Novex 4 12 tris glycine gradient gel

Example 13

HCoV EMC and SARS CoV S1 Fc proteins (2.5 μg) were mock incubated or incubated with 12.5 μg soluble DPP IV (sDPP IV) or soluble ACE2 (sACE2) in a total volume of 100 μl PBS. Precipitates were washed thrice with lysis buffer and once with PBS, and subjected to a NOVEX® 4 12% Tris Glycine gradient gel (Invitrogen) under non reducing conditions. Results are shown in FIG. 22.

Example 10

The immunoprecipitation protocol was essentially carried out as described before with some modifications (Li et al., 2003, Nature 426:450, included herein by reference). In short, Huh 7 cells were washed twice with ice cold PBS, scraped off the plastic with a rubber policeman, pelleted and lysed in ice cold lysis buffer (0.3% DDM in PBS) containing protease inhibitors (Roche Complete Mini) at a final density of ˜2.5×107 cells/mL. Cell lysates were precleared with protein A sepharose beads after which 10 micrograms of probe S1 Fc was added to 1 ml of cell lysate and incubated for 1 hour at 4° C. under rotation. Precipitates were washed thrice with lysis buffer and once with PBS and subjected to NOVEX® 4 12% Tris Glycine gradient gel (Invitrogen) under reducing and non reducing conditions. A distinct 110 kDa band precipitated with EMC S1 Fc was visualized by GelCodeBlue staining, excised from the gel, incubated with trypsin and analyzed by MS. Results are shown in FIG. 20 and results of target analyses are shown in FIG. 21.