Journal: Clinical and Experimental Vaccine Research

Article Title: Novel antigen panel for modern broad-spectrum recombinant rotavirus A vaccine

doi: 10.7774/cevr.2021.10.2.123

Figure Lengend Snippet: Characterization of rotavirus A (RVA) recombinant antigens. Lanes: VP8 * -[P8] (1), VP8 * -[P6] (2), VP8 * -[P4] (3), ep-875 (4), VP5 * (5). (A) Electrophoresis analysis, 8%–20% SDS-PAGE (sodium dodecyl sulphate–polyacrylamide gel electrophoresis), staining with Coomassie G-250. (B) Western blot membrane stained by Ponceau S. (C) Western blot analysis with commercial polyclonal antisera to 5 RVA strains: Wa-G1P[8], Ds-1 G2P[4], Ito-G3P[8], HOCHI-G4P[8], 69M-G8P[10] (Cat# MBS316568; MyBioSource Inc., San Diego, CA, USA) as primary antibodies and secondary anti-species antibodies conjugated with horseradish peroxidase. Positions of molecular weights (kDa) markers are indicated in the right.

Article Snippet: The membranes were next treated with commercial polyclonal antisera to five RVA strains: Wa-G1P[8], Ds-1 G2P[4], Ito-G3P[8], Hochi-G4P[8], 69M-G8P[10] (Cat#MBS316568; MyBioSource Inc., San Diego, CA, USA), at a 1:500 dilution and then with secondary anti-species antibodies conjugated with horseradish peroxidase (Cat#713-035-003; Jackson ImmunoResearch Laboratories Inc., West Grove, PA, USA) in a 1:10,000 dilution.

Techniques: Recombinant, Electrophoresis, SDS Page, Polyacrylamide Gel Electrophoresis, Staining, Western Blot, Membrane

Journal: Biophysical Journal

Article Title: SMIT1 Modifies KCNQ Channel Function and Pharmacology by Physical Interaction with the Pore

doi: 10.1016/j.bpj.2017.06.055

Figure Lengend Snippet: The KCNQ2 pore module is required for binding to SMIT1. (A) Topologies of SMIT1 (putative), KCNQ α-subunits, and KCNE (experimentally confirmed). (B–E). CHO cells were transfected with KCNQ2 fragments and/or SMIT1-FLAG, as indicated. All blots are each representative of n = 2 experiments. The immunoprecipitating (IP) antibodies are labeled FLAG, YFP, or GFP, and the immunoblot (IB) antibodies are labeled GFP or FLAG. The red “X” denotes fragments that did not bind to SMIT1, and the green check mark signifies SMIT1-binding fragments. (B) Cytosolic Q2 fragments. (Top) The N-terminal fragment KCNQ2(1–97) does not bind to SMIT1-FLAG. (Bottom) The C-terminal fragment KCNQ2 (321–852) does not bind to SMIT1-FLAG. (C) Transmembrane Q2 fragments. (Top) The N-terminus through the S4-S5 linker fragment (KCNQ2(1–224)) does not bind to SMIT1. (Bottom) Extension of the previous fragment to include the pore-forming S5-S6 region (KCNQ2(1–549)) enables binding to SMIT1-FLAG. (D) The S5–S6 pore-forming region fragment (KCNQ2(222–323)) binds to SMIT1-FLAG. (E) Summary of binding results. (F) Cartoon view from the extracellular side comparing the Kv channel and solute transporter approximate dimensions and hypothetical docking configurations.

Article Snippet: Proteins were resolved by sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred onto polyvinylidine fluoride membranes for immunoblotting with the following antibodies, as noted: KCNQ2 (Santa Cruz Biotechnology, Santa Cruz, CA), DDK (Origene, Rockville, MD), FLAG (Sigma-Aldrich, St. Louis, MO), SMIT1 (Santa Cruz, CA), YFP (Santa Cruz), GFP (Santa Cruz; Rockland Immunochemicals, Pottstown, PA).

Techniques: Binding Assay, Transfection, Labeling, Western Blot

Journal:

Article Title: Adenovirus Serotype 30 Fiber Does Not Mediate Transduction via the Coxsackie-Adenovirus Receptor

doi: 10.1128/JVI.76.2.656-661.2002

Figure Lengend Snippet: Analysis of Ad5GFP and Ad5GFPf30 fibers by Western blot analysis. Purified Ad5GFP and Ad5GFPf30 were subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and viral proteins were transferred to nitrocellulose membranes. Membranes were incubated with a primary antibody to the N terminus of Ad5 fiber and then with a peroxidase-conjugated goat anti-mouse secondary antibody. Membranes were developed with enhanced-chemiluminescence reagent. The gel is representative of at least three independent experiments from different viral isolates.

Article Snippet: The membrane was then washed four times for 5 min with PBS-0.1% Tween 20 and incubated with peroxidase-conjugated goat anti-mouse secondary antibody (Jackson ImmunoResearch Laboratories, Inc., West Grove, Pa.) diluted 1:2,500 in PBS-0.1% Tween 20 for 1 h at RT.

Techniques: Western Blot, Purification, Polyacrylamide Gel Electrophoresis, Incubation