The tissue samples were sliced at a thickness of 4 μm. The sections were incubated with primary antibody (STAT3, surviving, VEGF (all from Proteintech Group, USA) and cyclin D1 (Santa Cruz Biotechnology, USA)) overnight, followed by a biotin-labelled secondary antibody. The evaluation of immunostaining included both positivity rate and staining intensity of cells [22 (link)]. The proportion of positive cells was graded as follows: 0 for no staining, 1 for < 30%, 2 for 30–60%, and 3 for > 60%. The grading of staining density was 0 for no staining, 1 for light brown, 2 for brown, and 3 for dark brown. All specimen sections were analysed independently by two researchers with the double-blind method. The final score was obtained from the product of the percentage score and the staining intensity score. The final result was evaluated as positive when the score was ≥ 3 or negative when the score was < 3. The cholesteatoma tissues, control skin, and HaCaT cells were lysed with RIPA lysis buffer. Then, the proteins were separated and transferred to a polyvinylidene fluoride (PVDF) membrane (Millipore, MA, USA), which was incubated with primary antibody (STAT3, surviving, VEGF, β-actin (all from Proteintech Group, USA) and cyclin D1 (Santa Cruz Biotechnology, USA)). The enhanced chemiluminescence ECL detection system (Thermo Fisher Scientific, MA) was used to present the results.
Stat3
Manufactured by Proteintech
Sourced in United States, China
STAT3 is a recombinant protein that functions as a transcription factor. It is involved in cellular processes such as cell growth and differentiation.
Automatically generated - may contain errors
Lab products found in correlation
Gapdh, by Proteintech (13 mentions)
Pvdf membrane, by Merck Group (12 mentions)
P stat3, by Cell Signaling Technology (6 mentions)
P stat3, by Abcam (6 mentions)
P stat3, by Proteintech (6 mentions)
β actin, by Proteintech (6 mentions)
α sma, by Proteintech (5 mentions)
Erk1 2, by Proteintech (4 mentions)
β actin, by Merck Group (4 mentions)
P jak2, by Abcam (4 mentions)
E cadherin, by Proteintech (4 mentions)
N cadherin, by Proteintech (4 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (4 mentions)
Cyclin b1, by Proteintech (3 mentions)
Cyclin d1, by Proteintech (3 mentions)
Ecl reagent, by Keygen Biotech (3 mentions)
Vimentin, by Proteintech (3 mentions)
Bcl 2, by Proteintech (3 mentions)
P stat3, by Boster Bio (3 mentions)
Phosphatase inhibitor, by Aspen (2 mentions)
45 protocols using stat3
1
Immunohistochemical Analysis of Cholesteatoma
2
Galectin-3BP Signaling Pathway Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Beta-Lapachone (ARQ-501, LPC) was purchased from Selleck Chemicals Inc. RIPA buffer solution, TRIzol reagent, RIPA buffer solution and endotoxin-free plasmid small extraction medium volume extraction kit were obtained from Sparkjade Biotech (Shandong, China) AceQ qPCR SYBR Green Master Mix was purchased from Vazyme (Nanjing, China), and First-strand cDNA Synthesis super Mix kit was from TransGen Biotech (Beijing, China). HE staining kit was obtained from Beyotime Biotechnology (Hangzhou, China). EdU incorporation assay kit was obtained from Keygen Biotech (shanghai, China). Fetal Bovine Serum was purchased from Sigma-Aldrich (St. Louis, MO, United States). VECTASTAIN ABC Kit was purchased from Vector Laboratories. Human Galectin-3BP ELISA Kit was purchased from Abcam (Cambridge, MA, United States). Human IL6, IL6R ELISA kit were purchased from Boster Biological Technology (Nanjing, China). Primary antibodies we used were shown as following: ERK1/2 and β-Actin were purchased from abway. Gal-3, pSTAT3 and pERK1/2 were purchased from Cell Signaling Technology. Gal-3BP (LGALS3BP) was purchased from R&D Systems (Minneapolis, MN). STAT3 was purchased from Proteintech Group.
3
Molecular Mechanisms of JAK/STAT Signaling
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The following antibodies were used: anti-CEP55 (#23891-1-AP), MMP2 (#66366-1-Ig), MMP9 (#10375-2-AP), STAT3 (#60199-1-Ig), JAK1 (#66466-1-Ig), JAK2 (#17670-1-AP), and Actin (#60008-1-Ig) (Proteintech Group, Chicago, IL, USA); and p-JAK1 Tyr1034/1035 (#74129), p-JAK2 Tyr1007/1008 (#3771), and p-STAT3 Tyr705 (#9145) (Cell Signaling Technology, Beverly, MA, USA). The biological reagent IL-6 was acquired from Sigma Chemical (St. Louis, MO, USA). JAK2 inhibitor XL019 (#S7036), STAT3 inhibitor C188-9 (#S8605) was from Selleck (Shanghai, China).
4
Myricetin modulates autophagy pathways
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Myricetin (#HY-15097) was purchased from MedChemExpress (New Jersey, United States), dissolved in dimethyl sulfoxide (DMSO, #D8370, Solarbio, Beijing, China) at a concentration of 100 mM, and stored at −20 °C. MG132 (#HY-13259) and BafA1 (#HY-100558) were also bought from MedChemExpress and dissolved in DMSO. All antibodies were as follows: anti-Bcl-2 (#12789-1-AP), Ki-67 (#27309-1-AP), Stat3 (#10253-2-AP), LC3 (#14600-1-AP), P62 (#18420-1-AP), CyclinB1 (#55004-1-AP), CyclinD1 (#26939-1-AP), GAPDH (#10494-1-AP), and peroxidase-conjugated AffiniPure goat anti-rabbit IgG (H + L) (#SA00001-2) (Proteintech Group, Chicago, IL, United States); MARCH1 (#bs-9335R, Bioss, Beijing, China); p-Stat3 (#ab32143), p38 MAPK (#ab170099) (abcam, Cambridge, United Kingdom); p-p38 MAPK (#11581, Singalway Antibody, Maryland, United States); and MARCH1 (#YT2642, Immunoway, Newark, United States).
5
Western Blot Analysis of Signaling Pathways
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lung extracts were prepared in radioimmunoprecipitation assay (RIPA) cell lysis buffer (Solarbio, Beijing, China), and equal amounts of protein were separated on a 12% sodium dodecyl sulfate–polyacrylamide electrophoresis gel, electroblotted on nitrocellulose membranes, and probed with specific antibodies against JAK (ProteinTech Group, Wuhan, China, 1:1000), STAT3 (ProteinTech Group, Wuhan, China, 1:2000), Akt (ProteinTech Group, Wuhan, China, 1:1000), mTOR (ProteinTech Group, Wuhan, China, 1:1000), NF-κB (ProteinTech Group, Wuhan, China, 1:1000), cyclin D1 (ProteinTech Group, Wuhan, China, 1:2000) and GAPDH (ProteinTech Group, Wuhan, China, 1:2000). The secondary antibody used was Peroxidase-conjugated Affinipure Goat anti-rabbit IgG (H + L) (ProteinTech Group, 1:5000). Antibody binding was detected by enhanced chemiluminescence according to the manufacturer's protocol (Pierce, Rockford, IL). Band densities were quantified using Image software (NIH, Bethesda, MD, USA) and normalized according to the reference gene GAPDH. All values were then normalized to relative GAPDH expression.
6
Multiprotein Immunoblot Analysis
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Total cell protein lysate was made in RIPA buffer with proteinase and phosphatase inhibitor cocktails, resolved by SDS‐PAGE, and transferred onto an Immobilon‐PSQ Transfer Membrane, as previously described.20 Blots were incubated with antibodies against GAPDH (1:5000, Proteintech), VASN (2 μg/mL, R&D Systems), STAT3 (1:2000, Proteintech), p‐STAT3 (1:1000, Cell Signaling Technology), Notch1 (1:1000, 20687‐1‐AP, Proteintech), NICD (1:500, Cell Signaling Technology) and VEGF (1:1000, PeproTech) overnight at 4°C. All immunoblots were performed at least 3 times.
7
Gypenoside-Cisplatin Combination Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Gypenosides (GYP) were provided by our own laboratory (Qi et al., 2021 (link)); Cisplatin was purchased from TargetMol (United States). β-actin (Zsgb-Bio, TA-09, 1:1000), STAT3 (Proteintech, 60199-1-Ig, 1:2000), TYMS (Proteintech, 15047-1-AP, 1:1000), MAPK14 (Proteintech, 66234-1-Ig, 1:2000), EGFR (Proteintech, 66455-1-Ig, 1:10000).
8
Protein Profiling and Inhibition in Cell Line Study
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The antibodies and dilutions used were as follows: ERα (ab32063: immunohistochemistry, 1:150; immunoblotting, 1:750) from Abcam; GAPDH (CW0101: immunoblotting, 1:1000) from CWBIOTECH; PTEN (ab170941: immunoblotting, 1:1000) from Abcam; SFRS1 (12929‐2‐ap: immunoblotting, 1:750, RIP: 4 µg) from Proteintech; RBMX (ab190352: immunoblotting, 1:1000) from Abcam; p‐STAT3 (ab76315: immunoblotting, 1:2000) from Abcam; FAP (sc‐65398: immunofluorescence, 1:100) from Santa Cruz; α‐SMA (55135‐1‐ap: immunofluorescence, 1:50) from Proteintech; STAT3 (10253‐2‐ap: immunoblotting, 1:1000, ChIP: 4 µg) from Proteintech; CD63 (561925: flow cytometry, per test 20 µL, human) from BD bioscience; CD63 (143903: flow cytometry, per test 0.5 µg, mouse) from Biolegend; F‐actin (40735ES75: immunofluorescence, 1:100) from YEASEN.
The inhibitors were as follows: STAT3 inhibitors (HY‐15146: 30 × 10−6m for the in vitro assay) from MedChem Express; GW4869 (HY‐19363: 20 × 10−6m for the in vitro assay) from MedChem Express; JAK inhibitor baricitinib (HY‐15315: 50 × 10−6m for the in vitro assay) from MedChem Express.
The inhibitors were as follows: STAT3 inhibitors (HY‐15146: 30 × 10−6
9
Western Blot Analysis of Stem Cell Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Western blotting was used to detect the target protein in the sample. The total protein content in the sample was detected in a 96-well plate using a BCA protein concentration determination kit (Dingguochangsheng). Protein tracer sample buffer (reduction, 5×; CWBIO) was mixed with protein samples in a ratio of 1:4. The mixtures were then placed in a boiling water bath for 3 minutes. The samples were cooled to room temperature and centrifuged at 13 000×g at 4 °C for 30 seconds. Denatured proteins were directly loaded to a sodium dodecyl sulfate-page gel, and conventional electrophoresis (concentration gel voltage 60 V, separation gel voltage 120 V) and membrane transfer (current 200 mA) were performed using a Bio-Rad protein imprinting device. After completion, the transferred films were incubated at 4 °C overnight. The rabbit primary antibodies used were STAT3 (Proteintech), pSTAT3 (Tyr705; ABclonal), KLF4 (Proteintech), OCT4 (Proteintech), and β-actin (Proteintech). Samples were incubated with HRP-labeled goat anti-rabbit secondary antibody (Proteintech) at room temperature for 1 hour, washed with 1× TBST 3 times for 10 minutes each, and finally ECL development was performed.
10
Western Blot Analysis of Cardiac Markers
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Heart tissues or cells (NMVMs and CFs) were homogenized and lysed in cold RIPA lysis buffer (Thermo Fisher Scientific, USA) containing a protease and Phosphatase inhibitor cocktail (Beyotime, USA). Total protein for each sample was quantified by BCA Protein Assay Kit. Proteins were separated by SDS-PAGE and transferred to PVDF membranes (Millipore, USA). Western blot were performed by blocking the membrane with 5% skimmed milk, followed overnight incubation at 4° with indicated primary antibodies against Cad-11 (Thermo Fisher Scientific), Collagen I (Abclonal), α-SMA (Abcam), CTGF (CST), p-ERK1/2 (CST), ERK1/2 (CST), p-JNK (CST), JNK (CST), p-P38 (CST), P38 (CST), p-CaMKII (CST), CaMKII (CST), p-STAT3 (CST), STAT3 (CST), GAPDH (Proteintech). After rigorous washing, the membranes were incubated with horseradish peroxidase-conjugated anti-rabbit/mouse secondary antibodies and visualized on a Gel Imaging System (Tanon, China). The band intensity was quantified by ImageJ software (1.51) after normalization to their respective GAPDH loading control.
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!