Anti-p50 (sc-1190X), anti-c-Rel (sc-71X), anti-p65 (sc-372X), anti-IκBα (sc-371), anti-NOTCH1 (sc-6014-R), anti-GFP (sc-8334), anti-BCL10 (sc-9558 and sc5273), anti-CARMA1 (sc-20458), anti-IKKα (sc-7606), anti-IKKγ (sc-8330), normal goat IgG (sc-2043), and normal rabbit IgG (sc-2027) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA); cleaved N1 (#2421) and phospho-IκBα (#9241) antibodies were from Cell Signaling Technology (Beverly, MA, USA). GAPDH antibody (MAB374) was from Chemicon International, Inc. (Temecula, CA, USA). Anti-human CD3ε (145-2C11) and anti-human CD28 were obtained from R&D Systems (Minneapolis, MN, USA). Anti-CARMA1 (#3189) was obtained from ProSci, Inc. (Poway, CA, USA). Cholera toxin (CTX) B subunit, FITC-labeled (C 1655), monoclonal anti-VSV glycoprotein (V 5507), and anti-β-actin were purchased from Sigma-Aldrich (St. Louis, MO, USA).
Normal rabbit igg sc 2027
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Normal rabbit IgG (sc-2027) is a laboratory reagent used as an isotype control in immunoassays. It is purified from the serum of normal, non-immunized rabbits.
Automatically generated - may contain errors
Lab products found in correlation
Normal mouse igg sc 2025, by Santa Cruz Biotechnology (7 mentions)
Protein a g plus agarose, by Santa Cruz Biotechnology (3 mentions)
Alexa fluor 488 donkey anti rabbit igg antibody a21206, by Thermo Fisher Scientific (3 mentions)
Immobilized glutathione, by Thermo Fisher Scientific (2 mentions)
Anti erk2 c 14 sc 154, by Santa Cruz Biotechnology (2 mentions)
Rat anti dykddddk flag cat 200474, by Agilent Technologies (2 mentions)
Anti mlk3 d 11 sc 166639, by Santa Cruz Biotechnology (2 mentions)
Protein g resin, by GenScript (2 mentions)
Goat anti mouse igg hrp sc 2005, by Santa Cruz Biotechnology (2 mentions)
Protease inhibitor cocktail, by Roche (2 mentions)
Mouse 7076 and rabbit 7074 hrp conjugated secondary antibodies, by Cell Signaling Technology (2 mentions)
Alexa fluor 488, by Thermo Fisher Scientific (2 mentions)
Bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Anti human cd28, by R&D Systems (1 mentions)
Anti β actin, by Merck Group (1 mentions)
Ab4729, by R&D Systems (1 mentions)
Af2726, by R&D Systems (1 mentions)
Goat igg sc2028, by Santa Cruz Biotechnology (1 mentions)
α tubulin t5168, by Merck Group (1 mentions)
Ab18197, by Abcam (1 mentions)
36 protocols using normal rabbit igg sc 2027
1
Antibody Inventory for NF-κB Signaling
2
ChIP-seq protocol for epigenomic profiling
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ChIP was carried out as described (25 (link)). Briefly, K562 cells (2 × 107) were cross-linked with 1% formaldehyde and then nuclei were isolated by cell lysis. MNase digestion and sonication were performed with nuclei to produce chromatin at a mononucleosome size. Fragmented chromatin was incubated with antibodies after pre-clearing. Protein–DNA complexes were recovered with protein A or G agarose beads, and DNA was purified after reverse cross-linking.
Antibodies used in this study were purchased from Santa Cruz Biotechnology for TAL1 (sc-12984), GATA-1 (sc-1233), p45/NF-E2 (sc-22827), Brg1 (sc-10768), CBP (sc-369), Pol II (sc-899) and Ldb1 (sc-11198), from Millipore for H3ac (06-599), H3K4me2 (07-030 (link)) and H3K27me2 (07-452), from Abcam for H3K27ac (ab4729), H3K36me3 (ab9050), Rad21 (ab992) and SMC3 (ab9263) and from R&D systems for LMO2 (AF2726). Normal rabbit IgG (sc-2027) and goat IgG (sc-2028) were purchased from Santa Cruz Biotechnology.
Antibodies used in this study were purchased from Santa Cruz Biotechnology for TAL1 (sc-12984), GATA-1 (sc-1233), p45/NF-E2 (sc-22827), Brg1 (sc-10768), CBP (sc-369), Pol II (sc-899) and Ldb1 (sc-11198), from Millipore for H3ac (06-599), H3K4me2 (07-030 (link)) and H3K27me2 (07-452), from Abcam for H3K27ac (ab4729), H3K36me3 (ab9050), Rad21 (ab992) and SMC3 (ab9263) and from R&D systems for LMO2 (AF2726). Normal rabbit IgG (sc-2027) and goat IgG (sc-2028) were purchased from Santa Cruz Biotechnology.
3
Antibody Characterization and Validation
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GFP (#sc-8834), p53 (DO-1, #sc-126) p53 FL-393 (s#c-6243), Myc (9E10, #sc-40) and normal rabbit IgG (sc-2027) antibodies were purchased from Santa Cruz Biotechnologies. Flag (M2, #F3165) and α-tubulin (#T5168) antibodies were obtained from Sigma. ACTIN (C4, #69100) antibodies were purchased from MP Biomedicals and HA (3F10, #11867423001) antibodies from Roche. Ubiquitin (P4D1, #3936), and p53 phospho-Ser46 (#2521) antibodies were from Cell Signaling Technology. Cleaved PARP (Y34, #ab32561) was obtained from Abcam. The H4 (#07-108) antibodies were from Millipore. The affinity-purified HIPK2 antibody and the chicken SIAH1 antibody have been described previously (22 (link)). Rabbit polyclonal DAZAP2 antibodies were raised against the following keyhole limpet hemocyanine (KLH)-coupled peptide: H2N-MNSKGQYPTQPTYPVC-COOH. The rabbit sera were affinity-purified against the peptide prior to use. The phosphorylation specific DAZAP2 Ser77 antibody was raised against the following phospho-peptide: H2N-YLPMA-S(p)-VAVGPLC-COOH and the phospho-specific antibodies were subsequently affinity purified.
4
Immunohistochemical Analysis of Cell Proliferation in Atheromatous Aortic Valve Lesions
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Cell proliferation in atheromatous lesions from aortic valves was assessed using anti‐proliferating cell nuclear antigen (PCNA) antibody (rabbit polyclonal) (ab18197, Abcam). For double immunohistochemical staining, the sections were autoclaved in citrate buffer to restore antigenicity. A monoclonal anti‐macrophage MAC3 antibody (550292; BD Pharmingen, San Diego, CA) and polyclonal anti‐PCNA form antibodies (ab18197, anti‐PCNA antibody [rabbit polyclonal]; Abcam) were used as primary antibodies. Normal rabbit IgG (sc‐2027, Santa Cruz) was used for MAC3 antibody negative control. Goat anti‐rabbit IgG (H+L) (highly cross‐adsorbed; Alexa Fluor 488, A11034; Thermo Fisher Scientific Inc.) was used as a secondary antibody for anti‐PCNA. Normal rabbit IgG (sc‐2027, Santa Cruz) was used as the PCNA antibody‐negative control. Goat anti‐rat IgG (H+L) (Alexa Fluor 555, A21434; Thermo Fisher Scientific Inc.) was used as a secondary antibody for anti‐PCNA.
5
Synthesis and Characterization of FPD5
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The small chemical molecule FPD5 was synthesized in the laboratory of Professor Baoxiang Zhao (Shandong University, Jinan). The synthetic protocols were described in detail previously (ref. [23 ]).
Antibodies for ESD (sc-134333), GFP (sc-8334), ACTB (sc-47778), GAPDH (sc-47724), goat anti-mouse horseradish peroxidase (HRP)-conjugated IgG (GP016129) and goat anti-rabbit HRP-IgG (sc-2004) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Mouse anti RFP-Tag mAb (AE020) was from ABclonal. Antibodies for SQSTM1 (610833) were from BD Biosciences. Lipofectamine 2000 transfection reagent (11668-027) and acridine orange (AO, A3568) were from Invitrogen. Normal mouse IgG (sc-2025) and normal rabbit IgG (sc-2027), controls for immunofluorescence and immunoprecipitation assays, were from Santa Cruz Biotechnology. Secondary antibodies for immunofluorescence were goat anti-mouse IgG Alexa Fluor-488 (Invitrogen, A1101) and goat anti-rabbit IgG (Invitrogen, A21070). The antibodies ESD (ab133631) and FKBP25 (ab16654) were purchased from Abcam and mTORC1 (20657-1-AP) was from Proteintech. The antibodies p-mTORC1 (2971s), 4EBP1 (9452s), p-4EBP1 (9459S), P70S6K (9205s), and p-P70S6K (9202s) were from Cell Signaling Technology. SiRNA against ESD or FKBP25 was designed and synthesized by GenePharma (Shanghai). Scramble siRNA was used as a control (Santa Cruz Biotechnology, sc-37007).
Antibodies for ESD (sc-134333), GFP (sc-8334), ACTB (sc-47778), GAPDH (sc-47724), goat anti-mouse horseradish peroxidase (HRP)-conjugated IgG (GP016129) and goat anti-rabbit HRP-IgG (sc-2004) were purchased from Santa Cruz Biotechnology (Santa Cruz, CA). Mouse anti RFP-Tag mAb (AE020) was from ABclonal. Antibodies for SQSTM1 (610833) were from BD Biosciences. Lipofectamine 2000 transfection reagent (11668-027) and acridine orange (AO, A3568) were from Invitrogen. Normal mouse IgG (sc-2025) and normal rabbit IgG (sc-2027), controls for immunofluorescence and immunoprecipitation assays, were from Santa Cruz Biotechnology. Secondary antibodies for immunofluorescence were goat anti-mouse IgG Alexa Fluor-488 (Invitrogen, A1101) and goat anti-rabbit IgG (Invitrogen, A21070). The antibodies ESD (ab133631) and FKBP25 (ab16654) were purchased from Abcam and mTORC1 (20657-1-AP) was from Proteintech. The antibodies p-mTORC1 (2971s), 4EBP1 (9452s), p-4EBP1 (9459S), P70S6K (9205s), and p-P70S6K (9202s) were from Cell Signaling Technology. SiRNA against ESD or FKBP25 was designed and synthesized by GenePharma (Shanghai). Scramble siRNA was used as a control (Santa Cruz Biotechnology, sc-37007).
6
TEM1 Immunoprecipitation and Western Blot
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HNDF cultured with 10% FBS DMEM were lysed by the lysis buffer (0.1% Triton X-100, 0.1 M Tris-buffer, pH 8.1) with phosphatase inhibitors (1 mM Na3VO4 and 25 mM NaF) and protease inhibitors (Sigma-Aldrich, St. Louis, Missouri, USA). Cell lysates (1000 μg) were incubated with 1 μg anti-TEM1 antibody (18,160–1-AP; Proteintech, Cook County, Illinois, USA) or normal rabbit IgG (sc-2027; Santa Cruz Biotechnology, Dallas, Texas, USA) and 20 μL of protein A and G agarose beads (Millipore, Burlington, Massachusetts, USA) at 4 °C overnight. Immunoprecipitates were washed with lysis buffer 3 times and analyzed by Western blotting.
7
Antibody Characterization for Pax7 and Chromatin Remodeling
8
Characterization of CCL5/CCR5 Signaling
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Recombinant human CCL5 (278‐RN‐050CF) was purchased from R&D Systems (Minneapolis, Minnesota, USA). The primary antibodies used in western blot and immunohistochemistry (IHC) assays included rabbit anti‐CCR5 (ab32048; Abcam, Cambridge, Massachusetts, USA), anti‐AR (sc‐7305; Santa Cruz Biotechnology, Dallas, Texas, USA) and anti‐GAPDH (NB300‐221; Novus Biologicals, Littleton, Colorado, USA). MAB‐678 antihuman CCL5 monoclonal neutralizing antibody was obtained from R&D Systems. Normal rabbit IgG (sc‐2027) used as a control in the CCL5 neutralizing antibody assay was obtained from Santa Cruz Biotechnology.
9
ChIP-qPCR analysis of cell cycle
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ChIP was performed as described 37 (link). Briefly, about 3×106 MEFs were harvested at about passage 6 and fixed with 1% formaldehyde. After cell lysis, nuclei were separated and sonicated on ice to fragment DNA to an average length of 700 bp. Then the cross-liked and sonicated chromatin was subjected to immunoprecipitation with ChIP grade antibodies: cyclin A (H-432X), E2F1 (C-20X), E2F4 (A-20X) and normal rabbit IgG (SC-2027) (all from Santa Cruz Biotechnology). Following purification, about 5% of total precipitated DNA and 0.1% input were analyzed by qPCR. Data were analyzed as % of input. ChIP primer sequences were listed in Supplementary Table 1 .
10
Immunoprecipitation and GST Pull-down Assay
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