The tissue was homogenized in RIPA buffer containing a protease inhibitor cocktail (Beyotime Biotechnology). A BCA protein assay (Beyotime Biotechnology) was used to determine the protein concentrations. Equal amounts of protein were separated by SDS-PAGE and then transferred electrophoretically to polyvinylidene fluoride membranes (Bio-Rad) [29 (link), 30 (link)]. The membranes were incubated with the following primary antibodies for 2 h at room temperature: IL-6 (1:1000, Abcam, England), Gp130 (1:1000, Santa, America), pSTAT3 (1:1000, Bioss, China), and NMDA receptors (1:1000, Bioss, China). The membranes were then incubated with GAPDH (1:1000, Solarbio, China), goat anti-mouse IgG (Bioss, China, Gp130, 1:1000 and GAD67, 1:1000) or goat anti-rabbit IgG (Bioss, China, pSTAT3, 1:1000, NMDA receptors, 1:3000 and IL-6, 1:1000) secondary antibodies for 2 h at room temperature. Finally, the membranes were placed in a gel imaging analysis system for exposure and analysis (AlphaView FluorChem FC3).
Nmda receptors
Manufactured by Bioss Antibodies
Sourced in China
NMDA receptors are ionotropic glutamate receptors found in the central nervous system. They play a crucial role in synaptic transmission and plasticity, which are important for various neurological processes such as learning and memory.
Automatically generated - may contain errors
Lab products found in correlation
Pstat3, by Bioss Antibodies (6 mentions)
Interleukin 6 (il 6), by Bioss Antibodies (6 mentions)
Interleukin 6 (il 6), by Abcam (6 mentions)
Gapdh, by Solarbio (3 mentions)
Gp130, by Bioss Antibodies (3 mentions)
Goat anti mouse igg, by Bioss Antibodies (3 mentions)
Gad67, by Bioss Antibodies (3 mentions)
Goat anti rabbit igg, by Bioss Antibodies (3 mentions)
Bca protein assay, by Beyotime (3 mentions)
Protease inhibitor cocktail, by Beyotime (3 mentions)
Gp130, by Abcam (3 mentions)
Gp130, by Santa Cruz Biotechnology (3 mentions)
Nmda receptors, by Abcam (3 mentions)
Gad67, by Abcam (3 mentions)
P stat3, by Abcam (3 mentions)
Polyvinylidene fluoride membrane, by Bio-Rad (2 mentions)
Polyvinylidene uoride membrane, by Bio-Rad (1 mentions)
6 protocols using nmda receptors
1
Protein Expression Analysis in Biological Samples
2
Western Blot Analysis of Protein Expression
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The tissue was homogenized in RIPA buffer containing a protease inhibitor cocktail (Beyotime Biotechnology).
A BCA protein assay (Beyotime Biotechnology) was used to determine the protein concentrations. Equal amounts of protein were separated by SDS-PAGE and then transferred electrophoretically to polyvinylidene fluoride membranes (Bio-Rad) [29, 30] . The membranes were incubated with the following primary antibodies for 2 hours at room temperature: IL-6 (1:1000, Abcam, England), Gp130 (1:1000, Santa, America), pSTAT3 (1:1000, Bioss, China), and NMDA receptors (1:1000, Bioss, China). The membranes were then incubated with GAPDH (1:1000, Solarbio, China), goat anti-mouse IgG (Bioss, China, Gp130, 1:1000 and GAD67, 1:1000) or goat anti-rabbit IgG (Bioss, China, pSTAT3, 1:1000, NMDA receptors, 1:3000 and IL-6, 1:1000) secondary antibodies for 2 hours at room temperature. Finally, the membranes were placed in a gel imaging analysis system for exposure and analysis (AlphaView FluorChem FC3).
A BCA protein assay (Beyotime Biotechnology) was used to determine the protein concentrations. Equal amounts of protein were separated by SDS-PAGE and then transferred electrophoretically to polyvinylidene fluoride membranes (Bio-Rad) [29, 30] . The membranes were incubated with the following primary antibodies for 2 hours at room temperature: IL-6 (1:1000, Abcam, England), Gp130 (1:1000, Santa, America), pSTAT3 (1:1000, Bioss, China), and NMDA receptors (1:1000, Bioss, China). The membranes were then incubated with GAPDH (1:1000, Solarbio, China), goat anti-mouse IgG (Bioss, China, Gp130, 1:1000 and GAD67, 1:1000) or goat anti-rabbit IgG (Bioss, China, pSTAT3, 1:1000, NMDA receptors, 1:3000 and IL-6, 1:1000) secondary antibodies for 2 hours at room temperature. Finally, the membranes were placed in a gel imaging analysis system for exposure and analysis (AlphaView FluorChem FC3).
3
Western Blot Analysis of Signaling Proteins
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The tissue was homogenized in RIPA buffer containing a protease inhibitor cocktail (Beyotime Biotechnology). A BCA protein assay (Beyotime Biotechnology) was used to determine the protein concentrations. Equal amounts of protein were separated by SDS-PAGE and then transferred electrophoretically to polyvinylidene uoride membranes (Bio-Rad) [29, 30] . The membranes were incubated with the following primary antibodies for 2 hours at room temperature: IL-6 (1:1000, Abcam, England), Gp130 (1:1000, Santa, America), pSTAT3 (1:1000, Bioss, China), and NMDA receptors (1:1000, Bioss, China). The membranes were then incubated with GAPDH (1:1000, Solarbio, China), goat antimouse IgG (Bioss, China, Gp130, 1:1000 and GAD67, 1:1000) or goat anti-rabbit IgG (Bioss, China, pSTAT3, 1:1000, NMDA receptors, 1:3000 and IL-6, 1:1000) secondary antibodies for 2 hours at room temperature. Finally, the membranes were placed in a gel imaging analysis system for exposure and analysis (AlphaView FluorChem FC3).
4
Immunohistochemical Analysis of PVN Markers
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After the brain tissue was embedded in paraffin, the part between the optic chiasm and mammillary body was resected in the rostro-caudal direction. The tissue was serially sectioned on a paraffin slicer, and sections that were approximately 1.50 mm from the bregma were obtained [25 (link), 26 (link)]. After dewaxing the slices, 3% H2O2 was used to block endogenous peroxidase activity, and 0.01 M citric acid was used to retrieve the antigens prior to antibody incubation. Then, the slices were incubated with primary antibodies overnight at 4 °C [27 , 28 (link)]. The sections were immunohistochemically labelled to identify IL-6 (Bioss, China, 1:100), Gp130 (Santa Cruz, America, 1:20), pSTAT3 (Bioss, China, 1:50), NMDA receptors (Bioss, China, 1:50), and GAD67 (Abcam, England, 1:2000) and then incubated with secondary antibodies (anti-mouse for Gp130 and GAD67; anti-rabbit for NMDA receptors, pSTAT3 and IL-6) for 20 min at room temperature. For each rat, the positive neurons within the bilateral borders of the PVN were manually counted in three consecutive sections, and the average value is reported.
5
Immunohistochemical Analysis of PVN Markers
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After the brain tissue was embedded in para n, the part between the optic chiasm and mammillary body was resected in the rostro-caudal direction. The tissue was serially sectioned on a para n slicer, and sections that were approximately 1.50 mm from the bregma were obtained [25, 26] . After dewaxing the slices, 3% H 2 O 2 was used to block endogenous peroxidase activity, and 0.01 M citric acid was used to retrieve the antigens prior to antibody incubation. Then, the slices were incubated with primary antibodies overnight at 4°C [27, 28] . The sections were immunohistochemically labelled to identify IL-6 (Bioss, China, 1:100), Gp130 (Santa Cruz, America, 1:20), pSTAT3 (Bioss, China, 1:50), NMDA receptors (Bioss, China, 1:50), and GAD67 (Abcam, England, 1:2000) and then incubated with secondary antibodies (anti-mouse for Gp130 and GAD67; anti-rabbit for NMDA receptors, pSTAT3 and IL-6) for 20 minutes at room temperature. For each rat, the positive neurons within the bilateral borders of the PVN were manually counted in three consecutive sections, and the average value is reported.
6
Immunohistochemical Analysis of PVN Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After the brain tissue was embedded in paraffin, the part between the optic chiasm and mammillary body was resected in the rostro-caudal direction. The tissue was serially sectioned on a paraffin slicer, and sections that were approximately 1.50 mm from the bregma were obtained [25, 26] . After dewaxing the slices, 3% H 2 O 2 was used to block endogenous peroxidase activity, and 0.01 M citric acid was used to retrieve the antigens prior to antibody incubation. Then, the slices were incubated with primary antibodies overnight at 4°C [27, 28] . The sections were immunohistochemically labelled to identify IL-6 (Bioss, China, 1:100), Gp130 (Santa Cruz, America, 1:20), pSTAT3 (Bioss, China, 1:50), NMDA receptors (Bioss, China, 1:50), and GAD67 (Abcam, England, 1:2000) and then incubated with secondary antibodies (anti-mouse for Gp130 and GAD67; anti-rabbit for NMDA receptors, pSTAT3 and IL-6) for 20 minutes at room temperature. For each rat, the positive neurons within the bilateral borders of the PVN were manually counted in three consecutive sections, and the average value is reported.
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