Figure 2
From: Establishment of a highly sensitive sandwich ELISA for the N-terminal fragment of titin in urine
Characterization of antibodies, 53A1 and 144A2.
(a) 0.5 mL of culture supernatant of recombinant CHO-K1 cells expressing FLAG-Titin-N-His and 3 mL of urine from a healthy volunteer were immuno-precipitated with 2 μg of 53A1, 144A2 and 4A1 antibodies. The Flag-Titin-N-His fusion proteins in the conditioned medium of CHO-K1 cells (lane 1), the Flag-Titin-N-His fusion proteins immuno-precipitated from the conditioned medium of CHO-K1 cells with 53A1 (lane 2), 144A2 (lane 3), and 4A1 (lane 4) and the titin proteins immuno-precipitated from urine of healthy volunteer with 53A1(lane 5), 144A2 (lane 6) and 4A1 (lane 7) were applied on SDS-PAGE and blotted with anti-Titin-N (A69-S86) rabbit IgG Fab’-HRP. 4A1 (Anti-Titin-C fragment mouse IgG monoclonal antibody), denoted by asterisk was used as a negative control. (b) Culture supernatant of recombinant CHO-K1 cells expressing FLAG-Titin-N-His proteins was incubated with glycopeptidase F (GPF, Takara Bio), and subjected to SDS-PAGE (Lane 2) and the gel was blotted with 53A1, an anti-Titin-N Mouse IgG Fab’-HRP.
