Fluorescein‐conjugated dextran (D3306, molecular weight: 3 kDa, Dex‐3) was purchased from Invitrogen. Gd‐DPTA (molecular weight: 938 Da) was obtained from CONSUN. HiLyte Fluor‐488‐labeled recombinant human α‐syn (AS‐55457, molecular weight: 14 kDa HiLyte‐488‐α‐syn) was purchased from Anaspec. Adeno‐associated virus expressing A53T‐α‐syn (AAV‐A53T) was designed by OBio Technology. Primary antibody anti‐AQP4 (rabbit, AB3594), anti‐α‐syn (mouse, 36–008; Syn211), and anti‐tyrosine hydroxylase (rabbit, AB152; anti‐TH) were purchased from Millipore. Anti‐PDGF‐B (rabbit, DF6328) was purchased from Affinity Biosciences, and anti‐PDGFRb (mouse, sc‐374573) was purchased from Santa Cruz Biotechnology. β‐actin (mouse, ab008‐100), horseradish peroxidase‐conjugated anti‐rabbit and anti‐mouse IgG (70‐gam007, 70‐gar007) were purchased from Multisciences. Secondary antibodies, including Cy3‐conjugated donkey anti‐rabbit and Cy5‐conjugated donkey anti‐rabbit, were purchased from Jackson ImmunoResearch. DNaseI and 4′,6‐diamidine‐2′‐phenylindole dihydrochloride (DAPI) were purchased from Sigma‐Aldrich. RNAiso Plus (9109), PrimeScrip RT Master Mix (RR036A) and SYBR Premix Ex Taq II (RR820A) were purchased from TaKaRa.
β actin
Manufactured by MultiSciences Biotech
Sourced in China, United States
β-actin is a highly conserved cytoskeletal protein that is ubiquitously expressed in all eukaryotic cells. It is a key component of the cytoskeleton and plays a crucial role in various cellular processes, such as cell motility, structure, and integrity.
Automatically generated - may contain errors
Lab products found in correlation
Bcl2 associated x (bax), by Santa Cruz Biotechnology (2 mentions)
E cadherin, by Proteintech (2 mentions)
N cadherin, by Proteintech (2 mentions)
Vimentin, by Proteintech (2 mentions)
Pvdf membrane, by Merck Group (2 mentions)
Hrp conjugated secondary antibody, by MultiSciences Biotech (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
4 6 diamidine 2 phenylindole dihydrochloride dapi, by Merck Group (1 mentions)
Primescrip rt master mix, by Takara Bio (1 mentions)
Ab3594, by Merck Group (1 mentions)
Cy5 conjugated donkey anti rabbit, by Jackson ImmunoResearch (1 mentions)
Rnaiso plus 9109, by Takara Bio (1 mentions)
Anti pdgfrb, by Santa Cruz Biotechnology (1 mentions)
Sybr premix ex taq 2, by Takara Bio (1 mentions)
Dnase 1, by Merck Group (1 mentions)
Cy3 conjugated donkey anti rabbit, by Jackson ImmunoResearch (1 mentions)
Protein marker, by Thermo Fisher Scientific (1 mentions)
Isoflurane, by Merck Group (1 mentions)
Uvp biospectrum imaging system, by Analytik Jena (1 mentions)
Polyvinylidene difluoride pvdf membrane, by Merck Group (1 mentions)
14 protocols using β actin
1
Fluorescent Markers for Neurodegeneration
2
Protein Detection and Analysis Protocol
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Isoflurane, Tween 20, and 2,3,5-trihenylterzolium chloride (TTC) were purchased from Sigma-Aldrich (St. Louis, USA). Coated monofilament was purchased from Doccol Corporation (Sharon, USA). Chloral hydrate was purchased from Sinopharm Chemical Reagent Co. (Shanghai, China). Protein marker was purchased from Fermentas (Waltham, USA). Polyvinylidene difluoride (PVDF) membrane was purchased from Merck Millipore (Burlington, USA). Primary antibodies of SPK1 (sc-22702) and SPK2 (sc-22704) were purchased from Santa Cruz Biotechnology (Santa Cruz, USA). β-actin and IgG secondary antibody were purchased from Multi Sciences (Hangzhou, China). EZ-ECL kit was purchased from Biological Industries (Beit HaEmek, Israel); PH Indicator was purchased from Leici Device Works (Shanghai, China). A 5417R High-speed Freezing Centrifuge was purchased from Eppendorf (Hamburg, Germany). A 752 Ultraviolet Spectrophotometer was purchased from Puxi General Instruments (Beijing, China). A DYY-4C electrophoresis system was purchased from Tanon Science & Technology (Shanghai, China). A UVP BioSpectrum Imaging System was purchased from UVP (Upland, USA).
3
Western Blotting Analysis of Cellular Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
After the indicated treatment, the cells were harvested and lysed with ice-cold lysis buffer containing protease inhibitor cocktail (Roche, Mannheim, Germany). The protein concentration of the lysates was determined by a Protein Assay Kit (Thermo, Rockford, Illinois, USA), and equal amounts of protein were separated on a 10% sodium dodecyl sulfate–polyacrylamide electrophoresis gel and transferred to a nitrocellulose membrane. Nonspecific protein binding was blocked with 5% bovine serum albumin (BSA) for 1 h at room temperature. The membranes were then incubated with specific AMPKα (1:1000), phospho-AMPKα (p-AMPK; Thr172; 1:1000), p62 (1:1000), poly (ADP-ribose) polymerase (PARP), cleaved PARP (1:1000, Cell Signaling Technology, Massachusetts, USA), LC3 (1:1000), phospho-p70 S6 kinase (Thr389; 1:500, Abcam, Massachusetts, USA), cytochrome c (Cyt-c; 1:1000), Bcl-2 (1:1000), Bax (1:1000, Santa Cruz, State of California, USA), and β-actin (1:2000; MultiSciences, Hangzhou, China) primary antibodies overnight at 4 °C. The membranes were washed the next day and incubated with secondary antibodies at room temperature for 1 h. After being washed with phosphate-buffered saline (PBS) three times, the membranes were incubated with secondary antibodies. The immunoblots were detected by the Odyssey Infrared Imaging System, and the data were quantified with ImageJ software.
4
Western Blot Analysis of TLR4 Expression
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total protein was extracted by high performance liquid chromatography into radioimmunoprecipitation assay (RIPA) buffer with added phenylmethanesulfonyl fluoride (PMSF). The total protein concentration was determined using a bicinchoninic acid (BCA) assay. The proteins were separated by electrophoresis using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), transferred to polyvinylidene fluoride (PVDF) membranes, and blocked using 5% skim milk for 1 h. The membranes were then incubated with primary antibody overnight at 4°C. The primary antibodies used included those recognizing: TLR4 (1:1000) and β-actin (1:3000) (Multisciences Biotech Co., Hangzhou, China). After overnight incubation, the membranes were rinsed with TBST buffer (Tris buffer, NaCl, and Tween 20) and incubated with the corresponding secondary antibodies (1:2000) for 1 h at room temperature. Membranes were scanned using the ChemiDoc XRS+ imaging system (Bio-Rad, Hercules, CA, USA), and the gray values of the immunoreactive protein bands were analyzed using ImageJ 7.0 software (NIH, Bethesda, MD, USA). Relative protein expression levels were calculated in comparison with the level of β-actin.
5
Western Blot Analysis of Endoplasmic Reticulum Stress
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells and tissues were lysed in radioimmunoprecipitation assay (RIPA) lysis buffer (Solarbio, Beijing, China). Samples were then cleared by centrifugation (12,000 rpm, 15 min, 4 °C). Equal amount of proteins (10–20 μg) were separated by SDS-PAGE and transferred to polyvinylidene difluoride (PVDF) membranes (Millipore, Billerica, MA, USA). The following primary antibodies were used: iPLA2β (Santa Cruz Biotechnology, Dallas, TX, USA), GRP78 (Santa Cruz Biotechnology, Dallas, TX, USA), calnexin (Abclonal, Wuhan, China), p-IRE1α, ATF6 (Proteintech, Rosemont, IL, USA), ATF4 (Proteintech, Rosemont, IL, USA), p-PERK (Abclonal, Wuhan China), p-eIF2α (Abclonal, Wuhan, China), CHOP, cleaved caspase-3, Bax, p-CaMKII, SERCA2 (Santa Cruz Biotechnology, Dallas, TX, USA), β-actin (MultiSciences, Hangzhou, China), PERK (Abclonal, Wuhan, China), eIF2α (Abclonal, Wuhan, China), CaMKII and GAPDH (MultiSciences, Hangzhou, China). After incubation with the corresponding secondary antibodies conjugated with horseradish peroxidase (HRP) (MultiScience, Hangzhou, China), proteins were detected using a BioRad ChemiDoc MP Imaging system with enhanced chemiluminescence. All other antibodies were purchased from Cell Signaling Technology (Beverly, MA, USA).
6
Western Blot Analysis of SSADH Protein
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cells were lysed with RIPA lysis buffer (Beyotime, Beijing, China) containing 1% PMSF (Beyotime, Beijing, China). Protein (15 μg) was loaded on 10% SDS-PAGE gels for electrophoresis and transferred to PVDF membranes. The membranes were blocked with 5% non-fat dry milk in TBS containing 0.1% Tween 20 for 1.5 h at room temperature and then incubated with antibody for SSADH (AffinitY, Cat# DF12820) and β-actin (Multi Sciences, Hangzhou, China) at 4 °C overnight. The membranes were then incubated with HRP-conjugated secondary antibody (Multi Sciences, Hangzhou, China) for 2 h at room temperature. Signals were detected using film by darkroom exposure (Servicebio, G2019, China). In each experiment, each sample run on the SDS-PAGE gel in duplicate.
7
Detecting NF-κB Expression by Western Blot
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blot analysis was used to detect NF-κB. A PVDF membrane was obtained from EMD Millipore (Billerica, MA, USA); cat. no. IPVH00010; batch no. K5JA5013L). Anti-NF-kB p65 antibody was obtained from Abcam (Cambridge, MA, USA; cat. no. ab7970). β-actin was purchased from Multi Sciences (Lianke) Biotech, Co., Ltd. (cat. no. ab008-100). Goat anti-rabbit IgG was bought from Multi Sciences (Lianke) Biotech, Co., Ltd. (cat. no. GAR0072). Goat anti-mouse IgG was obtained from Bioworld Technology, Inc. (St. Louis Park, MN, USA; cat. no. BS12478). An electrophoresis system (Mini-Proten Tetra system) was purchased from Bio-Rad Laboratories, Inc. (Hercules, CA, USA). A gel imager was obtained from Bio-Rad Laboratories, Inc. (ChemiDoc XRS+ system).
8
Western Blot Analysis of EMT Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting analysis was conducted as described previously in our study (16 (link)). Total cellular proteins were harvested in IP lysis buffer (Beyotime, Shanghai, China) with a cocktail of proteinase and phosphatase inhibitors (Bimake.cn) and were measured by a BCA protein assay kit (Thermo Fisher Scientific, USA). Protein lysates were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transferred to nitrocellulose (NC) membrane (Millipore, USA). Then the membranes were blocked with 5% skimmed milk in Tris-Buffered saline (TBS) for 1 hour at RT and incubated with primary antibodies overnight at 4°C followed by incubation with DyLight fluorescent dye labelled secondary antibodies for 1 hour at RT. Finally, immunoblot signals were detected using an Odyssey Imaging System (LI-COR Biosciences, USA). Primary antibodies used in this analysis were listed as follows: TRIM50 (1:100 dilution, Abcam, ab174880), E-cadherin (1:5000, Proteintech, 20874-1-AP), N-cadherin (1:2000, Proteintech, 22018-1-AP), Vimentin (1:2000, Proteintech, 10366-1-AP), Snail1 (1:1000, Proteintech, 13099-1-AP), Snail2 (1:2000, Proteintech, 12129-1-AP), Twist1 (1:1000, Proteintech, 25465-1-AP), Twist2 (1:1000, Proteintech, 11752-1-AP), ZEB1 (1:1000, Proteintech, 21544-1-AP), ZEB2 (1:2000, Proteintech, 14026-1-AP), and β-actin (1:5000, MultiSciences, ab008).
9
KIFC1 Protein Expression Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Testicular tissue was homogenized in radio-immunoprecipitation assay lysis buffer (Solarbio, Shanghai, China) containing protease inhibitors. The lysate was centrifuged at 12,000 rpm for 20 min at 4 °C. After removal of the supernatant, 1× loading buffer was added to the sample. Protein concentration was measured using a bicinchoninic acid protein assay kit (Qiagen) according to the manufacturer’s instructions. Approximately 30 μg of protein was loaded on each gel and electrotransferred to polyvinylidene difluoride membranes using standard procedures. KIFC1 was detected using a rabbit polyclonal antibody to KIFC1 (1:200; Proteintech, Chicago, IL, USA), followed by a mouse anti-rabbit secondary antibody (1:3000; MultiSciencesBiotech, Chicago, IL, USA). Band intensity was quantified relative to that of β-actin (MultiSciencesBiotech) using Image Quant TL 7.0 software (GE Healthcare, Little Chalfont, UK).
10
Western Blotting Analysis of Glycogen Metabolism
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting analysis was performed according to the established protocols as previously described 35 (link). The primary antibodies used in this study were as follows: PYGL (1:1,000, 15851-1-AP), GYS1 (1:2,000, 10566-1-AP), GBE1 (1:3,000, 20313-1-AP), PYGM (1:5,000, 19716-1-AP), PYGB (1:1,000, 12075-1-AP), E-cadherin (1:1,000, 20874-1-AP), Occludin (1:2,000, 13409-1-AP), N-cadherin (1:2,000, 22018-1-AP), and Vimentin (1:2,000, 10366-1-AP) from Proteintech, and β-actin (1:5,000, ab008) from MultiSciences.
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!