Selected article for: "chemiluminescent substrate and nonfat dry milk"
Author: Lamborn, Ian T.; Jing, Huie; Zhang, Yu; Drutman, Scott B.; Abbott, Jordan K.; Munir, Shirin; Bade, Sangeeta; Murdock, Heardley M.; Santos, Celia P.; Brock, Linda G.; Masutani, Evan; Fordjour, Emmanuel Y.; McElwee, Joshua J.; Hughes, Jason D.; Nichols, Dave P.; Belkadi, Aziz; Oler, Andrew J.; Happel, Corinne S.; Matthews, Helen F.; Abel, Laurent; Collins, Peter L.; Subbarao, Kanta; Gelfand, Erwin W.; Ciancanelli, Michael J.; Casanova, Jean-Laurent; Su, Helen C.
Title: Recurrent rhinovirus infections in a child with inherited MDA5 deficiency
  • Document date: 2017_7_3
    • ID: vipx6t7e_37
    Snippet: Immunoblotting 5 million cells were lysed in 2% SDS or LDS sample loading buffer (Pierce), with 5% 2-ME (Sigma-Aldrich), and heated to 95°C for 10 min. Proteins contained within the supernatants were quantified by BCA (Pierce). Unless otherwise indicated, 40 µg of protein were loaded per lane and were separated on NuPAGE Bis-Tris SDS-PAGE gels with MOPS running buffer (Invitrogen), followed by semi-dry transfer onto nitrocellulose membranes (Bi.....
    Document: Immunoblotting 5 million cells were lysed in 2% SDS or LDS sample loading buffer (Pierce), with 5% 2-ME (Sigma-Aldrich), and heated to 95°C for 10 min. Proteins contained within the supernatants were quantified by BCA (Pierce). Unless otherwise indicated, 40 µg of protein were loaded per lane and were separated on NuPAGE Bis-Tris SDS-PAGE gels with MOPS running buffer (Invitrogen), followed by semi-dry transfer onto nitrocellulose membranes (Bio-Rad Laboratories). After blocking with 5% nonfat dry milk (Bio-Rad Laboratories) in PBS containing 0.1% Tween 20 (Sigma-Aldrich), membranes were incubated with the antibodies directed against MDA5 (clone #D74E4), RIG-I (clone #D14G6), MAVS (clone #3993; all from Cell Signaling Technologies), β-actin (clone #AC-15; Sigma-Aldrich), and HSP90 (clone 68; BD). Signal was detected by incubation with appropriate HRP-conjugated secondary antibodies (Jackson ImmunoResearch Laboratories or Southern Biotech), followed by application of SuperSignal West Pico Chemiluminescent substrate or Su-perSignal West Dura Extended Duration Substrate (Thermo Fisher Scientific) and exposing to film.

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