PREPARATION OF KODED SURFACES
Simply mix 1 part of cells/virions/particles/bacterium with one part of an FSL solution (containing 1 or more FSLs) and incubate for 10-120 minutes at 37°C (or at temperatures as low as 4°C). The constructs will spontaneously incorporate into the membrane or onto the surface and no further steps are required.
Kodecytes can also be created in vivo by injection of constructs directly into the circulation. However this process will modify all cells in contact with the constructs and usually require significantly more construct than in vitro preparation, as FSL constructs will preferentially associate with free lipids.
To facilitate description of FSL construct modified surfaces to be accurately described a series of Kode™ related terms have been adopted. These include:
- Kodecyte: A cell modified (koded) with an FSL construct.
- Kodevirion: A koded virus
- Kodesome: A koded liposome
- Koded: A cell, virus or surface (membrane), which has a coating of FSL constructs
- Koding: The process of contacting a surface/ membrane with an FSL construct
Some FSL constructs can be easily created in your own lab using FSLs bearing Reactive Functional Groups. One such product is FSL-RFG(Maleimide) which allows users to create peptides/proteins FSLs ready to use. The process is extremely simple and users can create an FSL-peptide overnight and ready for immediate use without purification.
FSL MODIFIED LIPID MEMBRANES
After labeling of the surface with the selected F bioactive(s) the constructs will be present and orientated at the membrane surface. It is expected that the FSL will be highly mobile within the membrane and the choice of lipid tail will effect is relative partitioning within the membrane. The construct unless it has flip-flop sequences is expected to remain surface presented. However, the modification is not permanent in living cells and constructs will be lost (consumed) at a rate proportional to the activity at the membrane and division rate of the cell (with dead cells remaining highly labeled). Additionally when present in vivo with serum lipids FSLs will elute from the membrane at a rate of about 1% per hour. In fixed cells or inactive cells (e.g. red cells) stored in serum free media the constructs are retained normally.
Liposomes are easy koded by simply adding FSL constructs into the preparation. Contacting kodesomes with microplates or other surfaces can cause the labeling of the surface.