Journal: Molecular medicine reports

Article Title: Preparation of a monoclonal antibody against the carcinoembryonic antigen, glypican‑3.

doi: 10.3892/mmr.2019.10019

Figure Lengend Snippet: Figure 3. Expression, purification and identification of recombinant GPC3 protein. (A) Induction expression of recombinant GPC3 protein by SDS‑PAGE analysis. Lane M, protein molecular weight marker; Lane 1, non‑induction of GPC3; lane 2, induction of GPC3. (B) SDS‑PAGE analysis of purification of recombinant GPC3 protein. M, standard protein molecular weight; lane 1, purified protein. (C) Confirmation of the recombinant protein by western blot analysis. The primary antibody was rabbit anti‑His‑tag monoclonal antibody. M, standard protein molecular weight; lane 1, purified protein. GPC3, glypican‑3.

Article Snippet: The recombinant plasmid was transfected into E. coli BL21 (DE3, Novagen; Merck KGaA) with the soluble fusion protein GPC3 expressed following induction with 0.1 mM isopropyl-β-Dthiogalactopyranoside (IPTG; Takara Bio, Inc.), and non-IPTG cells were used as control.

Techniques: Expressing, Purification, Recombinant, Molecular Weight, Marker, Western Blot

Journal: Biological chemistry

Article Title: Photoactivated inhibition of cathepsin K in a 3D tumor model

doi: 10.1515/hsz-2015-0274

Figure Lengend Snippet: (A) Taqman RT-PCR analysis of human CK gene expression in CK-1 and CK-2 clones. Data are normalized to HPRT1 and shown as average fold increase relative to EV (for 3 replicate experiments). (B) Western blot analysis of CK protein (37 kDa proenzyme and 28 kDa mature enzyme; top panel) and DDK (middle panel) expression in representative samples from PC3 prostate carcinoma cells stably transfected with empty vector (PC3-EV) or CTSK plasmids with DDK tag (PC3-CTSK); tubulin was used as loading control. (C) Immunofluorescent staining for DDK (green, left panel) indicative of CTSK expression; no primary antibody staining is shown as negative control (right panel); DAPI (blue) indicates nuclei; 40 × original magnification. (D) CK activity in CK clones. Assay was run against CK substrate Z-Gly-Pro-Arg-AMC the presence of cathepsin B inhibitor CA074. Data are shown as fold increase relative to CTSK activity in PC3-EV cells and a representative of three independent experiments. **p < 0.001; ***p < 0.0001 (values considered statistically significant).

Article Snippet: We confirmed minimal presence of CK mRNA in PC3 wild type cells (CT values > 30; data not shown) and utilized TrueORF ® Gold CK cDNA plasmids tagged with myc-DDK (OriGene) to establish stable CK-expressing clones (PC3-CK).

Techniques: Reverse Transcription Polymerase Chain Reaction, Expressing, Clone Assay, Western Blot, Stable Transfection, Transfection, Plasmid Preparation, Staining, Negative Control, Activity Assay