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Rabbit anti tlr4

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Rabbit anti-TLR4 is a primary antibody that specifically recognizes the Toll-like receptor 4 (TLR4) protein. TLR4 is a member of the Toll-like receptor family and plays a crucial role in the innate immune response, functioning as a pattern recognition receptor that detects pathogen-associated molecular patterns.

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Lab products found in correlation

Mouse anti β actin, by Santa Cruz Biotechnology (2 mentions) Rabbit anti gpr30, by Abcam (2 mentions) Rabbit anti iba1, by Fujifilm (2 mentions) Immobilon p pvdf membrane, by Merck Group (2 mentions) Streptavidin agarose bead, by Thermo Fisher Scientific (2 mentions) Lds sample buffer, by Thermo Fisher Scientific (2 mentions) Biotin labeled goat anti human igm, by Jackson ImmunoResearch (2 mentions) Rabbit anti tlr7, by Cell Signaling Technology (2 mentions) Goat anti igm hrp, by Fortis Life Sciences (2 mentions) Anti ikbα, by Cell Signaling Technology (1 mentions) Rabbit anti mif, by Santa Cruz Biotechnology (1 mentions) Goat anti cd74, by Santa Cruz Biotechnology (1 mentions) Mouse anti nf κb p65, by Cell Signaling Technology (1 mentions) Rabbit anti gfap, by Santa Cruz Biotechnology (1 mentions) Enhanced chemiluminescence detection reagent, by Cytiva (1 mentions) Gs 700 imaging densitometer, by Bio-Rad (1 mentions) Specific secondary antibodies conjugated to horseradish peroxidase, by Agilent Technologies (1 mentions) Mouse anti inos, by Santa Cruz Biotechnology (1 mentions) Alexa 647, by Thermo Fisher Scientific (1 mentions) Mouse anti tlr4, by Santa Cruz Biotechnology (1 mentions)

Spelling variants of this product

Anti-TLR4 (81 citations) Rabbit anti-human TLR4 antibody (3 citations) Rabbit anti-TM (2 citations) Rabbit anti-rat TLR4 (2 citations) Rabbit anti-TLR4 polyclonal antibody (2 citations)

8 protocols using rabbit anti tlr4

1

Western Blotting Protein Analysis

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Western blotting was performed as described previously.19, 20, 21 We used the proteins from the renal cortex for Western blotting analysis. The catalogue number of antibodies used for Western blot were as following: rabbit anti‐MIF (Santa Cruz sc‐20121), goat anti‐CD74 (Santa Cruz, sc‐5438), rabbit anti‐phosphorylated NF‐κB p65 (Cell Signaling, No. 3031), mouse anti‐NF‐κB p65 (Cell Signaling, No. 6965), rabbit anti‐phosphorylated IkBα (Cell Signaling, 2859), anti‐IKBα (Cell Signaling, 9242L), rabbit anti‐TLR4 (Santa Cruz, sc‐10741) and mouse anti‐β‐actin (Santa Cruz, sc‐69879).
Li J.H., Tang Y., Lv J., Wang X.H., Yang H., Tang P.M., Huang X.R., He Z.J., Zhou Z.J., Huang Q.Y., Klug J., Meinhardt A., Fingerle‐Rowson G., Xu A.P., Zheng Z.H, & Lan H.Y. (2019). Macrophage migration inhibitory factor promotes renal injury induced by ischemic reperfusion. Journal of Cellular and Molecular Medicine, 23(6), 3867-3877.
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2

Immunoblotting of Enteric Glial Markers

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Proteins were extracted from submucosal plexi deriving from rats and PPARα−/− mice at day 7 after diarrhea induction. The tissue was homogenized in ice-cold hypotonic lysis buffer to obtain cytosolic extracts and underwent electrophoresis through a polyacrilamide minigel. Proteins were transferred into nitrocellulose membrane that were saturated with non-fat dry milk and then incubated with either mouse anti-S100B (Neo-Marker, Milan, Italy), mouse anti-iNOS, rabbit anti-GFAP, rabbit anti-TLR4, and mouse anti-β-actin (all Santa Cruz Biotechnology, Santa Cruz, California, USA). Membranes were then incubated with the specific secondary antibodies conjugated to horseradish peroxidase (Dako, Milan, Italy). Immune complexes were revealed by enhanced chemiluminescence detection reagents (Amersham Biosciences, Milan, Italy). Blots were analyzed by scanning densitometry (GS-700 imaging densitometer; Bio-Rad). Results were expressed as OD (arbitrary units; mm2) and normalized on the expression of the housekeeping protein β-actin.
Sarnelli G., Seguella L., Pesce M., Lu J., Gigli S., Bruzzese E., Lattanzi R., D’Alessandro A., Cuomo R., Steardo L, & Esposito G. (2018). HIV-1 Tat-induced diarrhea is improved by the PPARalpha agonist, palmitoylethanolamide, by suppressing the activation of enteric glia. Journal of Neuroinflammation, 15, 94.
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3

Immunofluorescence Staining of Mouse Brain

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Immunofluorescence staining was performed on frozen coronal sections of mouse brains or on primary microglia plated on cover slips. The mouse brains were fixed with 4% paraformaldehyde. After post-fixation and concentration gradient dehydration, the brains were cut into 10-μm-thick sections using a Leica CM1900 frozen slicer. The cells were fixed with 4% paraformaldehyde for 30 min. The brain sections and cell cover slips were washed three times with PBS and then incubated overnight at 4 °C in a humidified atmosphere with primary antibodies. The following primary antibodies were used: mouse anti-TLR4 (1:100; Santa Cruz, USA), rabbit anti-TLR4 (1:100; Santa Cruz, USA), rabbit anti-GPR30 (1:200; Abcam, England), goat anti-Iba1 (1:200; Novus, USA), rabbit anti-Iba1 (1:500; Wako, Japan), and mouse anti-NeuN (1:200; Millipore, USA). Then, the samples were incubated with mixtures of Alexa-488 (red, Invitrogen) or Alexa-594 (red, Santa Cruz) and Alexa-647 (green, Invitrogen)-conjugated donkey anti-goat or anti-rabbit and donkey anti-mouse secondary antibodies for 2 h in the dark at room temperature. Finally, the sections were photographed using an Olympus BX51 (Japan) fluorescence microscope. The average area of single microglial cells was measured by ImageJ software.
Zhang Z., Qin P., Deng Y., Ma Z., Guo H., Guo H., Hou Y., Wang S., Zou W., Sun Y., Ma Y, & Hou W. (2018). The novel estrogenic receptor GPR30 alleviates ischemic injury by inhibiting TLR4-mediated microglial inflammation. Journal of Neuroinflammation, 15, 206.
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4

Immunoprecipitation and Western Blot Analysis of TLR Signaling

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HBL1, TMD8, OCI-Ly10 and OCI-Ly19 cells were lysed at 107 cells per ml in a modified RIPA buffer (0.5% Triton X-100, 0.25% deoxycholate, 0.025% SDS, 10 mM Tris, pH 8.0, 100 mM NaCl, 10 mM EDTA, 1 mM Na3VO4, 30 mM pyrophosphate, 10 mM glycerophosphate, 1 mM AEBSF, 0.02 U ml−1 aprotinin and 0.01% NaN3) for 10 min. on ice. Lysates were cleared by centrifugation at 14,000xg for 20 min. at 4°C. IgM was immunoprecipitated by incubating lysates on ice for 1 hour with 10 μg of biotin-labeled goat anti-human IgM (Jackson Immunoresearch), followed by the addition of 35 μl of pre-washed streptavidin-agarose beads (Invitrogen) and rotated for 30 min. at 4°C. Beads were washed 3X with cold 1X RIPA buffer, then solubilized by adding 2X LDS sample buffer (Invitrogen) with 1% β-mercaptoethanol and boiled for 5 min. Samples were separated on a 10% polyacrylamide gel and transferred to Immobilon-p PVDF membrane (Millipore) for western blot analysis. Membranes were probed with rabbit anti-TLR9 monoclonal XP, rabbit anti-TLR7 (Cell Signaling Technologies), rabbit anti-TLR4 (Santa Cruz Biotechnology) and goat anti-IgM-HRP (Bethyl).
Phelan J.D., Young R.M., Webster D.E., Roulland S., Wright G.W., Kasbekar M., Shaffer AL I.I.I., Ceribelli M., Wang J.Q., Schmitz R., Nakagawa M., Bachy E., Huang D.W., Ji Y., Chen L., Yang Y., Zhao H., Yu X., Xu W., Palisoc M.M., Valadez R.R., Davies-Hill T., Wilson W.H., Chan W.C., Jaffe E.S., Gascoyne R.D., Campo E., Rosenwald A., Ott G., Delabie J., Rimsza L.M., Rodriguez F.J., Estephan F., Holdhoff M., Kruhlak M.J., Hewitt S.M., Thomas C.J., Pittaluga S., Oellerich T, & Staudt L.M. (2018). A Multiprotein Supercomplex Controlling Oncogenic Signaling in Lymphoma. Nature, 560(7718), 387-391.
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5

Western Blotting Analysis of GPR30, Iba1, and TLR4

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Expression of the GPR30, Iba1, and TLR4 proteins in penumbra and primary microglia was measured by Western blotting as previously described [2 (link)]. The extracted proteins were separated by 10% SDS-PAGE and electrically transferred to polyvinylidene difluoride membranes. Then, the membranes were blocked with TBST containing 5% nonfat dry milk for 1 h at room temperature. The following primary antibodies were used: rabbit anti-GPR30 (1:1000; Abcam, England), rabbit anti-Iba1 (1:500; Wako, Japan), rabbit anti-TLR4 (1:200; Santa Cruz, USA), rabbit anti-NF-κB (Cell Signaling Technology; 1:500), mouse anti-β-actin (1:1000; Cell Signaling Technology, USA), rabbit anti-GAPDH (1:1000; Cell Signaling Technology, USA), and rabbit anti-Histone H2A.X (Cell Signaling Technology; 1:1000). The membranes were shaken at 60 rpm/min at 4 °C overnight and incubated with an IRDye secondary anti-rabbit or mouse antibody (Thermo Scientific, USA) for 1 h. Protein bands were visualized using the LI-COR Odyssey System (LI-COR Biotechnology, USA).
Zhang Z., Qin P., Deng Y., Ma Z., Guo H., Guo H., Hou Y., Wang S., Zou W., Sun Y., Ma Y, & Hou W. (2018). The novel estrogenic receptor GPR30 alleviates ischemic injury by inhibiting TLR4-mediated microglial inflammation. Journal of Neuroinflammation, 15, 206.
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6

Quantitative Western Blot Analysis of TLR4, HMGB1, and ATX

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Western blot analysis was performed as previously described [53 (link)] with minor alterations. Briefly, 100 μg of protein from tissue extracts were separated on a 12% SDS-PAGE gel. Following electrophoretic transfer onto a nitrocellulose membrane and blocking with 5% milk solution, the blots were incubated with primary antibody overnight at 4°C [rabbit anti-TLR4 (Santa Cruz, Dallas, TX, USA, #sc-10741; 1:500), mouse anti-HMGB1 (BioLegend, San Diego, CA, USA, #651402; 1:500), rabbit anti-ATX (Millipore, Billerica, MA, USA, #ABT28; 1:500), or mouse anti-β-actin (Sigma, USA, #A5441; 1:5,000)] and with horseradish peroxidase-labeled secondary antibody [anti-mouse or anti-rabbit (Santa Cruz, USA, #sc-2005 and #sc-2004, respectively; 1:5,000)] for 1 h at RT. Protein bands were detected by LumiGLO® (Cell Signaling, Danvers, MA, USA) and visualized by chemiluminescence with ChemiDoc (Bio-Rad, Hercules, CA, USA). Expression was quantified by computerized image analysis using Quantity One 1-D Analysis Software (Bio-Rad, Hercules, CA, USA).
Cardoso F.L., Herz J., Fernandes A., Rocha J., Sepodes B., Brito M.A., McGavern D.B, & Brites D. (2015). Systemic inflammation in early neonatal mice induces transient and lasting neurodegenerative effects. Journal of Neuroinflammation, 12, 82.
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7

Immunohistochemical Analysis of Immune Markers

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Immunohistochemistry was performed as previously described 5 (link). The following primary antibodies were used for immunohistochemical analysis: mouse anti-Ki-67 (Abcam, Cambridge, USA), rabbit anti-CD68 (Abbiotec, San Diego, CA, USA), mouse anti-CD68 (Dako, Glostrup, Denmark), anti-CD66b rabbit (Cell Signaling Technology, Danvers, USA), rabbit anti-hCAP18/LL-37 (Santa Cruz, CA, USA), rabbit anti-TLR2 (Santa Cruz, CA, USA), rabbit anti-TLR4 (Santa Cruz, CA, USA), and mouse anti-unphosphorylated β-catenin (Millipore, Billerica, USA). Secondary antibody incubation and staining were performed using the EnVision®+ System-HRP (DAB) kit (Dako, Glostrup, Denmark), according to instructions provided by the manufacturer. TUNEL staining was performed using the DeadEnd Colorimetric TUNEL System kit (Promega, Madison, USA).
Ji P., Zhou Y., Yang Y., Wu J., Zhou H., Quan W., Sun J., Yao Y., Shang A., Gu C., Zeng B., Firrman J., Xiao W., Bals R., Sun Z, & Li D. (2019). Myeloid cell-derived LL-37 promotes lung cancer growth by activating Wnt/β-catenin signaling. Theranostics, 9(8), 2209-2223.
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8

Immunoprecipitation and Western Blot Analysis of TLR Signaling

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HBL1, TMD8, OCI-Ly10 and OCI-Ly19 cells were lysed at 107 cells per ml in a modified RIPA buffer (0.5% Triton X-100, 0.25% deoxycholate, 0.025% SDS, 10 mM Tris, pH 8.0, 100 mM NaCl, 10 mM EDTA, 1 mM Na3VO4, 30 mM pyrophosphate, 10 mM glycerophosphate, 1 mM AEBSF, 0.02 U ml−1 aprotinin and 0.01% NaN3) for 10 min. on ice. Lysates were cleared by centrifugation at 14,000xg for 20 min. at 4°C. IgM was immunoprecipitated by incubating lysates on ice for 1 hour with 10 μg of biotin-labeled goat anti-human IgM (Jackson Immunoresearch), followed by the addition of 35 μl of pre-washed streptavidin-agarose beads (Invitrogen) and rotated for 30 min. at 4°C. Beads were washed 3X with cold 1X RIPA buffer, then solubilized by adding 2X LDS sample buffer (Invitrogen) with 1% β-mercaptoethanol and boiled for 5 min. Samples were separated on a 10% polyacrylamide gel and transferred to Immobilon-p PVDF membrane (Millipore) for western blot analysis. Membranes were probed with rabbit anti-TLR9 monoclonal XP, rabbit anti-TLR7 (Cell Signaling Technologies), rabbit anti-TLR4 (Santa Cruz Biotechnology) and goat anti-IgM-HRP (Bethyl).
Phelan J.D., Young R.M., Webster D.E., Roulland S., Wright G.W., Kasbekar M., Shaffer AL I.I.I., Ceribelli M., Wang J.Q., Schmitz R., Nakagawa M., Bachy E., Huang D.W., Ji Y., Chen L., Yang Y., Zhao H., Yu X., Xu W., Palisoc M.M., Valadez R.R., Davies-Hill T., Wilson W.H., Chan W.C., Jaffe E.S., Gascoyne R.D., Campo E., Rosenwald A., Ott G., Delabie J., Rimsza L.M., Rodriguez F.J., Estephan F., Holdhoff M., Kruhlak M.J., Hewitt S.M., Thomas C.J., Pittaluga S., Oellerich T, & Staudt L.M. (2018). A Multiprotein Supercomplex Controlling Oncogenic Signaling in Lymphoma. Nature, 560(7718), 387-391.
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