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. 2021 Jan 8;11:585361. doi: 10.3389/fimmu.2020.585361

Figure 1.

Figure 1

Characterization of purified C4BP. (A) SDS-PAGE (12% w/v acrylamide/bisacrylamide) was loaded with 5 µg of purified C4BP in lane 2, along with a protein ladder with a range of 250 to 10 kDa (Thermofisher) in lane 1. The denatured and reduced samples were run for 120 min at 90 V and stained with Coomassie brilliant blue to reveal protein bands corresponding to the α-chain (~70kDa) and a faint band corresponding to β-chain of C4BP (~45 kDa). (B) Immunoreactivity of the purified C4BP was analyzed by western blotting. A PVDF membrane with 1.25, 2.5, and 5 µg of purified C4BP in lanes 2, 3, and 4, respectively, along with a protein ladder with a range of 250 to 10 kDa (Thermofisher) in lane 1 was probed using rabbit-anti-human C4BP polyclonal antibodies (1:1,000) at room temperature for 1 h, followed by incubation with secondary goat anti-rabbit IgG HRP-conjugate (1:1,000) for 1 h at room temperature. Bands corresponding to α-chain (~70 kDa) and the β-chain (~45 kDa) of the C4BP were observed after developing the color using 3,3′-diaminobenzidine (DAB) substrate.