Western Blot:Article Title: Regulation of TAR DNA binding protein 43 (TDP-43) homeostasis by cytosolic DNA accumulation
Article Snippet: .. The following antibodies were used in this study: Mouse anti-TDP43 (R&D systems, MAB7778, epitope amino acids 1–103, 1:5000 for Western blot and 1:500 for immunostaining), Rabbit anti-TDP43 (Proteintech Group, 12892-1-AP, epitope amino acids 288–414, 1:5000 for Western blot), mouse anti-GAPDH (Proteintech Group, 60004-1-Ig, 1:5000 for Western blot), Rabbit anti-G3BP1 (Proteintech Group, 13057-2-AP, 1:300 for immunostaining), Rabbit anti-FUS (Proteintech Group, 11570-1-AP, 1:300 for immunostaining), Rabbit anti-GFAP (Proteintech Group, 16825-1-AP, 1:300 for immunostaining), rabbit anti-MAP2 (Proteintech Group, 17490-1-AP, 1:300 for immunostaining), rabbit anti-IBA-1 (Wako, 01,919,741, 1:300 for immunostaining), rat anti-mouse LAMP1 (1D4B) (BD Biosciences, 553,793, 1:300 for immunostaining), rabbit anti-EEA1 (Cell signaling, #3288, 1:200 for immunostaining), rabbit anti-phspho-Histone (S139) H2AX antibody (R&D systems, 4418-APC-100, 1:300 for immunostaining), rabbit anti-phospho-TDP-43 (Ser409/410) (Proteintech group, 80007-1-RR, 1:5000 for Western blot), rabbit anti-LaminB1 (Proteintech group, 66095-1-Ig, 1:5000 for Western blot), rabbit anti-KPNB1 (Proteintech group, 10077-1-AP, 1:3000 for Western blot and 1:300 for immunostaining), and rabbit anti-CRM1 (Proteintech group, 27917-1-AP, 1:5000 for Western blot and 1:300 for immunostaining). .. The following reagents were also used in the study: Dulbecco’s modified Eagle’s medium (DMEM) (Cellgro, 10–017-CV), HBSS (21–020-CV; Cellgro), DMEM/Ham’s F-12, (DMEM/F-12) (10–092-CV; Cellgro), 0.25% Trypsin (Corning, 25–053-CI), protease inhibitor (Roche, 05,056,489,001), Pierce BCA Protein Assay Kit (Thermo Scientific, 23,225), DNA damage inducer Etoposide (TCI America, E0675100 MG), PP2 (AdipoGen Life Sciences, AG-CR1-3563), Wortmannin (AdipoGen Life Sciences, AG-CN2-0023), Nu7441 (Apexbio Technology, 501,013,746), Lipopolysaccharides (LPS) (Sigma, L2654), polyinosinic-poly(C) potassium salt [poly(I: C)] (Sigma, 31,852–29–6), Imiquimod (Calbiochem, 99,011–02–6).
Article Title: Regulation of TAR DNA binding protein 43 (TDP-43) homeostasis by cytosolic DNA accumulation.
Article Snippet: .. The following antibodies were used in this study: Mouse anti-TDP43 (R&D systems, MAB7778, epitope amino acids 1-103, 1:5000 for western blot and 1:500 for immunostaining), Rabbit anti-TDP43 (Proteintech Group, 12892-1-AP, epitope amino acids 288-414, 1:5000 for western blot), mouse anti-GAPDH (Proteintech Group, 60004-1-Ig, 1:5000 for western blot), Rabbit anti-G3BP1 (Proteintech Group, 13057- 2-AP, 1:300 for immunostaining), Rabbit anti-FUS (Proteintech Group, 11570-1-AP, 1:300 for immunostaining), Rabbit anti-GFAP (Proteintech Group, 16825-1-AP, 1:300 for immunostaining), rabbit anti-MAP2 (Proteintech Group, 17490-1-AP, 1:300 for immunostaining), rabbit anti-IBA-1 (Wako, 01919741, 1:300 for immunostaining), rat anti-mouse LAMP1 (1D4B) (BD Biosciences, 553793, 1:300 for immunostaining), rabbit anti-EEA1 (Cell signaling, #3288, 1:200 for immunostaining), rabbit antiphspho-Histone (S139) H2AX antibody (R&D systems, 4418-APC-100, 1:300 for immunostaining), rabbit anti-phospho-TDP-43 (Ser409/410) (Proteintech group, 80007-1-RR, 1:5000 for western blot), rabbit anti- Jo urn al Pr e-p roo f 14 LaminB1 (Proteintech group, 66095-1-Ig, 1:5000 for western blot), rabbit anti-KPNB1 (Proteintech group, 10077-1-AP, 1:3000 for western blot and 1:300 for immunostaining), and rabbit anti-CRM1 (Proteintech group, 27917-1-AP, 1:5000 for western blot and 1:300 for immunostaining). .. The following reagents were also used in the study: Dulbecco’s modified Eagle’s medium (DMEM) (Cellgro, 10-017-CV), HBSS (21-020-CV; Cellgro), DMEM/Ham’s F-12, (DMEM/F-12) (10-092-CV; Cellgro), 0.25% Trypsin (Corning, 25-053-CI), protease inhibitor (Roche, 05056489001), Pierce BCA Protein Assay Kit (Thermo Scientific, 23225), DNA damage inducer Etoposide (TCI America, E0675100MG), PP2 (AdipoGen Life Sciences, AG-CR1-3563 ), Wortmannin (AdipoGen Life Sciences, AG-CN2-0023), Nu7441 (Apexbio Technology, 501013746), Lipopolysaccharides (LPS) (Sigma, L2654 ), polyinosinic-poly(C) potassium salt [poly(I: C)] (Sigma, 31852-29-6), Imiquimod (Calbiochem, 99011-02- 6).
Immunostaining:Article Title: Regulation of TAR DNA binding protein 43 (TDP-43) homeostasis by cytosolic DNA accumulation
Article Snippet: .. The following antibodies were used in this study: Mouse anti-TDP43 (R&D systems, MAB7778, epitope amino acids 1–103, 1:5000 for Western blot and 1:500 for immunostaining), Rabbit anti-TDP43 (Proteintech Group, 12892-1-AP, epitope amino acids 288–414, 1:5000 for Western blot), mouse anti-GAPDH (Proteintech Group, 60004-1-Ig, 1:5000 for Western blot), Rabbit anti-G3BP1 (Proteintech Group, 13057-2-AP, 1:300 for immunostaining), Rabbit anti-FUS (Proteintech Group, 11570-1-AP, 1:300 for immunostaining), Rabbit anti-GFAP (Proteintech Group, 16825-1-AP, 1:300 for immunostaining), rabbit anti-MAP2 (Proteintech Group, 17490-1-AP, 1:300 for immunostaining), rabbit anti-IBA-1 (Wako, 01,919,741, 1:300 for immunostaining), rat anti-mouse LAMP1 (1D4B) (BD Biosciences, 553,793, 1:300 for immunostaining), rabbit anti-EEA1 (Cell signaling, #3288, 1:200 for immunostaining), rabbit anti-phspho-Histone (S139) H2AX antibody (R&D systems, 4418-APC-100, 1:300 for immunostaining), rabbit anti-phospho-TDP-43 (Ser409/410) (Proteintech group, 80007-1-RR, 1:5000 for Western blot), rabbit anti-LaminB1 (Proteintech group, 66095-1-Ig, 1:5000 for Western blot), rabbit anti-KPNB1 (Proteintech group, 10077-1-AP, 1:3000 for Western blot and 1:300 for immunostaining), and rabbit anti-CRM1 (Proteintech group, 27917-1-AP, 1:5000 for Western blot and 1:300 for immunostaining). .. The following reagents were also used in the study: Dulbecco’s modified Eagle’s medium (DMEM) (Cellgro, 10–017-CV), HBSS (21–020-CV; Cellgro), DMEM/Ham’s F-12, (DMEM/F-12) (10–092-CV; Cellgro), 0.25% Trypsin (Corning, 25–053-CI), protease inhibitor (Roche, 05,056,489,001), Pierce BCA Protein Assay Kit (Thermo Scientific, 23,225), DNA damage inducer Etoposide (TCI America, E0675100 MG), PP2 (AdipoGen Life Sciences, AG-CR1-3563), Wortmannin (AdipoGen Life Sciences, AG-CN2-0023), Nu7441 (Apexbio Technology, 501,013,746), Lipopolysaccharides (LPS) (Sigma, L2654), polyinosinic-poly(C) potassium salt [poly(I: C)] (Sigma, 31,852–29–6), Imiquimod (Calbiochem, 99,011–02–6).
Article Title: Regulation of TAR DNA binding protein 43 (TDP-43) homeostasis by cytosolic DNA accumulation.
Article Snippet: .. The following antibodies were used in this study: Mouse anti-TDP43 (R&D systems, MAB7778, epitope amino acids 1-103, 1:5000 for western blot and 1:500 for immunostaining), Rabbit anti-TDP43 (Proteintech Group, 12892-1-AP, epitope amino acids 288-414, 1:5000 for western blot), mouse anti-GAPDH (Proteintech Group, 60004-1-Ig, 1:5000 for western blot), Rabbit anti-G3BP1 (Proteintech Group, 13057- 2-AP, 1:300 for immunostaining), Rabbit anti-FUS (Proteintech Group, 11570-1-AP, 1:300 for immunostaining), Rabbit anti-GFAP (Proteintech Group, 16825-1-AP, 1:300 for immunostaining), rabbit anti-MAP2 (Proteintech Group, 17490-1-AP, 1:300 for immunostaining), rabbit anti-IBA-1 (Wako, 01919741, 1:300 for immunostaining), rat anti-mouse LAMP1 (1D4B) (BD Biosciences, 553793, 1:300 for immunostaining), rabbit anti-EEA1 (Cell signaling, #3288, 1:200 for immunostaining), rabbit antiphspho-Histone (S139) H2AX antibody (R&D systems, 4418-APC-100, 1:300 for immunostaining), rabbit anti-phospho-TDP-43 (Ser409/410) (Proteintech group, 80007-1-RR, 1:5000 for western blot), rabbit anti- Jo urn al Pr e-p roo f 14 LaminB1 (Proteintech group, 66095-1-Ig, 1:5000 for western blot), rabbit anti-KPNB1 (Proteintech group, 10077-1-AP, 1:3000 for western blot and 1:300 for immunostaining), and rabbit anti-CRM1 (Proteintech group, 27917-1-AP, 1:5000 for western blot and 1:300 for immunostaining). .. The following reagents were also used in the study: Dulbecco’s modified Eagle’s medium (DMEM) (Cellgro, 10-017-CV), HBSS (21-020-CV; Cellgro), DMEM/Ham’s F-12, (DMEM/F-12) (10-092-CV; Cellgro), 0.25% Trypsin (Corning, 25-053-CI), protease inhibitor (Roche, 05056489001), Pierce BCA Protein Assay Kit (Thermo Scientific, 23225), DNA damage inducer Etoposide (TCI America, E0675100MG), PP2 (AdipoGen Life Sciences, AG-CR1-3563 ), Wortmannin (AdipoGen Life Sciences, AG-CN2-0023), Nu7441 (Apexbio Technology, 501013746), Lipopolysaccharides (LPS) (Sigma, L2654 ), polyinosinic-poly(C) potassium salt [poly(I: C)] (Sigma, 31852-29-6), Imiquimod (Calbiochem, 99011-02- 6).
Polyacrylamide Gel Electrophoresis:Article Title: Enhancing diabetic foot ulcer healing: Impact of the regulation of the FUS and ILF2 RNA-binding proteins through negative pressure wound therapy
Article Snippet: Total protein was extracted from cells and the granulation tissue from patients using radioimmunoprecipitation assay buffer (cat. no. P0013B; Beyotime Institute of Biotechnology), and protein concentrations were determined by bicinchoninic acid protein assay (cat. no. P0010; Beyotime Institute of Biotechnology). .. Equal amounts (20 μ g) of protein were loaded per lane and separated by 10% sodium dodecyl sulfate polyacrylamide gel electrophoresis, and transferred to polyvinylidene fluoride membranes, which were blocked with 5% skimmed milk at room temperature for 2 h. The membranes were probed overnight at 4°C with the following primary antibodies: Rabbit anti-FUS (1:5,000; cat. no. 11570-1-AP; Proteintech Group, Inc.), rabbit anti-ILF2 (1:1,000; cat. no. 14714-1-AP; Proteintech Group, Inc.) and mouse β-actin (1:1,000; cat. no. TA-09; ZSGB BIO; OriGene Technologies, Inc.). .. Subsequently, the membranes were incubated with horseradish peroxidase (HRP)-conjugated goat anti-rabbit secondary antibody (1:2,000; cat. no. ab205718; Abcam) or anti-mouse secondary antibody (1:5,000; cat. no. 550010; Chengdu Zen-Bioscience Co., Ltd.) at room temperature for 2 h. Enhanced chemiluminescence substrates (cat. no. SB-WB004; ShareBio; Shanghai Shenger Biotechnology Co., Ltd.) were used for signal detection, and ImageJ was used to identify the grayscale value of the target protein compared with the internal reference grayscale ratio.
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