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Ab25117

Manufactured by Abcam
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Sourced in United Kingdom, United States

Ab25117 is a laboratory equipment product. It is a high-quality tool designed for use in scientific research and experiments. The core function of this product is to provide a reliable and precise measurement or analysis capability, but further details on its intended use are not available.

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Lab products found in correlation

Ab124824, by Abcam (3 mentions) Enhanced chemiluminescence kit, by Cytiva (1 mentions) Ripa lysis buffer, by Santa Cruz Biotechnology (1 mentions) Ab28364, by Abcam (1 mentions) Oct compound, by Sakura Finetek (1 mentions) Ab197203, by Abcam (1 mentions) Ab104224, by Abcam (1 mentions) Ab1211, by Abcam (1 mentions) Ab10062, by Abcam (1 mentions) Ab6721, by Abcam (1 mentions) Bca protein assay kit, by Beyotime (1 mentions) Ab9485, by Abcam (1 mentions) Ab92486, by Abcam (1 mentions) Timp 3 ab39184, by Abcam (1 mentions) Ab178847, by Abcam (1 mentions) Iba 1, by Abcam (1 mentions) Pecam 1, by Santa Cruz Biotechnology (1 mentions) Ab36861, by Abcam (1 mentions) Ab3697, by Abcam (1 mentions) Ab41037, by Abcam (1 mentions)

11 protocols using ab25117

1

Cochlea Protein Analysis Protocol

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After decapitation of the animals, the bulla was quickly removed and placed in ice-cold 10 mM PBS. The bony capsule and the lateral wall of the cochlea were removed. Cochlea sample included the organ of corti, and the modiolus was dissected. Specimen was homogenized in a RIPA lysis buffer (sc-24948, Santa Cruz Biotechnology). Homogenate was centrifuged, and supernatant solution was collected. Total protein amount was determined according to the Bradford method (Bradford, 1976) using bovine serum albumin as a protein concentration standard. Protein was separated using SDS gel and transferred into a nitrocellulose membrane. After blocking, the membrane was incubated with primary antibodies for rabbit polyclonal to SDF-1 (AB 25117, 1:1,000; Abcam, Cambridge, UK) and β-actin (AB, ab8227, 1:2,000; Abcam) (as an internal control) at 4°C overnight. After rinsing to eliminate unbound primary antibody, the membrane was incubated with horseradish peroxidase-conjugated anti-rabbit IgG secondary antibody (AB, 6721, 1:2,000; Abcam) for 2 hours at room temperature. The protein bands were visualized using an enhanced chemiluminescence kit (Amersham Pharmacia Biotech Inc., Piscataway, NJ, USA) and quantified by TotalLab software (Wales, UK) (Niknazar et al., 2016, 2017).
Peyvandi A.A., Roozbahany N.A., Peyvandi H., Abbaszadeh H.A., Majdinasab N., Faridan M, & Niknazar S. (2018). Critical role of SDF-1/CXCR4 signaling pathway in stem cell homing in the deafened rat cochlea after acoustic trauma. Neural Regeneration Research, 13(1), 154-160.
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2

Evaluating Wound Vascularization and Healing

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Upon wound closure, mice were euthanized and wounds were harvested with an 8 mm biopsy punch (Integra Miltex, Plainsboro, New Jersey). Skin tissues were fixed in 4% paraformaldehyde overnight followed by serial dehydration in ethanol and embedding in paraffin. In order to assess dermal thickness, 5-μm sections were then stained for H&E and photographed at 10X magnification.
Immunofluorescence staining for CD31 and SDF-1 was then performed to assess wound vascularization. Frozen tissue samples were prepared by OCT Compound (Sakura Finetek US, Torrance, California) and cryosectioned for immunohistochemistry.27 Slides underwent antigen retrieval using 0.01M sodium citrate buffer in PBS solution, followed by blocking for 1 hour with 5% goat serum in PBS. Sections were then incubated in CD31 antibody (ab28364; Abcam, Cambridge, Massachusetts) or SDF-1 antibody (ab25117; Abcam, Cambridge, Massachusetts) overnight, followed by a secondary antibody. Lastly, slides were DAPI-stained for cell nuclei and mounted. ImageJ was used to binarize immunofluorescent images taken with the same excitation, gain, and exposure settings.28 (link) Intensity threshold values were set automatically and quantification of staining was determined by pixel-positive area normalized to dermal area. All histology was conducted in quadruplicate.
WHITTAM A.J., MAAN Z.N., DUSCHER D., BARRERA J.A., HU M.S., FISCHER L.H., KHONG S., KWON S.H., WONG V.W., WALMSLEY G.G., GIACCO F., JANUSZYK M., BROWNLEE M., LONGAKER M.T, & GURTNER G.C. (2018). Small molecule inhibition of dipeptidyl peptidase-4 enhances bone marrow progenitor cell function and angiogenesis in diabetic wounds. Translational research : the journal of laboratory and clinical medicine, 205, 51-63.
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3

Recombinant CXCL12 Peptide and Inhibitors

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Rat recombinant CXCL12 peptide (Z02860) was purchased from Genscript (Piscataway, NJ, USA). CXCR4 antagonists AMD3100 (A5602) and Astrocyte metabolic inhibitor fluorocitrate (F9634) were purchased from Sigma (St. Louis, MO, USA). Anti‐CXCL12 neutralizing antibody (ab25117) and control IgG (ab133470) antibody were purchased from Abcam (Cambridge, MA, USA). AMD3100, Anti‐CXCL12 neutralizing antibody and control IgG were dissolved in sterilized saline. CXCL12 peptide and fluorocitrate were prepared in 1% dimethyl sulfoxide (DMSO).
Antibodies against GFAP (ab10062), CXCL12 (ab25117),14 CXCR4 (ab197203),15 NeuN (ab104224), and OX42 (ab1211) were purchased from Abcam. Antibodies against CXCR4 (11073‐2‐AP)16 were purchased from Proteintech (San Ying biotechnology, Wuhan, China). Antibody against Actin (sc‐8432) was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA). Anti‐mouse IgG (H + L) (#4408) and anti‐Rabbit IgG (H + L) (#4413) were purchased from Cell Signaling Technology (CST) (Danvers, MA, USA).
Liu Z., Song Z., Guo S., He J., Wang S., Zhu J., Yang H, & Liu J. (2019). CXCL12/CXCR4 signaling contributes to neuropathic pain via central sensitization mechanisms in a rat spinal nerve ligation model. CNS Neuroscience & Therapeutics, 25(9), 922-936.
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4

Protein Extraction and Western Blotting

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A protein extraction kit (Beyotime, P0013J) was used to extract the total protein from rat skin ulcer tissues according to the manufacturer’s protocol. Prior to centrifugation at 12,000g for 15 min at 4 °C, TissueLyser beads were added to the centrifugation tubes to assist with protein cleavage. The upper layer of lipid was removed, and a BCA Protein Assay Kit (Beyotime, China) was using to determine the protein concentration in each extract. Western blotting was performed according to a standard WB protocol (Yukhananov, Chimento & Marlow, 2022 (link)). ImageJ software (v 1.8.0) was used to quantify protein levels, and GAPDH expression served as an internal standard. The primary antibodies used for WB were as follows: anti-CXCR4 (ab124824, 1:100; Abcam), anti-SDF1 (ab25117, 1:1,000; Abcam), and anti-GAPDH (ab9485, 1:2500; Abcam). The secondary antibody was goat anti-rabbit IgG H&L (HRP) (ab6721, 1:2000; Abcam).
Ou S., Wu X., Yang Y., Xia C., Zhang W., Qu Y., Li J., Chen B., Zhu L., Xu C, & Qi Y. (2023). Tibial cortex transverse transport potentiates diabetic wound healing via activation of SDF-1/CXCR4 signaling. PeerJ, 11, e15894.
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5

Osteoarthritis Treatment Protocol

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AMD3100 (ab120718, Purity: > 99%), LY303511 (ab145193) and primary antibodies for CXCL12 (ab25117), CXCR4 (ab124824), aggrecan (ab36861) ADAMTS-5 (ab41037), Smad3(ab52903), TIMP-3 (ab39184) and TGF-β1 (ab92486) were purchased from Abcam (Cambridge, UK), p-Akt (ab38449), p-Smad3 (ab122028) were purchased from Optomics (Hermiston, OR, USA).CXCL12a and CXCL12b ELISA kits were purchased from Tsz Biosciences (Boston, MA, USA). Alzet osmotic mini-pumps (2006) were purchased from DURECT Corporation (Cupertino, CA, USA). Recombinant rat CXCL12a, IL-1 and other reagents were purchased from Beyotime (Shanghai, China).
Lu W., He Z., Shi J., Wang Z., Wu W., Liu J., Kang H., Li F, & Liang S. (2020). AMD3100 Attenuates Post-Traumatic Osteoarthritis by Maintaining Transforming Growth Factor-β1-Induced Expression of Tissue Inhibitor of Metalloproteinase-3 via the Phosphatidylinositol 3-Kinase/Akt Pathway. Frontiers in Pharmacology, 10, 1554.
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6

Fixation and Cryosectioning of Posterior Eyecup

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After the anterior segments were removed, the posterior eyecup was fixed in 4% PFA overnight and treated in 10% sucrose for two hours before being incubated in 30% sucrose overnight. Eyecups were imbedded in optimal cutting temperature compound (OCT, Tissue Tek, USA) and cryosections were cut at 10 μm intervals. Cells grown on glass coverslips or tissue slices were fixed for 10 minutes at room temperature in 4% paraformaldehyde. PBS containing 1% bovine serum albumin and 0.2% Triton X-100 were added for 30 minutes. The following primary antibodies were used: Pecam1 (Santa Cruz, sc-46694), Iba1 (Abcam, ab178847), Sdf1 (Abcam, ab25117), Cxcr4 (Abcam, ab181020), Ki67 (CST, 9129). Then, the samples were stained with secondary antibodies for 1 hours at 37 °C).
Chen X., Wang X., Cui Z., Luo Q., Jiang Z., Huang Y., Jiang J., Qiu J., Li Y., Yu K, & Zhuang J. (2023). M1 Microglia-derived Exosomes Promote Activation of Resting Microglia and Amplifies Proangiogenic Effects through Irf1/miR-155-5p/Socs1 Axis in the Retina. International Journal of Biological Sciences, 19(6), 1791-1812.
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7

Osteoarthritis Molecular Mechanism Analysis

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AMD3100 (ab120718) and primary antibodies for CXCL12 (ab25117), CXCR4 (ab124824), ACAN (ab36861), SOX-9 (ab3697), RUNX-2 (ab76957), ADAMTS-4/5 (ab185722, ab41037), the aggrecan neo-epitope 374ARGSV (ab3773), PNCA (ab29), SB202190 (ab120638), U-0126 (ab120241), and C59 (ab142216) were purchased from Abcam (Cambridge, UK). Antibodies for ERK (#9102), phospho-ERK (#4377), JNK (#9258), phospho-JNK (#4668), p38 (#8690), phospho-p38 (#4511), p65 (#8242), phospho-p65 (#3033), IκBα (#4812), phospho-IκBα (#2859), β-catenin (#9582), GSK-3β (#12456), SURVIVIN (#2808), Cyclin D1 (#2978), and MMP-7 (#3301) were purchased from Cell Signaling Technology (Beverly, MA, USA). CXCL12a and CXCL12b ELISA kits were purchased from Tsz Biosciences (Boston, MA, USA). Alzet osmotic mini-pumps (2004) were purchased from DURECT Corporation (Cupertino, CA, USA). Recombinant rat CXCL12a and other reagents were purchased from Beyotime (Shanghai, China).
Lu W., Shi J., Zhang J., Lv Z., Guo F., Huang H., Zhu W, & Chen A. (2016). CXCL12/CXCR4 Axis Regulates Aggrecanase Activation and Cartilage Degradation in a Post-Traumatic Osteoarthritis Rat Model. International Journal of Molecular Sciences, 17(10), 1522.
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8

Western Blot Analysis of SDF-1 and CXCR4

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The proteins were extracted from heart samples using RIPA Lysis Buffer [Sangon Biotech (Shanghai), Co., Ltd.]. Western blotting was performed with anti-SDF-1 (ab25117, abcam), anti-CXCR4 (ab124827, abcam), and anti-GAPDH (Santa Cruz Biotechnology, Inc., Santa Cruz, CA, United States) primary antibodies and with relevant secondary anti-bodies (Santa Cruz Biotechnology, Inc.) as described previously (Chen et al., 2015 (link)).
Chen J., Wei J., Huang Y., Ma Y., Ni J., Li M., Zhu Y., Gao X, & Fan G. (2018). Danhong Injection Enhances the Therapeutic Efficacy of Mesenchymal Stem Cells in Myocardial Infarction by Promoting Angiogenesis. Frontiers in Physiology, 9, 991.
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9

Western Blot Analysis of Protein Expression

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Protein expression levels were analysed by Western blot analysis as described previously.15, 18 The primary antibodies used were as follows: anti‐SDF‐1α (ab25117; Abcam), anti‐fibronectin (F3648; Sigma‐Aldrich), anti‐α‐SMA (ab5694; Abcam), anti‐collagen I (ab34710; Abcam), anti‐vimentin (SAB1305447, Sigma‐Aldrich), anti‐CXCR4 (BA0747‐2; Boster), anti‐p‐GSK3β (ser 9) (5558; Cell Signaling Technology), anti‐β‐catenin (610 154; BD Transduction Laboratories), anti‐PAI‐1 (AF3828; R&D Systems), anti‐snail1 (ab180714; Abcam), anti‐MMP‐7 (104 658; GTX), anti‐E‐cadherin (3195; Cell Signaling Technology), anti‐active β‐catenin (19807s; Cell Signaling Technology), anti‐Kim1 (BA3537; Boster, Wuhan, China), anti‐α‐tubulin (RM2007, Ray Antibody Biotech, Beijing, China) and p‐STAT antibody sampler kit (9914; Cell Signaling Technology), p‐JAK family antibody sampler kit (97 999; Cell Signaling Technology), STAT antibody sampler kit (9939; Cell Signaling Technology) and anti‐GAPDH (RM2000, Ray Antibody Biotech, Beijing, China).
Liu Y., Feng Q., Miao J., Wu Q., Zhou S., Shen W., Feng Y., Hou F.F., Liu Y, & Zhou L. (2020). C‐X‐C motif chemokine receptor 4 aggravates renal fibrosis through activating JAK/STAT/GSK3β/β‐catenin pathway. Journal of Cellular and Molecular Medicine, 24(7), 3837-3855.
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10

Intrathecal Drug Delivery in Rat Pain Model

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Drugs were delivered intrathecally. The intrathecal catheterization was performed according to our previous method.22 (link) In brief, a polyethylene-10 (OD, 0.61 mm; ID, 0.28 mm) catheter was inserted into the rat’s subarachnoid space through L5–L6 intervertebral space, and the tip of the catheter was located at the L5 spinal segmental level. The CXCR4-specific antagonist AMD3100 and the NF-κB activation inhibitor PDTC were purchased from Sigma (St. Louis, USA) and freshly dissolved daily in normal sterile saline prior to use. Anti-CXCL12 neutralizing antibody (Abcam, ab25117) and anti-IgG antibody (for control) were purchased from Abcam (Massachusetts, USA) and diluted with sterile artificial cerebrospinal fluid containing 126.6 mM NaCl, 2.5 mM KCl, 2.0 mM MgCl2, and 1.3 mM CaCl2. The specific MEK (extracellular signal-regulated kinase (ERK)) inhibitor PD98059 was purchased from Sigma and was dissolved in sterile saline containing 10% DMSO. The intrathecal injection of drug was performed on 8:00 a.m. daily, the time which was 30 min before behavioral test. The doses of AMD3100 (5, 10, 20 µg/10 μl),9 (link),13 (link) anti-CXCL12 neutralizing antibody (4 µg/10 μl),8 (link) PDTC (0.5 µg/10 μl),23 (link),24 (link) and PD98059 (10 µg/10 μl)25 (link) used in this experiment were based on those used in previous studies.
Xing F., Kong C., Bai L., Qian J., Yuan J., Li Z., Zhang W, & Xu J.T. (2017). CXCL12/CXCR4 signaling-mediated ERK1/2 activation in spinal cord contributes to the pathogenesis of postsurgical pain in rats. Molecular Pain, 13, 1744806917718753.
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