Rabbit anti src

Manufactured by Cell Signaling Technology

Sourced in United Kingdom, United States

Rabbit anti-Src is a primary antibody produced in rabbit that specifically binds to the Src protein. Src is a non-receptor tyrosine kinase that plays a key role in signal transduction pathways involved in cell growth, proliferation, differentiation, and survival. This antibody can be used to detect and study the Src protein in various experimental systems.

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Anti-Src (73 citations) Rabbit anti-phospho-Src (6 citations) Rabbit anti-phospho-Src (Tyr416, (3 citations) Rabbit polyclonal anti-Src antibody, (2 citations) Anti-Src rabbit monoclonal (2 citations) Rabbit anti-p-Src (2 citations) Anti-phospho-Src (Tyr 416) rabbit polyclonal (2 citations) Rabbit anti p-Src (Tyr416) (2 citations)

16 protocols using rabbit anti src

EV Protein Level Analysis via Western Blotting

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For analysis of protein levels in EVs, isolated EVs were lysed with RIPA buffer, followed by addition of 1X Laemlli buffer and incubation at 95°C for 5 min. Samples were subject to standard Western blotting with chemiluminescence detection. The following antibodies were used as recommended by the manufacturer: rabbit anti‐Src (Cat#: 2109, 1:1000), rabbit anti‐calnexin (Cat#: 2679, 1:5000), rabbit anti‐CD‐9 (Cat#: 13403 for human species, 1:1000), rabbit anti‐GAPDH (Cat#: 2118, 1:10,000), rabbit anti‐gamma‐tubulin (Cat#: 5886, 1:1000), and mouse anti‐Cas9 (Cat#: 14697, 1:1000) were all purchased from Cell Signaling Technology. Mouse anti‐VSV‐G (Cat#: EB0010, Kerafast, 1:1000), rabbit anti‐syntenin (Cat#: ab19903, Abcam, 1:1000), mouse anti‐CD63 (Cat#: 556019, BD Pharmingen, 1:500). Secondary antibodies anti‐rabbit IgG HRP (Cat#: 7074, 1:5000), anti‐mouse IgG HRP (Cat# 7076, Cell Signaling Technology, 1:5000). The anti‐myristoylated octapeptide antibody described in this study was used at dilutions of 1:250, 1:500, or 1:1000. Cas9 recombinant protein was purchased from Sigma Aldrich (Cat# Cas9Prot). EV protein analysis was done in accordance with the MISEV 2018 guidelines (Théry et al., 2018 (link)). The band intensity was quantified by Image J software.

Whitley J.A., Kim S., Lou L., Ye C., Alsaidan O.A., Sulejmani E., Cai J., Desrochers E.G., Beharry Z., Rickman C.B., Klingeborn M., Liu Y., Xie Z, & Cai H. (2022). Encapsulating Cas9 into extracellular vesicles by protein myristoylation. Journal of Extracellular Vesicles, 11(4), e12196.

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Western Blot Analysis of Metabolic Enzymes

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Mouse tissues were homogenized and lysed by using RIPA buffer (Santa Cruz Biotechnology). Protein contents were measured by using the Bradford assay (Pierce Biotechnology). Detailed steps for western blot analysis were as previously described^{59 (link)}. The primary antibodies included goat anti-KYNU, rabbit anti-KMO, mouse anti-KAT1, mouse anti-Cyclin B1, mouse anti-GAPDH (all above antibodies from Santa Cruz Biotechnology); mouse anti-YAP1, mouse anti-phospho-YAP (ser 127), rabbit anti-Erk1/2 (Thr202/Tyr204), rabbit anti-Erk1/2, rabbit anti-c-Myc, rabbit anti-phospho-Src (Tyr527) and rabbit anti-Src (all seven antibodies from Cell Signaling). The relative protein levels were calculated by normalizing to GAPDH protein as a loading reference by using ImageJ.

Zhang D., Ning J., Ramprasath T., Yu C., Zheng X., Song P., Xie Z, & Zou M.H. (2022). Kynurenine promotes neonatal heart regeneration by stimulating cardiomyocyte proliferation and cardiac angiogenesis. Nature Communications, 13, 6371.

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Oncogenic Src Kinase Inhibition and Drug Sensitivity

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3, 3′-dithiobis sulfosuccinimidylpropionate (DTSSP) and beta-D-Lactose were purchased from Thermo Scientific (Pittsburgh, PA). Sucrose was purchased from MP Biomedicals (Solon, OH). Ouabain octahydrate, cis-diammineplatinum dichloride (CDDP), doxorubicin hydrochloride (DXR) and thiazolyl blue tetrazolium bromide (MTT) were purchased from Sigma-Aldrich. Pyrazolo pyrimidine (PP2), a Src kinase inhibitor, was purchased from Tocris Bioscience (Bristol, UK).
Customized polyclonal rabbit anti-Gal-3 antibody was created by Pierce Biotechnology; mouse anti-V5 and purified mouse IgG were purchased from Invitrogen; polyclonal goat anti-Na⁺/K⁺-ATPase alpha1 and monoclonal mouse anti-Mdr-1 were purchased from Santa Cruz Biotechnology (Santa Cruz, CA); polyclonal rabbit anti-Mdr was purchased from Oncogene Research Products (Cambridge, UK); mouse anti-beta-actin was purchased from Sigma-Aldrich; rabbit anti-Src and anti-p-Src were purchased from Cell Signaling Technology (Beverly, MA); purified rabbit IgG was purchased from ZYMED (San Francisco, CA); monoclonal mouse anti-phosphoserine was purchased from abcam (Cambridge, MA).

Harazono Y., Kho D.H., Balan V., Nakajima K., Hogan V, & Raz A. (2015). Extracellular galectin-3 programs multidrug resistance through Na+/K+-ATPase and P-glycoprotein signaling. Oncotarget, 6(23), 19592-19604.

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Tissue Lysis and Protein Extraction

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Tissues were washed once in 500 μl of ice-cold Bio-Rad BioPlex Cell Wash Buffer, cut into 3×3 mm pieces, and homogenized in 500 μl of ice-cold Bio-Rad BioPlex Cell Lysis Buffer using an IKA T10 Basic Ultra-Turrax homogenizer (dispersing element S10N-5G) for 30 s at minimum speed and 15 s at maximum speed. Homogenates were frozen at −80°C to complete the lysis, thawed, and centrifuged at 4°C, 8000 rcf, for 20 min, followed by isolation of the soluble layer. Protein concentrations were measured using Bio-Rad DC Protein Assay on NanoDrop 2000. The following primary antibodies were used for immunoblotting: 1:10,000 rabbit anti-PCNA (Novus Biologicals), 1:2000 rabbit anti-Src (Cell Signaling Technology), 1:500 goat anti-Grb2 (R&D Systems), 1:1000 rabbit anti-ERK1/2 (R&D Systems), 1:2000 rabbit anti-IER3 (Novus Biologicals), 1:5000 rabbit anti-KRAS (Novus Biologicals), 1:1000 rabbit-anti-HoxA1 (Abcam).

Brock A., Krause S., Li H., Kowalski M., Goldberg M.S., Collins J.J, & Ingber D.E. (2014). Prevention of mammary tumor progression by silencing HoxA1 via intraductal injection of nanoparticle-formulated siRNA. Science translational medicine, 6(217), 217ra2.

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Striatal Protein Analysis Post-Collagenase ICH

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A 4-mm coronal section containing the striatum was collected at 24 h after collagenase-induced ICH, as described previously (Chang et al., 2014 (link)). Twenty-microgram protein samples were separated by 4–12% SDS-PAGE and transferred onto polyvinylidene difluoride membranes. The membranes were blocked and probed with the following primary antibodies: rabbit anti-cleaved caspase 3 (1:1000; Cell Signaling, Danvers, MA), rabbit anti-caspase 3 (1:1000; Cell Signaling), mouse anti-nitrotyrosine (1:40,000; Millipore), rabbit anti-Src (1:1000; Cell Signaling), rabbit anti-phospho-Src (Tyr416, 1:1000; Cell Signaling), and β-actin (1:5000; Santa Cruz Biotechnology, Santa Cruz, CA). Resulting protein bands were scanned and analyzed with ImageJ software (version 1.42q, NIH). Optical density values were normalized to the corresponding loading control intensity on each gel and were expressed as fold change over values from sham-operated mice.

Zhao X., Wu T., Chang C.F., Wu H., Han X., Li Q., Gao Y., Li Q., Hou Z., Maruyama T., Zhang J, & Wang J. (2015). Toxic Role of Prostaglandin E2 Receptor Ep1 After Intracerebral Hemorrhage in Mice. Brain, behavior, and immunity, 46, 293-310.

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Western Blot Analysis of Signaling Proteins

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Proteins obtained as described previously^{58 (link)} were blotted onto polyvinylidene fluoride membrane (Calbiochem/Merck Millipore) and incubated with following antibodies: rabbit anti-NTSR-1 (ANT-015), rabbit anti-NTSR-2 (ANT-016), Alomone Labs; rabbit anti-Bcl-2 (sc-783, Santa Cruz Biotechnology, Dallas, TX, USA); rabbit anti-Bcl-xL (#2764S), rabbit anti-phospho-Akt (Ser473) (#4060S), mouse anti-Akt (pan; #2920S), rabbit anti-phospho-Src family (Tyr416; #6943S), rabbit anti-Src (#2108S), rabbit anti-phospho-p38 MAPK (Thr180/Tyr182; #9211S), rabbit anti-p38 MAPK (#8690S), rabbit anti-phospho-SAPK/JNK (Thr183/Tyr185; #4668S), and rabbit anti-SAPK/JNK (#9252), all from Cell Signaling Technology (Danvers, MA, USA); mouse anti-Giα1/2 antibody (06-236, Calbiochem/Merck Millipore); and anti-actin (A5441, Sigma-Aldrich). After washing (tris-buffered saline/0.1% Tween-20), the immunoreactions were detected by incubation for 2 h at RT with horseradish peroxidase-conjugated secondary Ab against mouse or rabbit Ig (P0447 and P0448, respectively, Agilent Technologies, Santa Clara, CA, USA), revealed with the Immobilon Western Chemiluminescent HRP Substrate (Merck Millipore). Western blot were detected using Bioimaging Systems (GeneSnap and GeneTool; Syngene, Cambridge, UK). Densitometric analyses were performed using the ImageJ software (NIH, Bethesda, MD, USA).

Abbaci A., Talbot H., Saada S., Gachard N., Abraham J., Jaccard A., Bordessoule D., Fauchais A.L., Naves T, & Jauberteau M.O. (2017). Neurotensin receptor type 2 protects B-cell chronic lymphocytic leukemia cells from apoptosis. Oncogene, 37(6), 756-767.

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Antibody-Based Protein Expression Analysis

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The detailed protocol has been published[24 (link)]. Primary antibodies used were rabbit anti-phospho-AKT (Ser473), rabbit anti-AKT, rabbit anti-phospho-FAK (Tyr397), rabbit anti-FAK, rabbit anti-eIF2α, rabbit anti-phospho-MEK1/2 (Ser217/221), rabbit anti-MEK1/2, rabbit anti-phospho-p38-MAPK (Thr180/Tyr182), rabbit anti-p38-MAPK, rabbit anti-phospho-Src (Tyr416), rabbit anti-Src (1:1000, Cell Signaling), goat anti-phospho-eIF2α (Ser51), rabbit anti-MMP2 (1:1000, Abgent), mouse anti-MMP9 (1:500, Abgent), rabbit anti-tubulin (1:1000, BioLegend). Secondary antibodies used were goat anti-mouse, goat anti-rabbit or donkey anti-goat IgG HRP conjugates (1:5000, Santa Cruz Biotechnology), accordingly. Relative levels from three independent experiments were presented as mean ± SEM.

Marín-Ramos N.I., Thein T.Z., Cho H.Y., Swenson S.D., Wang W., Schönthal A.H., Chen T.C, & Hofman F.M. (2018). NEO212 Inhibits Migration and Invasion of Glioma Stem Cells. Molecular cancer therapeutics, 17(3), 625-637.

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Molecular Profiling of Hemorrhagic Stroke

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Brain tissue from the whole hemorrhagic hemisphere was obtained 24 hours after ICH. Twenty-microgram protein samples were separated by 4–12% sodium dodecyl sulfate polyacrylamide gel electrophoresis and transferred onto polyvinylidene difluoride membranes. The membranes were probed with the following primary antibodies: rabbit anti-HMGB1 (1:500, Abcam, Cambridge, MA), rabbit anti-Src (1:1000, Cell Signaling Technology, Danvers, MA), rabbit anti-phospho-Src (Tyr416, 1:1000, Cell Signaling), rabbit anti-cyclooxygenase (COX)-2 (1:100, Cayman Chemical, Ann Arbor, MI), rabbit anti-interleukin (IL)-1β (1:200, Abcam), and rabbit anti-β-actin (1:5000, Abcam). Resulting protein bands were scanned and analyzed with ImageJ software. Data were expressed as fold change over the loading control. We examined five brains per treatment group and three brains per sham group.

Wu H., Wu T., Hua W., Dong X., Gao Y., Zhao X., Chen W., Cao W., Yang Q., Qi J., Zhou J, & Wang J. (2015). PGE2 receptor agonist misoprostol protects brain against intracerebral hemorrhage in mice. Neurobiology of aging, 36(3), 1439-1450.

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Antibody Panel for PEDV Detection

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Rabbit anti-TfR1 (Abcam). Mouse monoclonal anti-p-Tyr (PY20) (Santa Cruz Biotechnology). Rabbit anti-Src (Cell Signaling Technology). Rabbit anti-pAPN (ABclonal Biotechnology). Mouse monoclonal antibodies to His, HA, and GFP (CMCTAG, Milwaukee, USA). Mouse monoclonal anti-PEDV N protein was purchased from Medgene labs (FACS, 1:100; IF, 1:200; Western-blot, 1:1000). FITC-conjugated anti-PEDV polyclonal antibody was purchased from VMRD (1:200, PC-IFA-PEDV). Dylight 488 goat anti-mouse IgG (H+L) and Dylight 649 goat anti-rabbit IgG (H+L) were from MultiSciences (Lianke) Biotech, CO., LTD. Rabbit monoclonal anti-GAPDH and goat anti-rabbit IgG-HRP were from Bioworld Technology Inc (St. Louis Park, MN, USA). HRP-conjugated goat anti-mouse IgG (H+L) (Vazyme, Nanjing, China). Anti-rabbit IgG antibody (Beyotime).

Zhang S., Cao Y, & Yang Q. (2020). Transferrin receptor 1 levels at the cell surface influence the susceptibility of newborn piglets to PEDV infection. PLoS Pathogens, 16(7), e1008682.

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Immunoblotting and Immunoprecipitation Protocols

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Immunoblotting was performed as described (Jayabal et al., 2017 (link)). Immunoprecipitation was performed as described (Shiio and Eisenman, 2003 (link)). The following antibodies were used: rabbit anti-FLI-1 (ab15289, Abcam); rabbit anti-GDF6 (Novus, NBP 1-91934); rabbit anti-caspase 3 (9665, Cell Signaling Technology); rabbit anti-PARP (sc-7150, Santa Cruz Biotechnology); mouse anti-CD99 (MS-1633, Lab Vision); sheep anti-CD99L2 (AF5185, R&D Systems); mouse anti-p21 (BD PharMingen, 556430); mouse anti-tubulin (Developmental Studies Hybridoma Bank); rabbit anti-GAPDH (sc-25778, Santa Cruz Biotechnology); rabbit anti-CSK (HPA028425, Atlas Antibodies); mouse anti-GST (MS-707-P0, Lab Vision); mouse anti-GFP (sc-9996, Santa Cruz Biotechnology); rabbit anti-actin (8457, Cell Signaling Technology); rabbit anti-Src (2123, Cell Signaling Technology); rabbit anti-phospho-Src (2101, Cell Signaling Technology); rabbit anti-phospho-Cortactin (4569, Cell Signaling Technology); rabbit anti-phospho-p130 Cas (4011, Cell Signaling Technology); rabbit anti-phospho-STAT3 (9145, Cell Signaling Technology); rabbit anti-phospho-Smad1/5 (9516, Cell Signaling Technology); rabbit anti-HA (3724, Cell Signaling Technology); and mouse anti-FLAG (F1804, Sigma-Aldrich).

Zhou F., Elzi D.J., Jayabal P., Ma X., Chiu Y.C., Chen Y., Blackman B., Weintraub S.T., Houghton P.J, & Shiio Y. (2020). GDF6-CD99 Signaling Regulates Src and Ewing Sarcoma Growth. Cell reports, 33(5), 108332.

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