Hippocampal MF and CF proteins from 8-week-old male C57BL/6 mice treated with ketamine for 2 h, 14 h, 24 h and 72 h were fractionated by SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot was performed based on standard protocols. After electrophoresis, proteins were transferred to PVDF membranes (Immobilon-P, Millipore, Billerica, USA). Primary antibodies were against adenosinmonophosphate-activated protein kinase (AMPK, Abcam, Cambridge, UK), phosphorylated AMPK (pAMPK, Cell Signaling, Merck, Darmstadt, Germany), peroxiredoxin 1 (Prdx1, Abcam, Cambridge, UK), peroxiredoxin 3 (Prdx3, Abcam, Cambridge, UK). Anti-rabbit, anti-mouse and anti-goat ECL horseradish peroxidase-linked secondary antibodies (GE Healthcare Life Sciences, Little Chalfont, Buckinghamshire, UK) were used. The densitometric analyses were performed with the Image Lab software (Bio-Rad, Munich, Germany).
Prdx3
Manufactured by Abcam
Sourced in United Kingdom
Prdx3 is a protein that is a member of the peroxiredoxin family. Peroxiredoxins are involved in the reduction of peroxides. Prdx3 is found in the mitochondria and plays a role in protecting cells from oxidative stress.
Automatically generated - may contain errors
Lab products found in correlation
Immobilon p, by Merck Group (1 mentions)
Peroxiredoxin 1, by Abcam (1 mentions)
Prdx1, by Abcam (1 mentions)
Image lab software, by Bio-Rad (1 mentions)
Phospho p44 42 mapk erk1 2, by Cell Signaling Technology (1 mentions)
Mouse anti mouse β actin, by Cell Signaling Technology (1 mentions)
Pvdf membrane, by Bio-Rad (1 mentions)
Chemidoc, by Bio-Rad (1 mentions)
Trxr1, by Cell Signaling Technology (1 mentions)
β actin, by Thermo Fisher Scientific (1 mentions)
Hsp90, by Cell Signaling Technology (1 mentions)
α tubulin, by Santa Cruz Biotechnology (1 mentions)
Prdx1, by Cell Signaling Technology (1 mentions)
Tomm20, by Cell Signaling Technology (1 mentions)
Sirt3, by Cell Signaling Technology (1 mentions)
Cleaved caspase 3, by Beyotime (1 mentions)
Gel pro analyzer version 4, by Media Cybernetics (1 mentions)
β actin, by Beyotime (1 mentions)
Enhanced chemiluminescence kit, by Cytiva (1 mentions)
Glyceraldehyde 3 phosphate dehydrogenase, by Proteintech (1 mentions)
5 protocols using prdx3
1
Ketamine Regulates AMPK and Peroxiredoxins in Hippocampus
2
Oxidative Stress Protein Expression in PFC Homogenates
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Equal protein concentrations of PFC homogenates were separated on 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and then transferred to PVDF membranes (Bio-Rad Laboratories, Hercules, CA). Membranes were blocked in 5% skim milk in 1X Tris-buffered saline with Tween 20 (TBST) for 1 h at room temperature. Membranes were then incubated overnight (4 °C) with either mouse anti-mouse OxPhos antibody cocktail (Thermo Fisher 45–8099, 1:200), mouse anti-mouse β-actin (Cell Signaling 4970, 1:1000), rabbit anti-mouse (mAb) SOD-2 (Cell Signaling 13141, 1:1000), rabbit mAb HO-1 (Cell Signaling 86806, 1:1000), rabbit mAb PRDX-3 (Abcam ab73349, 1:1000), rabbit mAb GPX-4 (Cell Signaling 52455, 1:1000), Bcl-1 rabbit mAb Cell Signaling 42406, 1:1000), p62 rabbit mAb Cell Signaling 23214, 1:1000), BDNF (Cell Signaling 47808, 1:1000), p-TrkB (Abcam ab229908, 1:1000), p44/42 MAPK (Erk1/2) (Cell Signaling L34F12, 1:1000), Phospho-p44/42 MAPK (Erk1/2) (Cell Signaling D13.14.4E, 1:1000). After washing three times with 1X TBST, the membranes were incubated in a secondary antibody goat anti-rabbit IgG or goat anti-mouse (Bio-Rad 1706515 and 1706516) in 1X TBST for 1 h. Blots were subsequently developed with an enhanced chemiluminescence (ECL) system (Bio-Rad 1705061) and visualized using a Chemi-Doc (Bio-Rad) imaging device.
3
Protein Expression Analysis in Cells
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The following antibodies were used: NRF2 (Cell Signaling Technologies, D1Z9C, Cat #12721), NQO1 (Sigma Aldrich, Cat #HPA007308), β-actin (Thermo Fisher, clone AC-15, Cat #A5441), α-tubulin (Santa Cruz, TU-02, Cat #sc-8035), SOD2 (Cell Signaling Technologies, Cat #13194S), Prdx3 (Abcam, Cat #ab73349), Prdx1 (Cell Signaling Technologies, D5G12, Cat# 50-191-580), HSP90 (Cell Signaling Technologies, Cat #4874S), ME1 (Thermo Fisher Scientific, Cat #PA5-21550), IDH1(Cell Signaling Technologies, Cat #8137S), SDHA (Cell Signaling Technologies, D6J9M, Cat #11998), GSR (Santa Cruz Biotechnology, Cat #sc-133245), TRXR1 (Cell Signaling Technologies, Cat #15140S), TRXR2 (Cell Signaling Technologies, Cat #12029S). Total Histone H2A.X (Cell Signaling Technologies, D17A3, Cat #9718), Gamma Histone H2A.X (Cell Signaling Technologies, 20E3, Cat #7631S), Aconitase 1 GeneTex, Cat #GTX128976), Aconitase 2 (GeneTex, Cat #GTX109736).
4
Western Blot Analysis of Mitochondrial Proteins
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Western blotting was conducted with primary antibodies against PRDX3 (Abcam, Ltd., Cambridge, UK), SIRT3 (Cell Signaling Technology, MA, USA), cleaved caspase-3 (Beyotime Institute of Biotechnology, Shanghai, China), TOMM20 (Cell Signaling Technology, MA, USA) and β-actin (Beyotime Institute of Biotechnology, Shanghai, China). Protein quantification was performed using Gel-Pro Analyzer version 4.0 (Media Cybernetics, MD, USA).
5
Western Blot Analysis of Apoptosis Markers
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Cells in each group were collected and lysed with RIPA buffer for 15 min on ice. After centrifugation, the supernatant was taken for quantification, and protein electrophoresis buffer was added for denaturation at 100°C for 10 min. A 20 μg denatured protein was separated on a 10% sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to a polyvinylidene fluoride membrane. After blocking with 5% skimmed milk in Tris-buffered saline-Tween 20 (TBST) for 1 h at room temperature, the primary antibody and the internal reference protein glyceraldehyde 3-phosphate dehydrogenase (Proteintech) were added overnight at 4°C, respectively. The primary antibodies were PRDX3 (Abcam), cleaved caspase-3 (Affinity), and Bax (ABclonal). After washing with TBST, the membranes were incubated with antirabbit (ABclonal) or antimouse (ABclonal) secondary antibodies for 2 h at room temperature and developed using an enhanced chemiluminescence kit (Amersham). The protein band gray values were analyzed using ImageJ.
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