AAV – purity and integrity

Monitoring each step in the purification of AAV

Undesired AAV sample characteristics to consider when optimizing the downstream process

  • Aggregation
  • Failure to remove host-cell debris or residual DNA
  • Loss of particle morphology or integrity

ALl these types of contaminants can easily be identified and quantified by using the automated image analysis software tools in MiniTEM

  After 1st purification step

After 1st purification step

After 2nd purification step

After 2nd purification step

After final purification step

After final purification step

Types of contaminants that may appear

Large size impurities

Large size impurities

Small size impurities

Small size impurities

Aggregates

Aggregates

Residual DNA

Residual DNA

Automated MiniTEM analysis of purity and level of debris

MiniTEM automatically images, detects and classifies particles and debris. Since automation enables detection and analysis of a far larger number of particles than is feasible with manual operation, the results are more consistent, statistically significant and reproducible.

Undetected particles

Undetected particles

Detected primary particles and debris

Detected primary particles and debris

   Quantified  particles

Quantified particles

Time saving through automated MIniTEM analysis

Time saving through automated MIniTEM analysis

Capsid integrity analysis with MiniTEM

The proportion of broken AAV capsids in a sample can be measured using MiniTEM as negative stained samples of AAV particles display variations in staining patterns correlated to  capsid integrity status. 

Capsid integrity analysis with MiniTEM 1

Capsid integrity analysis with MiniTEM 2

Fig small top

Intact AAV particles appear as lacking internal staining as opposed to broken particles that exhibit internal dark staining due to disrupted virus capsid structures.