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Socs3

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SOCS3 is a laboratory equipment product manufactured by Thermo Fisher Scientific. It is a protein that functions as a negative regulator of cytokine signaling. The core function of SOCS3 is to inhibit the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, which is involved in the regulation of various cellular processes.

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Lab products found in correlation

High capacity cdna reverse transcription kit, by Thermo Fisher Scientific (2 mentions) Lipofectamine rnaimax reagent, by Thermo Fisher Scientific (1 mentions) Socs1, by Santa Cruz Biotechnology (1 mentions) Opti mem, by Thermo Fisher Scientific (1 mentions) Stat3, by Cell Signaling Technology (1 mentions) Ripa lysis buffer, by Beyotime (1 mentions) Taqman real time pcr master mix, by Thermo Fisher Scientific (1 mentions) Interleukin 6 (il 6), by Thermo Fisher Scientific (1 mentions) Mmp 2, by Thermo Fisher Scientific (1 mentions) Mmp 9, by Thermo Fisher Scientific (1 mentions) Rneasy mini micro kit, by Qiagen (1 mentions) Viia 7 real time pcr system, by Thermo Fisher Scientific (1 mentions) Sh ptp1, by Santa Cruz Biotechnology (1 mentions) Ptyr705 stat3, by Cell Signaling Technology (1 mentions) Socs3, by Cell Signaling Technology (1 mentions) P jak2, by Thermo Fisher Scientific (1 mentions) Pepck, by Thermo Fisher Scientific (1 mentions) Stat3, by R&D Systems (1 mentions) Pser727 stat3, by Cell Signaling Technology (1 mentions) Sybr green chemistry, by Thermo Fisher Scientific (1 mentions)

Spelling variants of this product

SOCS3 siRNA (4 citations) SOCS3 Mm01249143 (2 citations)

8 protocols using socs3

1

Dendritic Cell Silencing for Infection Studies

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DCs were transfected with small interfering RNAs (siRNAs) targeting SOCS1 (Santa Cruz Biotechnology, Inc.), SOCS2 (ThermoFisher), SOCS3 (ThermoFisher) or Allstars negative control (Qiagen). For transfection, Lipofectamine RNAiMAX reagent (Life Technologies) was used according to the manufacturer’s instructions. In brief, 2 × 105 DCs were seeded in 100 μl DC medium and transfected with 100 μl Opti-MEM (Life Technologies) containing 50 to 100 pmol siRNA and 1 μl transfection reagent. Subsequently, DCs were incubated for 6 h before 800 μl of fresh DC medium was added. After 48 h of incubation, DCs were used for infection experiments after silencing efficiency was assessed by qRT-PCR.
Sarajlic M., Neuper T., Vetter J., Schaller S., Klicznik M.M., Gratz I.K., Wessler S., Posselt G, & Horejs-Hoeck J. (2020). H. pylori modulates DC functions via T4SS/TNFα/p38-dependent SOCS3 expression. Cell Communication and Signaling : CCS, 18, 160.
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2

Western Blot Analysis of Lung Proteins

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Total protein from lung tissues was extracted using RIPA lysis buffer (Beyotime Biotechnology, Shanghai, China). Extracted proteins were separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and transferred to polyvinylidene difluoride (PVDF) membranes. The relative expression of proteins was determined using an ECL detection system (Li et al., 2021 (link)). The primary antibodies were those against GR (Cell Signaling Technology, Danvers, MA, United States), p-GR (Ser211; Cell Signaling Technology), SOCS3 (Thermo Scientific), and STAT3 (124H6; Cell Signaling Technology).
Xue Y., Zhou Y., Bao W., Fu Q., Hao H., Han L., Zhang X., Tian X, & Zhang M. (2021). STAT3 and IL-6 Contribute to Corticosteroid Resistance in an OVA and Ozone-induced Asthma Model with Neutrophil Infiltration. Frontiers in Molecular Biosciences, 8, 717962.
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3

RNA Extraction and qRT-PCR Analysis

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RNA extraction from whole tissues and FACS-purified macrophages was performed using RNeasy Mini/Micro kits (Qiagen) according to the manufacturer's instructions. RNA (2 μg) was used to generate cDNA with the High Capacity cDNA Reverse Transcription Kit (Applied Biosystems) according to the manufacturer's instructions. Quantitative RT-PCR analysis was performed on duplicate samples with TaqMan Real-Time PCR Master mix (Life Technologies) using the Viia7 Real-Time PCR System (Life Technologies) for 40 cycles (95°C for 15 seconds, 60°C/1 minute) and following an initial holding stage (50°C/2 minutes, 95°C/10 minutes). 18S or Gapdh were used as housekeepers and fold changes in gene expression were obtained using the 2−ΔΔCt method (13 (link)).
TaqMan probes used were mouse 18s (Mm04277571_s1), Gapdh (Mm99999915_g1), Il6 (Mm00446190_m1), Il11 (Mm00434162_m1), Socs3 (Mm00545913_s1), Mmp2 (Mm00439498_m1), Mmp9 (Mm00442991_m1), Il10 (Mm01288386_m1), Arg1 (Mm00475988_m1), and Vegfα (Mm00437306_m1) from Thermo Fisher Scientific.
Poh A.R., Dwyer A.R., Eissmann M.F., Chand A.L., Baloyan D., Boon L., Murrey M.W., Whitehead L., O'Brien M., Lowell C.A., Putoczki T.L., Pixley F.J., O'Donoghue R.J, & Ernst M. (2020). Inhibition of the SRC Kinase HCK Impairs STAT3-Dependent Gastric Tumor Growth in Mice. Cancer immunology research, 8(4), 428-435.
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4

Immunoblotting of Hepatic Signaling

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Western blots for immunocomplexes and hepatic homogenates were performed using antibodies against phosphorylated (p) Tyr705-signal transducer and activator of transcription factor 3 (pTyr705-STAT3), pSer727-STAT3 and SOCS3 from Cell Signaling Technology (Danvers, MA), pTyr1007/1008 Janus kinase 2 (pJAK2), JAK2 and regulatory subunit of PI3K (p85) from Millipore (Temecula, CA), STAT3 from R&D Systems (Minneapolis, MN) and aquaglyceroporin-9 (AQP9), the beta chain of insulin receptor (IRb), catalytic subunit of PI3K (p110), GLUT2, long form of the leptin receptor (OBeRb), phosphoenolpyruvate carboxykinase (PEPCK) and SH-PTP1 from Santa Cruz Biotechnology (Santa Cruz, CA). The proteins were detected by chemiluminiscence using an ECL system. Quantification of the bands was carried out by densitometry using a Kodak Gel Logic 1500 Image Analysis system and Molecular Imaging software 4.0 (Rochester, NY, USA). AQP9, GLUT2, IRb, OB-Rb, p85, PEPCK, SOCS3 and SH-PTP1 were normalized with actin (Thermo Scientific, Fremont, CA), whereas pJAK2, pTyr705-STAT3 and p-Ser727-STAT3 were normalized with their respective total forms.
Burgos-Ramos E., Canelles S., Rodríguez A., Gómez-Ambrosi J., Frago L.M., Chowen J.A., Frühbeck G., Argente J, & Barrios V. (2015). Chronic central leptin infusion modulates the glycemia response to insulin administration in male rats through regulation of hepatic glucose metabolism. Molecular and cellular endocrinology, 415.
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5

Tocilizumab Modulates IL-6 Response in Monocytes

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Human monocytes from healthy donors were pre-incubated with or without 0.5 mg/mL tocilizumab and then incubated with or without recombinant human IL-6 (0.1ug/ml), normal donor plasma, or patient plasma for an additional 30 minutes in IMDM supplemented with 1.0% human A/B serum. Tocilizumab (Actemra™, Genentech) was purchased through the Hospital of the University of Pennsylvania Pharmacy. Following stimulation, culture medium was removed, cells were lysed directly in TRIzol, and RNA was isolated using a Qiagen RNeasy Kit per manufacturer instructions. cDNA was synthesized from 0.5 – 1.0 μg of RNA per sample (Applied Biosystems, Foster City, CA), and primers for qRT-PCR were designed using the Primer 3 online program (25 (link), 26 (link)) and synthesized by Integrated DNA Technologies (SOCS3) or purchased from Applied Biosystems (GAPDH). Relative quantification was measured using either SYBR Green chemistry (Applied Biosystems; SOCS3) or Taqman chemistry (Applied Biosystems; GAPDH). SOCS3 expression was normalized to GAPDH and relative expression was calculated using the ΔCT formula. The fold increase or decrease in expression of treated samples relative to no treatment controls was calculated (ΔΔCT). Primer sequences for human SOCS3 are as follows: (i) Forward: 5′-CAAGGACGGAGACTTCGATT-3′, (ii) Reverse: 5′-AACTTGCTGTGGGTGACCAT-3′.
Long K.B., Tooker G., Tooker E., Luque S.L., Lee J.W., Pan X, & Beatty G.L. (2017). IL-6 receptor blockade enhances chemotherapy efficacy in pancreatic ductal adenocarcinoma. Molecular cancer therapeutics, 16(9), 1898-1908.
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6

Systematic Gene Expression Analysis in Muscle Tissue

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RNA was extracted from TA and diaphragm muscles using TRIzol as previously described [16 (link)]. Isolated total RNA was subsequently purified using an RNeasy Mini kit (Qiagen, Valencia, CA), according to manufacturer’s instructions. The resulting quantity and purity of total RNA was tested through absorbance spectrophotometry at 230, 260 and 280 nm, and the quality of RNA was tested on a 1% denaturing agarose gel. Synthesis of cDNA and qRT-PCR analyses from RNA isolated from the TA and diaphragm were performed as described previously [28 (link)] using a 7300 real-time PCR system and the following primers from Applied Biosystems (Austin, TX): Fbxo30 (NM_027968.3), Fbxo31 (NM_133765.4), Bach2 (NM_001109661.1), Socs3 (NM_007707.3), Ubr2 (NM_146078.3), Psma2 (NM_008944.2), Ubqln1 (NM_026842.4), Fos (NM_010234.2), Cebpb (NM_009883.3), Stat3 (NM_011486.4), Col6a2 (NM_146007.2), Myoz3 (NM_133363.3), atrogin-1/MAFbx/Fbxo32 (NM_026346.2), MuRF1/Trim63 (NM_001039048.2), Bcl3 (NM_033601.3), and Maff (NM_010755.3).
Judge S.M., Wu C.L., Beharry A.W., Roberts B.M., Ferreira L.F., Kandarian S.C, & Judge A.R. (2014). Genome-wide identification of FoxO-dependent gene networks in skeletal muscle during C26 cancer cachexia. BMC Cancer, 14, 997.
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7

Quantitative Gene and miRNA Expression

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Total RNA was extracted from cells with TRIzol Reagent (Invitrogen, Calsbad, CA) following the manufacturer’s protocol. cDNA was synthesized with High Capacity cDNA Reverse Transcription Kit or MicroRNA Reverse Transcription Kit (Applied Biosystems, Foster City, CA) according to the manufacturer’s instructions. Quantitative real-time PCR was performed by Prism 7300 Real Time PCR System (Applied Biosystems) using TaqMan Gene Expression Master Mix (Applied Biosystems) in a final reaction volume of 20 µl with each primer. The following human primers were purchased from Applied Biosystems: 18S (Hs 99999901_sl), tp53 (Hs01034249_m1), Bcl-2 (Hs00236808_s1), BAX (Hs00180269_m1), Atg5 (Hs00169468_m1), Beclin-1 (Hs00387943_m1), PPARγ (Hs00234592_m1), SOCS3 (Hs02330328_s1), U6 snRNA (001973), hsa-miR-203b-3p (464535-mat), hsa-miR-34a-3p (002316), and hsa-miR-21-3p (002438). A negative control without cDNA did not produce any amplicons. Data were analyzed with 2−ΔΔCt value calculation using 18S or U6 RNA for normalization.
Li P., Guo Y., Bledsoe G., Yang Z., Chao L, & Chao J. (2016). Kallistatin induces breast cancer cell apoptosis and autophagy by modulating Wnt signaling and microRNA synthesis. Experimental cell research, 340(2), 305-314.
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8

Cortical Gene Expression in TBI Mice

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Around 5 mm tissue of ipsilateral cortex around the site of injury from TBI mice (1 day, 3 day and 7 day post-TBI) or the corresponding tissue of same volume around the same cortical region from sham mice were dissected and processed as described23 (link). Total RNA isolated using miRNeasy Mini Kit (Qiagen, Cat No. 217004) was converted into cDNA using High Capacity RNA to cDNA kit (Applied Biosystem, Cat. No. 4387406) as per manufacturer's instruction. cDNA TaqMan Universal Master Mix II (Applied Biosystems, Cat. No. 4440040) was used to perform quantitative real-time PCR amplification as described previously23 (link) using 20 × TaqMan® Gene Expression Assay (Applied Biosystems) for the following mouse genes: Gapdh (Mm99999915_g1), Nlrp3 (Mm00840904_m1), Cybb (Mm01287743_m1), Nos2 (Mm00440502_m1), Tnf (Mm00443258_m1), Ifnb1 (Mm00439552_s1), Il1b (Mm00434228_m1), Arg1 (Mm00475988_m1), Socs3 (Mm00545913_s1), Chil3 (Mm00657889_mH) and Il4r (Mm01275139_m1) (Applied Biosystems). Reactions were amplified and quantified by using a 7900HT Fast Real-Time PCR System and corresponding software (Applied Biosystems). Relative gene expression normalized to Gapdh was calculated based on the comparative Ct method45 (link). For this study n = 5/group for all time points.
Hegdekar N., Lipinski M.M, & Sarkar C. (2021). N-Acetyl-l-leucine improves functional recovery and attenuates cortical cell death and neuroinflammation after traumatic brain injury in mice. Scientific Reports, 11, 9249.
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