Journal: PLoS ONE

Article Title: Functional Characterization of a CRH Missense Mutation Identified in an ADNFLE Family

doi: 10.1371/journal.pone.0061306

Figure Lengend Snippet: To investigate intracellular distribution of CRH, cell were fixed in PFA and probed with mouse polyclonal anti-GM130 (red) for Golgi visualization and rabbit polyclonal anti-CRH (green) antibodies.

Article Snippet: After washing twice with PBS buffer, wells were probed overnight in PBS containing 5% (w/v) BSA with anti-CRH rabbit polyclonal antibody (1∶1000) (Source Biosciences).

Techniques:

Journal: Scientific Reports

Article Title: Microtubule stabilising peptides rescue tau phenotypes in-vivo

doi: 10.1038/srep38224

Figure Lengend Snippet: For each phospho-tau antigen, intensity of signal (pixels/mm ) was normalised to total tau levels. There was no significant change in the levels of the phospho-tau epitopes detected by AT180 ( a ), AT8 ( b ), and PHF-1 ( c ) after treatment with 5 μg/ml SAL (green bars) and 10 μg/ml SAL (dark green bars) SAL. Total dtau levels were not altered by SAL treatment ( d ). Representative blots are shown (a-d). All lanes were run on the same gel. Data were analysed by one-way ANOVA with Bonferroni’s correction. Error bars represent mean ± S.E.M; n = htau 0N3R (5), 5 μg/ml SAL (5), 10 μg/ml SAL (5).

Article Snippet: Blots were probed with the following primary antibodies: anti-human tau (1:15,000, Dako, UK), anti-dtau (1:500, gifted by Prof. St. Johnston, University of Cambridge, UK), anti-actin (1:5000, Abcam, UK) and anti-phospho-tau: PHF-1 (1:2000, gifted by Dr. Peter Davies, Albert Einstein College of Medicine; Bronx, NY), AT180 (1:100, Source, Biosciences, UK), AT8 (1:800, Source, Biosciences, UK).

Techniques:

Journal: Cell chemical biology

Article Title: Hypomorph mutation-directed small-molecule protein-protein interaction inducers to restore mutant SMAD4-suppressed TGF-β signaling

doi: 10.1016/j.chembiol.2020.11.010

Figure Lengend Snippet: (A) Schematic illustration of the design of the TR-FRET assay for monitoring the SMAD4/SMAD3 PPI to discover small-molecule PPI inducers. Anti-Flag-Tb coupled with Flag-SMAD3 serves as the TR-FRET donor and anti-His-D2 coupled with His-SMAD4R361H serves as the acceptor. At the basal level, R361H impairs SMAD4 interaction with SMAD3, yielding low TR-FRET signal. Upon treatment with a PPI inducer, the induced SMAD3/SMAD4R361H complex formation brings two fluorophores into close proximity (<10 nm), generating a high TR-FRET signal.

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Tb cryptate-conjugated mouse monoclonal anti-Flag antibody Cisbio Bioassays Cat# 61FG2TLB D2-conjugated anti-HIS antibody Cisbio Bioassays Cat# 61HISDLF Mouse monoclonal anti-FLAG-HRP antibody Sigma-Aldrich Cat# A8592; RRID: AB_439702 Rabbit polyclonal anti-GST-HRP antibody Sigma-Aldrich Cat# A7340; RRID: AB_258340 Mouse monoclonal anti-β-Actin antibody Sigma-Aldrich Cat# A5441; RRID: AB_476744 Rabbit monoclonal anti-p21 antibody Cell signaling Cat# 2947S; RRID: AB_823586 Mouse monoclonal anti-p15 antibody Santa Cruz Cat# sc-271791; RRID: AB_10709436 SMAD4 (D3M6U) Rabbit mAb Cell signaling Cat# 38454; RRID: AB_2728776 SMAD3 mouse monoclonal Antibody (38-Q) Santa Cruz Cat# sc-101154; RRID: AB_1129525 Mouse Anti-Rabbit IgG (Light-Chain Specific) (D4W3E) mAb (HRP Conjugate) Cell signaling Cat# 93702; RRID: AB_2800208 Peroxidase AffiniPure Goat Anti-Rabbit IgG(H+L) secondary antibody Jackson ImmunoResearch Cat# 111–035-003 Peroxidase AffiniPure Goat Anti-Mouse IgG(H+L) secondary antibody Jackson ImmunoResearch Cat# 115–035-003 Chemicals, Peptides, and Recombinant Proteins Recombinant Human TGF-beta 1 Protein R&D Systems Cat# 240-B Ro-31-8220 Cayman Cat# 138489–18-6 Go-6983 Cayman Cat# 133053–19-7 Ro-31-7549 Cayman Cat# 138516–31-1 Ro-31-8425 Cayman Cat# 145317–11-9 Ro-32-0432 Cayman Cat# 145333–02-4 UCN-1028c Cayman Cat# 121263–19-2 PKC-412 Cayman Cat# 120685–11-2 EEBL Emory ECBDC ( Mo et al., 2019 ) N/A FuGENE Promega Cat# E2311 SBE4-Luc gift from Bert Vogelstein ( Zawel et al., 1998 ) Addgene plasmid # 16495 Dual-Glo luciferase kit Promega Cat# E2920 Oligonucleotides p21-F: 5′-ATGTGTCCTGGTTCCCGTTTC-3′; IDT N/A p21-R: 5′-CATTGTGGGAGGAGCTGTGA-3′; IDT N/A p15-F: 5′- GGACTAGTGGAGAAGGTGCG-3′; IDT N/A p15-R: 5′-GGGCGCTGCCCATCATCATG-3′; IDT N/A GAPDH-F: 5′-GAAGGTGAAGGTCGGAGT-3′; IDT N/A GAPDH-R: 5′-GAAGATGGTGATGGGATTTC-3′.

Techniques:

Journal: Cell chemical biology

Article Title: Hypomorph mutation-directed small-molecule protein-protein interaction inducers to restore mutant SMAD4-suppressed TGF-β signaling

doi: 10.1016/j.chembiol.2020.11.010

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Tb cryptate-conjugated mouse monoclonal anti-Flag antibody Cisbio Bioassays Cat# 61FG2TLB D2-conjugated anti-HIS antibody Cisbio Bioassays Cat# 61HISDLF Mouse monoclonal anti-FLAG-HRP antibody Sigma-Aldrich Cat# A8592; RRID: AB_439702 Rabbit polyclonal anti-GST-HRP antibody Sigma-Aldrich Cat# A7340; RRID: AB_258340 Mouse monoclonal anti-β-Actin antibody Sigma-Aldrich Cat# A5441; RRID: AB_476744 Rabbit monoclonal anti-p21 antibody Cell signaling Cat# 2947S; RRID: AB_823586 Mouse monoclonal anti-p15 antibody Santa Cruz Cat# sc-271791; RRID: AB_10709436 SMAD4 (D3M6U) Rabbit mAb Cell signaling Cat# 38454; RRID: AB_2728776 SMAD3 mouse monoclonal Antibody (38-Q) Santa Cruz Cat# sc-101154; RRID: AB_1129525 Mouse Anti-Rabbit IgG (Light-Chain Specific) (D4W3E) mAb (HRP Conjugate) Cell signaling Cat# 93702; RRID: AB_2800208 Peroxidase AffiniPure Goat Anti-Rabbit IgG(H+L) secondary antibody Jackson ImmunoResearch Cat# 111–035-003 Peroxidase AffiniPure Goat Anti-Mouse IgG(H+L) secondary antibody Jackson ImmunoResearch Cat# 115–035-003 Chemicals, Peptides, and Recombinant Proteins Recombinant Human TGF-beta 1 Protein R&D Systems Cat# 240-B Ro-31-8220 Cayman Cat# 138489–18-6 Go-6983 Cayman Cat# 133053–19-7 Ro-31-7549 Cayman Cat# 138516–31-1 Ro-31-8425 Cayman Cat# 145317–11-9 Ro-32-0432 Cayman Cat# 145333–02-4 UCN-1028c Cayman Cat# 121263–19-2 PKC-412 Cayman Cat# 120685–11-2 EEBL Emory ECBDC ( Mo et al., 2019 ) N/A FuGENE Promega Cat# E2311 SBE4-Luc gift from Bert Vogelstein ( Zawel et al., 1998 ) Addgene plasmid # 16495 Dual-Glo luciferase kit Promega Cat# E2920 Oligonucleotides p21-F: 5′-ATGTGTCCTGGTTCCCGTTTC-3′; IDT N/A p21-R: 5′-CATTGTGGGAGGAGCTGTGA-3′; IDT N/A p15-F: 5′- GGACTAGTGGAGAAGGTGCG-3′; IDT N/A p15-R: 5′-GGGCGCTGCCCATCATCATG-3′; IDT N/A GAPDH-F: 5′-GAAGGTGAAGGTCGGAGT-3′; IDT N/A GAPDH-R: 5′-GAAGATGGTGATGGGATTTC-3′.

Techniques: Recombinant, Plasmid Preparation, Luciferase, Viability Assay, Software