HSPC/Chol/DSPE-mPEG2000/DSPE-mPEG2000-maleimide/MPL (49/45/4/1/1, mol/mol) Liposomes

The maleimide-thiol reaction is a click chemistry technique useful for the selective bioconjugation of molecules containing sulfhydryl groups to a secondary molecule containing maleimide moieties. Thiols are most notably found in nature via the amino acid residue cysteine; thus, maleimide-thiol coupling is frequently employed for bioconjugation of cysteine-containing biomolecules such as proteins and peptides. Moreover, synthetic oligonucleotides can be engineered to contain thiol groups, enabling maleimide coupling to nucleic acids [1].

These liposomes are prepared containing 1 mol% of DSPE-mPEG2000-maleimide, in an optimal buffer pH of 6.8. The reaction is relatively straight forward: maleimide- and thiol- containing compounds are allowed to react overnight at room temperature or 4oC. It is common to exclude oxygen from the system by flushing with an inert gas such as nitrogen or by applying vacuum. Additonally, as thiols have the tendency to form disulfide bonds, which will not react with maleimides, a reducing agent such as tris(2-carboxyethyl)phosphine (TCEP) is generally included [2]. A saturated TCEP solution is available as an add-on to your order if desired. After conjugation, the external buffer may be exchanged to remove TCEP by using dialysis or a centrifugal filtration device.

These liposomes contain 1 mol% MPL, which may be useful as an adjuvant for stimulating immune responses against the antigen to be conjugated by maleimide-thiol coupling. MPL is a less toxic analog of the bacterial lipopolysaccaride (LPS) derived from the Salmonella enterica serotype minnesota Re 595. MPL elicits Th1 responses and is reported to induce activation of T cells, B cells, and macrophages [3].