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. 2013 Jan 30;8(1):e55338. doi: 10.1371/journal.pone.0055338

Figure 1. Overexpression of human LF in HEK293 cells using Flp-In T-Rex system.

Figure 1

(A) End-point RT-PCR analysis of LF. RT-PCR was conducted for RNAs from HEK293 cells transfected with T-Rex LF expression vectors with signal sequence (sLF) or without signal sequence and 26 N-terminal amino acid residues (ΔLF), non-treated with (Tet(−), top panel) or treated with tetracycline (Tet(+), bottom panel) for 24 or 36 h. C, vector alone. M, size marker. (B) Real-time RT-PCR analysis of LF. Each reaction per clone was carried out in duplicate, and the average of 15 clones is presented along with the standard error. (C) Immunoblot analysis of LF. LF protein expressed from recombinant vector was immunobloted using anti-LF antibody from cell lysate and culture media. LF, commercially available sLF (80 kDa). C, vector alone. LFs expressed from vectors with and without the signal sequence and 26 N-terminal amino acid residues are marked by arrows. Signals below the LF bands appearing in all samples in the medium are non-specifically-reacted serum proteins. The result of ELISA for the LF in medium is shown at the bottom.