Anti gfap

Manufactured by ABclonal

Anti-GFAP is a primary antibody that binds to the GFAP (Glial Fibrillary Acidic Protein) antigen. GFAP is a type III intermediate filament protein that is expressed in astrocytes and other glial cells in the central nervous system. The Anti-GFAP antibody can be used to identify and study astrocytes in various research applications.

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4 protocols using anti gfap

Immunohistochemical Analysis of Neural Markers

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We used a method described recently.³³, ³⁴ Sections were first deparaffinized and were then rehydrated using ethanol, boiled in 0.01 mol/L sodium citrate buffer (pH = 6.0) for 25 minutes. Sections were blocked by PBS containing 5% bovine serum albumin (BSA) for 30 minutes and incubated overnight at 4°C with the following antibodies: anti‐NeuN (ABN781:1000; Millipore), anti‐MAP2 (MAB3418, 1:500; Millipore), anti‐GFAP (A14673; 1:500; Abclonal), and anti‐Iba1 (019‐19741; 1:500; Wako). On the next day, the sections were incubated for 1 hour at room temperature in PBS with secondary antibodies (1:500) including Alexa Fluor 488 goat anti‐mouse/anti‐rabbit and Alexa Fluor 594 goat anti‐mouse/anti‐rabbit (Invitrogen). Images were captured and analyzed using a ZEISS LSM‐880 confocal laser‐scanning microscope.

Bi H., Zhou C., Zhang Y., Cai X., Ji M., Yang J., Chen G, & Hu Y. (2020). Neuron‐specific deletion of presenilin enhancer2 causes progressive astrogliosis and age‐related neurodegeneration in the cortex independent of the Notch signaling. CNS Neuroscience & Therapeutics, 27(2), 174-185.

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Protein Expression Analysis by Western Blotting

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Western blotting was performed with specific antibodies against human. All antibodies were used at the dilution according to manufacturer including anti-ST2 (HPA007917; Sigma), anti-IL-33 (Nessy-1; Enzo) and anti-GFAP (A14673; ABclonal), anti-POU5F1 (A7920; ABclonal), anti-Sox2 (A0561; ABclonal), anti-Nestin (19483-1-AP; Proteintech), anti-β-catenin (17565-1-AP; Proteintech) and anti-CD133 (64326; Cell Signaling Technology), anti-E-cadherin (3195; Cell Signaling Technology), anti-N-cadherin (13116; Cell Signaling Technology), anti-Vimentin (5741; Cell Signaling Technology), anti-SAPK (JNK) (9252; Cell Signaling Technology), anti-P38 (8690; Cell Signaling Technology), anti-P44/42 (4695; Cell Signaling Technology), anti-NF-κB (10745-1-AP; Proteintech), anti-p-SAPK (JNK) (4668; Cell Signaling Technology), anti-p-P44/42 (4377; Cell Signaling Technology), anti-p-P38 (4511; Cell Signaling Technology), anti-p-NF-κB (3033; Cell Signaling Technology) and anti-GAPDH (Cell Signaling Technology). Following incubation in HRP labeled secondary antibody (Cell Signaling Technology). Specific protein bands were scanned with ECL detection reagents and detected by Gel Doc 2000 (Bio-Rad). The bands of western blots were quantified by ImageJ and shown in the Supplementary Figure5.

Lin L., Li Y., Liu M., Li Q., Liu Q, & Li R. (2020). The Interleukin-33/ST2 axis promotes glioma mesenchymal transition, stemness and TMZ resistance via JNK activation. Aging (Albany NY), 12(2), 1685-1703.

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Immunofluorescence Analysis of Neuroinflammation Markers

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Spinal cord sections were dewaxed, conducted to antigen retrieval (Improved Citrate Antigen Retrieval Solution, P0083, Beyotime Biotechnology, Shanghai, CHN), treated with 3% hydrogen peroxide for 10 min, blocked with immunofluorescence blocking solution (P0102, Beyotime Biotechnology, Shanghai, CHN, room temperature) for 1 h, and then incubated with primary antibody overnight at 4°C, subsequently, incubated with fluorescent secondary antibody at room temperature for 1 h and observed under a fluorescence microscope (IX73, Olympus Corp, Tokyo, JPN). The fluorescence intensities were analyzed using ImageJ 1.51j8 (National Institutes of Health, US). The following primary antibodies were used: anti-IL-1β (1:100, A19635), anti-GFAP (1:100, A0237), anti-NLRP3 (1:100, A5652), anti-caspase-1 (1:100, A0964), anti-Bcl-2 (1:100, A0208) and anti-GSK-3β (1:100, A6164) were from Abclonal (Wuhan, CHN); anti-NF-κB (1:100, BF8005) and anti-DHODH (1:100, DF3991) were from Affinity (Jiangsu, CHN). The secondary antibodies used for immunofluorescence analysis were goat anti-mouse IgG H&L (1:100, FITC, ab6785) and goat anti-rabbit IgG H&L (1:100, FITC, ab6717) were purchased from Abcam (Cambridge, UK).

Yang H.Y., Sun X., Zhen S.Q., Yu L.Z., Ding J.Q., Liu L., Xie M, & Zhu H.L. (2023). GSK-3β inhibition alleviates arthritis pain via reducing spinal mitochondrial reactive oxygen species level and inflammation. PLOS ONE, 18(4), e0284332.

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Western Blotting Analysis of Neuroinflammation

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For western blotting analysis, total proteins were collected from striatum, SN, and BV2 cells and stored at −80 °C. 40 ug of total protein lysate was loaded onto a 12% sodium-dodecyl-sulfate polyacrylamide gel in each lane, then transferred onto a polyvinylidene-difluoride membrane (Millipore, United Ststes). The primary antibodies were incubated overnight at 4 °C included anti-TH (1:500; Santa Cruz, sc-25269), anti-solute carrier family 6 member 3 (SLC6A3) (1:1,000; ABclonal, A152360), anti-dopamine receptor D2 (DRD2) (1:1,000; ABclonal, A12930), anti-AIF1 (1:1,000; Santa Cruz, sc-32725), anti-GFAP (1:1,000; ABclonal, A14673), anti-NLRP3 (1:200; AdipoGen, AG-20B-0014-C100), anti-PYCARD (1:1,000; Immunoway, T0365), anti-CASP1 (1:1,000; ABclonal, A0964), anti-IL1B (1:1,000; ABclonal, A12688), anti-MAP1LC3B (1:1,000; ABclonal, A11282), anti-BECN1 (1:1,000; Cell Signaling Technology, 3738S) and anti-ACTB (1:3,000; ABclonal, AC026). HRP-conjugated anti-Rabbit antibody (1:5,000; ABclonal, AS014) or anti-Mouse antibody (1:5,000; ABclonal, AS003) was used as secondary antibody. ImageJ software was used to quantify the target bands and ACTB as an internal control.

Huang S., Liu H., Lin Y., Liu M., Li Y., Mao H., Zhang Z., Zhang Y., Ye P., Ding L., Zhu Z., Yang X., Chen C., Zhu X., Huang X., Guo W., Xu P, & Lu L. (2021). Berberine Protects Against NLRP3 Inflammasome via Ameliorating Autophagic Impairment in MPTP-Induced Parkinson’s Disease Model. Frontiers in Pharmacology, 11, 618787.

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