Synthesis of LLO-s-s PEI
1. Dissolve 10 mg PEI1.8 in 0.5 ml of methylene chloride.
2. Add 0.5 ml (N-Succinimidyl 3-(2-pyridyldithio)-propionate) (SPDP) in methylene chloride dropwise to the PEI solution.
3. React at room temperature for 6 h.
4. Remove methylene chloride under vacuum at room temperature.
5. Extract dried mixtures with 1 mL of phosphate buffer.
6. Filter through a 0.45 ÎŒm syringe filter.
7. Purify PEI pyridyldithiopropionate (PEI1.8-PDP) by size exclusion chromatography (Sephadex G10, 1Ã35 cm) with deionized water.
8. Determine PEI content using Cu2+ against PEI1.8 standard at 630 nm.
9. Determine the number of PDP groups per average chain of PEI.
9.1. Treatment of PEI1.8-PDP at 50 mM DTT
9.2. Purify thiolated PEI using Sephadex G10.
9.3. Determine pyridine-2-thione at 343 nm (molar absorptivity=8080 cmâ1 Mâ1)