Bovine carbonic anhydrase cab

The phospholipids we used were 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC, MW = 734.04), 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC, MW = 790.15). Besides, we used DPPC incorporating cholesterol (MW = 386.65) (DPPC:cholesterol = 66 mol%:33 mol%). DPPC, DSPC, and cholesterol were purchased from Avanti Polar Lipids (Alabaster, AL, USA). Target proteins we used were bovine carbonic anhydrase (CAB, MW = 28,400) and lysozyme (MW = 17,307), which were purchased from Sigma Aldrich (St. Louis, MO, USA). The fluorescent molecule encapsulated in liposomes was calcein (MW = 622.53), which was also purchased from Sigma Aldrich. Silpot184 and Silpot184 CAT (hardener) purchased from Toray Dow Corning (Tokyo, Japan) were used for the fabrication of polydimethylsiloxane (PDMS)-based chips. The gel beads, Sepharose 4B, which are used for filtering out the free calcein during liposome preparation, were purchased from GE Healthcare (Uppsala, Sweden). The filtration buffer, phosphate buffered saline (PBS), were purchased from Thermo Fisher Scientific (Yokohama, Japan). All the chemical reagents were of analytical grade.

1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC) and egg yolk phosphatidyl-ethanolamine (EPE) were purchased from Avanti Polar Lipids (Alabaster, AL, USA). Chitosan-hexamer was purchased from Seikagaku Co. (Tokyo, Japan). Bovine carbonic anhydrase (CAB) and 16-mercaptohexadecanoic acid were obtained from Sigma (St. Louis, MO, USA). Hen egg white lysozyme, cholesterol, guanidinium hydrochloride (GuHCl), dithiothreitol (DTT), N-hydroxysuccinimide (NHS), dioxane and potassium hexacyano-ferrate (II) were purchased from Wako (Osaka, Japan). 1-ethyl-3-(3-dimethlaminopropyl)-carbodiimide hydrochloride (WSC) and 1-(4-trimethyl -ammoniumphenyl)-6-diphenyl-1,3,5-hexatriene (TMA-DPH) were purchased from Dojindo Laboratories (Kumamoto, Japan). All other chemicals used were of analytical reagent grade.