产品图片  Sample:Optic nerve (Rat)cell Lysate at 40 ug Primary: Anti-GFAP(bs-0199R)at 1/300 dilution Secondary: IRDye800CW Goat Anti-RabbitIgG at 1/20000 dilution Predicted band size: 48 kD Observed band size: 53 kD  Sample: U251 Cell Lysate at 40 ug Primary: Anti- GFAP (bs-0199R) at 1/300 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/10000 dilution Predicted band size: 48 kD Observed band size: 50 kD  Sample: Cerebellum (Rat) Lysate at 40 ug Cerebellum (Mouse) Lysate at 40 ug Eye (Mouse) Lysate at 40 ug Primary: Anti-GFAP (bs-0199R) at 1/300 dilution Secondary: IRDye800CW Goat Anti-Rabbit IgG at 1/20000 dilution Predicted band size: 48 kD Observed band size: 48 kD  Tissue/cell: rat brain tissue; 4% Paraformaldehyde-fixed and paraffin-embedded; Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min; Incubation: Anti-GFAP Polyclonal Antibody, Unconjugated(bs-0199R) 1:400, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining  Paraformaldehyde-fixed, paraffin embedded (mouse cerebellum); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.  Paraformaldehyde-fixed, paraffin embedded (rat cerebellum); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.  Paraformaldehyde-fixed, paraffin embedded (mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.  Paraformaldehyde-fixed, paraffin embedded (rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:200 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.  Paraformaldehyde-fixed, paraffin embedded (Mouse brain); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:400 overnight at 4°C, followed by operating according to SP Kit(Rabbit) (sp-0023) instructionsand DAB staining.  Tissue/cell: rat brain tissue; 4% Paraformaldehyde-fixed and paraffin-embedded; Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Block endogenous peroxidase by 3% Hydrogen peroxide for 30min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min; Incubation: Anti-GFAP Polyclonal Antibody, Unconjugated(bs-0199R) 1:500, overnight at 4°C, followed by conjugation to the secondary antibody(SP-0023) and DAB(C-0010) staining  Tissue/cell:SH-SY5Y cell; 4% Paraformaldehyde-fixed; Triton X-100 at room temperature for 20 min; Blocking buffer (normal goat serum, C-0005) at 37°C for 20 min; Antibody incubation with (GFAP) polyclonal Antibody, Unconjugated (bs-0199R) 1:100, 90 minutes at 37°C; followed by a FITC conjugated Goat Anti-Rabbit IgG antibody at 37°C for 90 minutes, DAPI (blue, C02-04002) was used to stain the cell nuclei.  Paraformaldehyde-fixed, paraffin embedded (Human brain glioma); Antigen retrieval by boiling in sodium citrate buffer (pH6.0) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:400 overnight at 4°C, followed by a conjugated Goat Anti-Rabbit IgG antibody (bs-0295G-FITC) for 90 minutes, and DAPI for nuclei staining.  Paraformaldehyde-fixed, paraffin embedded (rat brain); Antigen retrieval by boiling in sodium citrate buffer (pH6) for 15min; Block endogenous peroxidase by 3% hydrogen peroxide for 20 minutes; Blocking buffer (normal goat serum) at 37°C for 30min; Antibody incubation with (GFAP) Polyclonal Antibody, Unconjugated (bs-0199R) at 1:200 overnight at 4°C, followed by a conjugated secondary (bs-0295G-Cy3) at [1:500] for 90 minutes and DAPI staining of the nuclei.  Tissue/cell: rat brain tissue;4% Paraformaldehyde-fixed and paraffin-embedded; Antigen retrieval: citrate buffer ( 0.01M, pH 6.0 ), Boiling bathing for 15min; Blocking buffer (normal goat serum,C-0005) at 37℃ for 20 min; Incubation: Anti-GFAP Polyclonal Antibody, Unconjugated(bs-0199R) 1:400, overnight at 4°C; The secondary antibody was Goat Anti-Rabbit IgG, Cy3 conjugated(bs-0295G-Cy3)used at 1:200 dilution for 40 minutes at 37°C. DAPI(5ug/ml,blue,C-0033) was used to stain the cell nuclei  Blank control: RSC96(blue). Primary Antibody:Rabbit Anti- GFAP antibody(bs-0199R), Dilution: 1μg in 100 μL 1X PBS containing 0.5% BSA; Isotype Control Antibody: Rabbit IgG(orange) ,used under the same conditions ); Secondary Antibody: Goat anti-rabbit IgG-PE(white blue), Dilution: 1:200 in 1 X PBS containing 0.5% BSA. Protocol The cells were fixed with 2% paraformaldehyde (10 min) , then permeabilized with 90% ice-cold methanol for 30 min on ice. Primary antibody (bs-0199R, 1μg /1x10^6 cells) were incubated for 30 min on the ice, followed by 1 X PBS containing 0.5% BSA + 1 0% goat serum (15 min) to block non-specific protein-protein interactions. Then the Goat Anti-rabbit IgG/PE antibody was added into the blocking buffer mentioned above to react with the primary antibody at 1/200 dilution for 30 min on ice. Acquisition of 20,000 events was performed. 已发表文献 [IF=11.42]Zhao, Yan G., et al. "The autophagy gene Wdr45/Wipi4 regulates learning and memory function and axonal homeostasis." Autophagy (2015).IHC-P, Mouse [IF=10.82]Zhao, Hongyu, et al. "Mice deficient in Epg5 exhibit selective neuronal vulnerability to degeneration." The Journal of Cell Biology (2013).IHC-P, Mouse [IF=10.53]Ma, Benyu, et al. "Dapper1 promotes autophagy by enhancing the Beclin1-Vps34-Atg14L complex formation." Cell Research (2014).IHC-F, Mouse [IF=7.58]Shan, Chun-Lei, et al. 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