bind your capsid with a standard of the same serotype. Engineered and chimeric capsids frequently fall outside every catalogue kit, and the honest answer there is a custom assay rather than the closest available.
Why does my capsid to genome ratio look wrong?
Most often the two numbers came from different samples, different dilutions or different laboratories. Measure both on the same aliquot before concluding anything about the empty fraction.