Purification of Prototype Foamy Virus Intasomes by Size Exclusion Chromatography

Begin with a suspension containing precipitates of prototype foamy virus or PFV intasomes, monomeric integrases, DNA oligomers, and protein aggregates.

PFV intasomes are tetramers of viral integrases complexed with synthetic oligomers that mimic viral genomic DNA.

Add a salt solution to the suspension and incubate on ice. The salt reduces the electrostatic interaction between the intasomes, enabling the precipitates to solubilize.

Pipette gently at regular intervals to promote intasome solubilization.

Centrifuge to pellet remaining precipitate and collect the supernatant containing soluble intasomes.

Take a pre-equilibrated size-exclusion chromatography column packed with cross-linked agarose beads and load the supernatant onto the column.

Due to their large size, protein aggregates and intasomes bypass the bead pores and elute earlier.

Smaller monomeric integrases and oligomers enter the bead pores and elute later.

Collect the early-eluting fractions containing intasomes for further downstream analysis.

Prepare two wide-bore pipette tips by removing two to four milliliters from the end of a 200 microliter tip with a new razor or scalpel blade on a clean surface. Remove the dialysis tubing