A Simple Method for the Design and Development of Flavivirus NS1 Recombinant Proteins Using an In Silico Approach

Figure 4

SDS polyacrylamide gel electrophoresis (PAGE) analysis showing the example of Ni-NTA purification of flavivirus NS1 recombinant protein by the Escherichia coli expression system (a), western blot analysis of DENV NS1 (b), and western blot analysis of ZIKV NS1 recombinant proteins (c). (a) Each fraction was loaded as follows: 1, E. coli lysate expressing DENV 1 NS1 recombinant protein after IPTG induction; 2, fraction (unbound) through-out in Ni-NTA column; 3, washing fraction at 50 mM imidazole; 4, elute 1 fraction at 150 mM imidazole; 5, elute 2 fraction at 150 mM imidazole; 6, E. coli lysate expressing DENV type 2 NS1 recombinant protein; 7, fraction (unbound) through-out in Ni-NTA column; 8, washing fraction; 9, elute 1 fraction; 10, elute 2 fraction, M: PageRuler protein ladder (Abcam plc., Cambridge, UK). (b) Anti-DENV NS1 primary antibody (titer 1 : 2,000) and anti-mouse IgG secondary antibody (1 : 20,000) were used. 1–4; DENV type 1 NS1 recombinant protein, 5-6; DENV type 2 NS1 recombinant protein, 7-8; DENV type 3 and 4 NS1 recombinant protein, 9–12; DENV type 1–4 NS1 recombinant protein (Native Antigen Company), 13; JEV-NS1 (Native Antigen Company), 14; TBEV NS1 (Native Antigen Company), 15; WNV-NS1 (Native Antigen Company), 16; YFV-NS1 (Native Antigen Company), 17; ZIKV Uganda strain NS1 (Native Antigen Company), 18; ZIKV Suriname strain NS1 (Native Antigen Company). (c) Anti-ZIKV NS1 primary antibody (titer 1 : 2,000) and anti-mouse IgG secondary antibody (1 : 20,000) were used. 1; WNV NS1 recombinant protein, 2; YFV NS1 recombinant protein, 3; ZIKV NS1 recombinant protein, 4; JEV NS1 recombinant protein, 5; DENV NS1 recombinant protein, 6; ZIKV Uganda strain NS1 (Native Antigen Company), 7; DENV type 1 NS1 (Native Antigen Company), 8; DENV type 2 (Native Antigen Company), 9; DENV type 3 (Native Antigen Company), 10; DENV type 4 (Native Antigen Company), M; PageRuler protein ladder (Abcam).

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