The antibodies used in this study included polyclonal rabbit anti-tyrosine hydroxylase (Novus Biologicals, Littleton, CO, USA), polyclonal rabbit anti-inducible nitric oxide synthase (iNOS), anti-cyclooxygenase-2 (COX-2), and anti-glial fibrillary acidic protein (GFAP) (Abcam, Cambridge, MA, USA), polyclonal rabbit anti-ionized calcium binding adaptor molecule-1 (Iba-1) (Wako Chemicals, Richmond, VA, USA), biotinylated secondary anti-rabbit antibody (Jackson Immunoresearch, West Grove, PA, USA), and Alexa fluor 488-conjugated goat anti-rabbit secondary antibodies (Life Technologies, Grand Island, NY, USA). The test compound, thymol was procured from Santa Cruz Biotechnology Inc, CA, USA. ROT, the chemical to induce PD in rats were purchased from Sigma Aldrich, St. Louis, MO, USA. The ELISA assay kits for antioxidant enzymes and glutathione (GSH) as well as other analytical grade reagents were also obtained from Sigma Aldrich, St. Louis, MO, USA.
Anti glial fibrillary acidic protein
Manufactured by Abcam
Sourced in United States, United Kingdom
Anti-glial fibrillary acidic protein (GFAP) is a Type III intermediate filament protein that is expressed by numerous cell types of the central and peripheral nervous systems, including astrocytes and Schwann cells. GFAP is a reliable biomarker for the activation and proliferation of astrocytes, which can occur in response to central nervous system injury or disease.
Automatically generated - may contain errors
Lab products found in correlation
Anti neun, by Abcam (3 mentions)
Biotinylated anti rabbit secondary antibody, by Jackson ImmunoResearch (2 mentions)
Iba 1, by Abcam (2 mentions)
Cm3050, by Leica (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Horseradish peroxidase conjugated secondary antibody, by GE Healthcare (1 mentions)
Anti cd68, by Abcam (1 mentions)
Pvdf membrane, by Bio-Rad (1 mentions)
Western lightning plus ecl, by PerkinElmer (1 mentions)
Triton x 100, by Avantor (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by ZSGB-BIO (1 mentions)
Quick antigen retrieval solution for frozen sections, by Beyotime (1 mentions)
Anti p nf κb, by Abcam (1 mentions)
Anti rage, by Abcam (1 mentions)
Anti cd45, by Abcam (1 mentions)
Anti robo1, by Santa Cruz Biotechnology (1 mentions)
Anti neuronal nuclei neun, by Abcam (1 mentions)
Anti mpo, by Santa Cruz Biotechnology (1 mentions)
Anti slit2, by Santa Cruz Biotechnology (1 mentions)
Anti paxillin, by Santa Cruz Biotechnology (1 mentions)
15 protocols using anti glial fibrillary acidic protein
1
Immunohistochemical Analysis of Thymol's Neuroprotective Effects in Rotenone-Induced Parkinson's Disease
2
Protein Expression Analysis in Neuroscience
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Bicinchoninic Acid Protein (BCA) Assay was used to quantify the amount of protein in each sample. Equivalent amounts of protein were resolved by 12.5% Tricine polyacrylmide gel electrophoresis. Proteins were then transferred onto polyvinylidenedifluoride (PVDF) membranes (Bio-Rad, cat. no. 1620177). After the transfer, the membranes were blocked in 5% Blocking Milk (Carnation) in TBS (Tris-Base, NaCl) and incubated with various primary antibodies at room temperature for 1 hour or overnight at 4°C. Membranes are then washed with TBS-T (TBS and Tween) then incubated with horseradish peroxidase conjugated secondary antibodies (GE Healthcare UK Limited) at room temperature for 1 hour. Western blots were visualized using Western Lightning® Plus-ECL (PerkinElmer, LAS Inc.) Antibodies to the following proteins were used for this study: Anti-Neuronal Specific Enolase (NSE) (Abcam, cat. co. ab53025), Anti-Glial Fibrillary Acidic Protein (GFAP) (Abcam, cat. no. ab7260), Anti-CD68 (Abcam, cat. no. ab31630), and Actin (Santa Cruz Biotechnology Inc).
3
Immunohistochemical Analysis of Rat Spinal Cord Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Animals were anesthetized with sodium pentobarbital (60 mg/kg body weight) and
perfused with phosphate-buffered saline (PBS) followed by fresh 4%
paraformaldehyde. L3-5 SDHs were collected from rats, fixed in 4%
paraformaldehyde overnight and cryopreserved in 30% sucrose at 4°C overnight.
Tissues were mounted and sectioned on a cryostat at a thickness of 12 µm. Tissue
sections were permeabilized with 0.3% Triton X-100 (Amresco, Solon, USA) in PBS
for 15 min, followed by antigen retrieval with Quick Antigen Retrieval Solution
for Frozen Sections (Beyotime, Jiangsu, China). Then, the sections were
incubated with 3% BSA for 1 h at room temperature and then with primary
antibodies overnight at 4°C. The following primary antibodies were used:
anti-glial fibrillary acidic protein (GFAP; Abcam, Cambridge, UK), anti-ionized
calcium binding adaptor molecule 1 (IBA1; Abcam, Cambridge, UK), anti-NeuN
(Abcam, Cambridge, UK), anti-p-NF-κB (Abcam, Cambridge, UK) and anti-RAGE
(Abcam, Cambridge, UK). The tissue sections were washed three times and
incubated with the appropriate secondary antibodies for 1 h at room temperature.
After the slides were washed in PBS, coverslips were applied with mounting
medium with DAPI (ZSGB-BIO, Beijing, China). The sections were examined on an
Olympus fluorescence microscope (Olympus, Tokyo, Japan).
perfused with phosphate-buffered saline (PBS) followed by fresh 4%
paraformaldehyde. L3-5 SDHs were collected from rats, fixed in 4%
paraformaldehyde overnight and cryopreserved in 30% sucrose at 4°C overnight.
Tissues were mounted and sectioned on a cryostat at a thickness of 12 µm. Tissue
sections were permeabilized with 0.3% Triton X-100 (Amresco, Solon, USA) in PBS
for 15 min, followed by antigen retrieval with Quick Antigen Retrieval Solution
for Frozen Sections (Beyotime, Jiangsu, China). Then, the sections were
incubated with 3% BSA for 1 h at room temperature and then with primary
antibodies overnight at 4°C. The following primary antibodies were used:
anti-glial fibrillary acidic protein (GFAP; Abcam, Cambridge, UK), anti-ionized
calcium binding adaptor molecule 1 (IBA1; Abcam, Cambridge, UK), anti-NeuN
(Abcam, Cambridge, UK), anti-p-NF-κB (Abcam, Cambridge, UK) and anti-RAGE
(Abcam, Cambridge, UK). The tissue sections were washed three times and
incubated with the appropriate secondary antibodies for 1 h at room temperature.
After the slides were washed in PBS, coverslips were applied with mounting
medium with DAPI (ZSGB-BIO, Beijing, China). The sections were examined on an
Olympus fluorescence microscope (Olympus, Tokyo, Japan).
4
Immunofluorescence Staining of Slit2 and Robo1
5
Immunofluorescence Characterization of Vascular and Neural Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Briefly, rats were perfused with PBS and formalin after which the brain samples were collected and stored in formalin. Using a cryostat (CM3050S; Leica Microsystems, Bannockburn, IL), 10 µm thick sections were obtained. Immunofluorescence staining procedure was performed as described previously33 (link), 34 (link). The following primary antibodies were applied onto sections and incubated at 4 °C overnight: anti-Robo4 (1:100) or anti-Paxillin (1:100) (both from Santa Cruz Biotechnology, Dallas, TX) and co-localized with anti-von willibrand factor (vWF) (1:100) (Santa Cruz Biotechnology, Dallas, TX), anti-glial fibrillary acidic protein (GFAP) (1:100) or anti-neuronal nuclear protein (NeuN) (1:100) (both from Abcam, Cambridge, MA). Next, appropriate FITC- and Rhodamine Red-conjugated secondary antibodies (1:100) (Jackson Immuno Research, West Grove, PA) were applied at room temperature for 2 h. A fluorescence microscope (Olympus BX51) was used to visualize the sections.
6
Neuroinflammation and Oxidative Stress
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Polyclonal rabbit anti-cyclooxygenase-2 (COX-2), anti-inducible nitric oxide synthase (iNOS), and anti-glial fibrillary acidic protein (GFAP) antibodies were purchased from Abcam, Cambridge, MA, USA. Anti-ionized calcium-binding adaptor molecule-1 (Iba-1) polyclonal rabbit antibody was procured from Wako Chemicals, Richmond, VA, USA. Polyclonal rabbit anti-tyrosine hydroxylase antibody was obtained from Novus Biologicals, Littleton, CO, USA. Alexa fluor 488 conjugated secondary goat anti-rabbit antibodies were purchased from Life Technologies, Grand Island, NY, USA. Biotinylated secondary anti-rabbit antibody was purchased from Jackson Immunoresearch, West Grove, PA, USA. Nerolidol was purchased from Santa Cruz Biotechnology Inc., CA, USA. ROT and the assay kit for reduced glutathione (GSH) and other reagents of analytical grade were purchased from Sigma-Aldrich, St. Louis, MO, USA.
7
Histological Analysis of Spinal Cord Tissues in SCI Rats
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The spinal cord tissues were collected from SCI-induced rats for histological analysis on day 28 (n = 3 per group). The tissues were fixed in 10% neutral-buffered formalin, embedded in paraffin, and sectioned (thickness: 4 μm). Hematoxylin and eosin staining of the sections was conducted using Dako CoverStainer (Agilent, Santa Clara, CA, USA). The following antibodies were used for immunohistochemistry analysis: anti-ionized calcium-binding adapter molecule 1 (Iba-1) (Abcam, Cambridge, UK) and anti-glial fibrillary acidic protein (GFAP) (Abcam) to determine neuroinflammation. All stained slides were scanned using a Pannoramic SCAN II (3DHISTECH Kft., Budapest, Hungary). Photomicrographs were captured using CaseViewer software 2.5 (3DHISTECH Kft.). The signal was quantified using the ImageJ software (NIH, Bethesda, MD, USA).
8
IHC Analysis of p22 phox and GFAP
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For IHC analysis, the free-floating method was used for the selected tissue in each treatment group. After washing the tissue 3× for 10 min each in 0.01 M PBS, the tissue was incubated at 80 °C for 60 min in a beaker containing 0.01 M sodium citrate, followed by blocking in 10% normal donkey serum (Millipore Sigma, Burlington, MA, USA). The tissue sample was then reacted with primary antibodies anti-p22 phox (Santa Cruz Biotechnology, dilution: 1:500) and anti-Glial fibrillary acidic protein (GFAP) (Abcam, dilution: 1:500) for 12 h at 4 °C. The following day, the tissue was washed 3× for 5 min each in 0.01 M PBS and then reacted with HRP-conjugated goat anti-mouse secondary antibody (Santa Cruz Biotechnology, dilution: 1:5000) at room temperature for 2 h. Finally, the tissue was incubated at room temperature using a Vectastain-Elite ABC kit (Vector Laboratories, Burlingame, CA, USA) solution, and the results were visualized using the DAB Peroxidase Substrate Kit (Vector Laboratories). Each tissue slice was mounted on a slide using a mounting medium (Vector Laboratories) and inspected using a light microscope (Leica Microsystems).
9
Immunofluorescence Staining of Tissue and Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
For immunofluorescence staining, OCT-embedded frozen section (tissue) or cell climbing side (cell) was fixed with pre-cooled acetone for 10 min at RT, followed by blocking with goat serum for 1 h. Samples were incubated with primary antibodies at 4℃ overnight. These primary antibodies were used: anti-NeuN (Millipore-Chemicon), anti-CD31 (RD Systems), anti-caveolin-1, anti-glial fibrillary acidic protein (GFAP) (Abcam), anti-desmin (Abclonal), anti-cleaved caspase 3 (CST) and anti-albumin. After washing in PBS three times, the corresponding secondary antibody was subsequently used in the sample at RT for 1 h. The secondary antibodies included Alexa Fluor 594 (goat anti-rabbit) (Abcam), Alexa Fluor 488 (Goat anti-mouse) (Abcam), and Alexa Fluor 488 (Goat anti-chicken) (Invitrogen). Finally, the sample was washed and stained with diaminobenzidine/imidazole (Sigma). Immunofluorescence images were acquired by a fluorescence microscope (Nikon).
10
Immunohistochemical Analysis of Spinal Cord
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The spinal cord tissues of the TRANSPLANT+PEG group and the TRANSPLANT group were cut into 10‐μm longitudinal sections, while the bridging tissue of the spinal cord of one beagle in the TRANSPLANT+PEG group was cut into 10‐μm coronal sections. The sections were then mounted onto silanized glass slides. Slides were air‐dried for 15 min and washed in PBS before blocking in 10% normal bovine serum and 0.1% Triton X‐100 in PBS at room temperature. Primary antibodies diluted in Dako antibody diluent were applied overnight in a humidified chamber at 4°C. Primary antibodies included anti‐neurofilament heavy polypeptide (1:100, Abcam) and anti‐glial fibrillary acidic protein (GFAP; 1:1000, Abcam). Slides were then washed and secondary antibodies (IgG‐H&L, Alexa Fluor 594; 1:500, Abcam) that were diluted in PBS were applied for 45 min at room temperature. This procedure was repeated again for a duration of 30 min using IgG‐H&L, Alexa Fluor 488 (1:500, Abcam). Finally, the slides were washed in PBS prior to being mounted with a coverslip using VECTASHIELD (Vector Laboratories).
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!