What media can I use to form transfection complexes?

As biologists know, any and all components of an experiment can affect successful outcomes. Today we’ll discuss a sometimes overlooked variable of the transfection workflow: complex formation solution.

The complex formation solution serves as the vehicle where nucleic acids and TransIT® reagents coalesce into transfection complexes. This first step in any transfection application can set the stage for success of the downstream experiment. For Mirus reagents, there are a few important considerations to keep at top of mind for complexation to occur correctly.

1. Beware ‘helpful’ additives
   Many media that are classified as ‘serum-free’ have components that are detrimental to transfection. Dextran sulfate is a common media additive, but other components with a strong positive charge in media can also be problematic and interfere with the condensation of nucleic acid and transfection reagent.
2. Serum-free is the way to be
   Serum contains negatively charged proteins, which can inhibit complex formation, and nucleases, which can degrade nucleic acid cargo prior to being protected by the transfection reagent. After the complex formation step, the transfection complex may be added to cells growing in the presence of serum.
3. Keep it in neutral
   The pH of the complex formation solution can directly affect the dynamics of complex formation. Compare complex growth in Opti-MEM® at pH 7.4 versus pH 8.5 in the figure below, which displays data collected from dynamic light scattering (DLS).
4. Saline is serene
   Transfection complexes consisting of multiple copies of nucleic acid cargo must aggregate to a certain size to achieve effective transfection. When utilizing cationic lipid and polymer transfection reagents in the absence of salt, this aggregation of complexes does not occur (see Water and HEPES conditions, Figure below).

Growth of Transfection Complexes in Various Complex Formation Media. Transfection complexes were formed in the indicated media. The size of the transfection complex was measured over time with DLS.

Mirus recommends forming transfection complexes in buffered, saline solutions such as Opti-MEM® or, in some cases, PBS or other serum-free basal media. Here is a list of our favorites:

• Opti-MEM® I Reduced-Serum Medium
• PBS 
• CHOgro® Complex Formation Solution
• RPMI 1640
• DMEM
• Ham’s F12
• MEM

NOTE: ‘complex formation solution’ and ‘cell culture media’ serve different purposes. The tips above apply to the complex formation solution, which is required solely for the unity of the nucleic acid and transfection reagent; therefore, the complex formation solution should not include additives that can interfere with this process. On the other hand, cell culture media, which provides the cells with the proper environment to support their health and growth can contain nutrients and additives such as serum and antibiotics. TransIT® Transfection Reagents are low toxicity, serum-compatible reagents, which eliminates the need for cell culture media changes.

Have a question about complex formation solutions or other transfection components? Let us know by email (techsupport@mirusbio.com)!