Rabbit TrueBlot®: Anti-Rabbit IgG HRP
Cat# 18-8816-31
Size : 50ul
Brand : Rockland Immunochemicals
Contact local distributor :
Phone : +1 850 650 7790
Specifications for Rabbit TrueBlot®: Anti-Rabbit IgG HRP
Product Details
Rabbit TrueBlot®: Anti-Rabbit IgG HRP - 18-8816-31
Anti-Rabbit IgG HRP, TrueBlot, HRP TrueBlot ULTRA, Peroxidase TrueBlot, TrueBlot for IP/WB, TrueBlot for immunoprecipitation, TrueBlot for western blotting
Mouse
Peroxidase (HRP) ULTRA
Monoclonal
IgG
Immunoprecipitation Kit
Target Details
Rabbit
Rabbit TrueBlot® Antibody Peroxidase Conjugate was prepared from tissue culture supernatant by Protein G affinity chromatography. Assay by immunoelectrophoresis resulted in a single precipitin arc against Anti-Rabbit Serum. Reactivity is observed against native Rabbit IgG by both Western blot and ELISA.
Application Details
ELISA, IP, WB
ChIP, FISH, IF - View References
Rabbit IgG HRP TrueBlot has been tested in ELISA, Western blot, and immunoprecipitation and may also be used for detection in immunoblotting assays that do not employ immunoprecipitation. Rabbit IgG TrueBlot® is provided as 1000X solution. To conserve reagent, we recommend incubating the blots from minigels in sealed bags (removing as much air as possible) with minimal volume (2-3 mLs). If used conservatively at 2.5 mLs/blot will yield enough reagent for 20 blots. Note that there are three key procedural considerations: 1. Protein A or G should not be used for the immunoprecipitation. Use of protein A or G beads with the rabbit TrueBlot will result in contaminating bands. For immunoprecipitation, Anti-Rat IgG beads, or Anti-Rabbit IgG beads should be used for rat or rabbit immunoprecipitating antibodies, respectively. 2. Immunoprecipitate should be completely reduced. 3. BLOTTO/Milk should be used as the blocking protein for the immunoblot. All recommended dilutions for listed applications are intended as an initial recommendation, specific conditions for each protein and antibody combination should be specifically optimized by the end user.
Formulation
1.0 mg/mL by UV absorbance at 280 nm
0.02 M Potassium Phosphate, 0.15 M Sodium Chloride, pH 7.2
Proclin is added as an antimicrobial agent.
0.1 mg/ml Bovine Serum Albumin (BSA) - IgG and Protease free, 50% (v/v) Glycerol
Shipping & Handling
Wet Ice
Store at -20°C. This product is guaranteed for 6 months upon receipt, when handled and stored as instructed.
Expiration date is six (6) months from date of receipt.
Background
Rabbit IgG TrueBlot® is a unique horseradish peroxidase conjugated Anti-Rabbit IgG monoclonal secondary antibody. Rabbit IgG TrueBlot® enables detection of immunoblotted target protein bands, without hindrance by interfering immunoprecipitating immunoglobulin heavy and light chains. It is easy to generate publication-quality IP/Western Blot data with Rabbit IgG TrueBlot®, simply substitute the conventional HRP Anti-Rabbit IgG blotting reagent with Rabbit IgG TrueBlot® and follow the prescribed protocol for sample preparation and immunoblotting. Rabbit IgG TrueBlot® is ideal for use in protocols involving immunoblotting of immunoprecipitated proteins. TrueBlot preferentially detects the non-reduced form of rabbit IgG over the reduced, SDS-denatured form of IgG. When the immunoprecipitate is fully reduced immediately prior to SDS-gel electrophoresis, reactivity of Rabbit IgG TrueBlot® with the 55 kDa heavy chains and the 23 kDa light chains of the immunoprecipitating antibody is minimized thereby eliminating interference by the heavy and light chains of the immunoprecipitating antibody in IP/Western blot applications. Applications include studies examining post-translational modification (e.g., phosphorylation or acetylation) or protein-protein interactions.
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Predescu et al. (2018). Epsin15 Homology Domains: Role in the Pathogenesis of Pulmonary Arterial Hypertension. Frontiers in Physiology
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Pazos et al. (2018). Z-ring membrane anchors associate with cell wall synthases to initiate bacterial cell division. Nature Communications
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Gong L, Pan X, Lim CB, de Polo A, Little JB, Yuan ZM. (2018). A functional interplay between Δ133p53 and ΔNp63 in promoting glycolytic metabolism to fuel cancer cell proliferation. Oncogene.
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Hou W, Cruz-Cosme R, Wen F, et al. (2018). Expression of Human Cytomegalovirus IE1 Leads to Accumulation of Mono-SUMOylated PML That Is Protected from Degradation by Herpes Simplex Virus 1 ICP0. J Virol.
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Lahaye X, Gentili M, Silvin A, et al. (2018). NONO Detects the Nuclear HIV Capsid to Promote cGAS-Mediated Innate Immune Activation. Cell. 2018;175(2):488-501.e22. doi:10.1016/j.cell.2018.08.062 Cell.
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Zhang et al. (2017). Protein Phosphotyrosine Phosphatase 1B (PTP1B) in Calpain-dependent Feedback Regulation of Vascular Endothelial Growth Factor Receptor (VEGFR2) in Endothelial Cells: IMPLICATIONS IN VEGF-DEPENDENT ANGIOGENESIS AND DIABETIC WOUND HEALING. Journal of Biological Chemistry
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Prevost et al. (2017). The Legionella effector WipB is a translocated Ser/Thr phosphatase that targets the host lysosomal nutrient sensing machinery. Scientific Reports
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Iwahori S, Umaña AC, VanDeusen HR, Kalejta RF. (2017). Human cytomegalovirus-encoded viral cyclin-dependent kinase (v-CDK) UL97 phosphorylates and inactivates the retinoblastoma protein-related p107 and p130 proteins. J Biol Chem.
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Schering L, Albrecht E, Komolka K, Kühn C, Maak S. (2017). Increased expression of thyroid hormone responsive protein (THRSP) is the result but not the cause of higher intramuscular fat content in cattle. Int J Biol Sci.
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Iwahori S, Kalejta RF. (2017). Phosphorylation of transcriptional regulators in the retinoblastoma protein pathway by UL97, the viral cyclin-dependent kinase encoded by human cytomegalovirus. Virology.
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Perez KJ, Martínez FP, Cosme-Cruz R, Perez-Crespo NM, Tang Q. Correction for Perez et al., (2017). A Short cis-Acting Motif in the M112-113 Promoter Region Is Essential for IE3 To Activate M112-113 Gene Expression and Is Important for Murine Cytomegalovirus Replication. J Virol.
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Aly et al. (2016). RNA Exosome Complex Regulates Stability of the Hepatitis B Virus X-mRNA Transcript in a Non-stop-mediated (NSD) RNA Quality Control Mechanism. Journal of Biological Chemistry
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Hwang et al. (2016). Leucine Carboxyl Methyltransferase 1 (LCMT-1) Methylates Protein Phosphatase 4 (PP4) and Protein Phosphatase 6 (PP6) and Differentially Regulates the Stable Formation of Different PP4 Holoenzymes. Journal of Biological Chemistry
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Faouzi M, Hague F, Geerts D, et al. (2016). Functional cooperation between KCa3.1 and TRPC1 channels in human breast cancer: Role in cell proliferation and patient prognosis. Oncotarget.
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Masser DR, Clark NW, Van Remmen H, Freeman WM. (2016). Loss of the antioxidant enzyme CuZnSOD (Sod1) mimics an age-related increase in absolute mitochondrial DNA copy number in the skeletal muscle. Age (Dordr).
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Chung et al. (2015). Regulation of C-X-C chemokine gene expression by keratin 17 and hnRNP K in skin tumor keratinocytes. Journal of Cell Biology
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Kumar et al. (2015). MEI4 – a central player in the regulation of meiotic DNA double-strand break formation in the mouse. Journal of Cell Science
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Wani et al. (2015). Maturation of the proteasome core particle induces an affinity switch that controls regulatory particle association. Nature Communications
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Marsh NM, Wareham A, White BG, Miskiewicz EI, Landry J, MacPhee DJ. (2015). HSPB8 and the Cochaperone BAG3 Are Highly Expressed During the Synthetic Phase of Rat Myometrium Programming During Pregnancy. Biol Reprod.
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Albrecht E, Kuzinski J, Komolka K, Gotoh T, Maak S. (2015). Localization and abundance of early markers of fat cell differentiation in the skeletal muscle of cattle during growth--are DLK1-positive cells the origin of marbling flecks? Meat Sci.
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Iwahori S, Hakki M, Chou S, Kalejta RF. (2015). Molecular Determinants for the Inactivation of the Retinoblastoma Tumor Suppressor by the Viral Cyclin-dependent Kinase UL97. J Biol Chem.
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Applications
WB, IB, PCA PubMed
Yousif AS et al. (2014). Differential regulation of S-region hypermutation and class switch recombination by noncanonical functions of uracil DNA glycosylase. Proc Natl Acad Sci USA
Applications
ChIP; IP, Co-IP PubMed
Dusonchet J et al. (2014). A Parkinson's disease gene regulatory network identifies the signaling protein RGS2 as a modulator of LRRK2 activity and neuronal toxicity. Hum Mol Genet.
Applications
WB, IB, PCA PubMed
Ogino H, Ishino S, Haugland GT, Birkeland NK, Kohda D, Ishino Y. (2014). Activation of the MCM helicase from the thermophilic archaeon, Thermoplasma acidophilum by interactions with GINS and Cdc6-2. Extremophiles.
Applications
WB, IB, PCA PubMed
Amigo-Jiménez I, Bailón E, Ugarte-Berzal E, Aguilera-Montilla N, García-Marco JA, García-Pardo A. (2014). Matrix metalloproteinase-9 is involved in chronic lymphocytic leukemia cell response to fludarabine and arsenic trioxide. PLoS One.
Applications
WB, IB, PCA PubMed
Puglisi R, Maccari I, Pipolo S, Mangia F, Boitani C. (2014). The nuclear form of glutathione peroxidase 4 colocalizes and directly interacts with protamines in the nuclear matrix during mouse sperm chromatin assembly. Spermatogenesis.
Applications
WB, IB, PCA PubMed
Komolka K, Albrecht E, Schering L, Brenmoehl J, Hoeflich A, Maak S. (2014). Locus characterization and gene expression of bovine FNDC5: is the myokine irisin relevant in cattle? PLoS One.
Applications
WB, IB, PCA PubMed
Totsuka N, Kim YG, Kanemaru K, et al. (2014). Toll-like receptor 4 and MAIR-II/CLM-4/LMIR2 immunoreceptor regulate VLA-4-mediated inflammatory monocyte migration. Nat Commun.
Applications
WB, IB, PCA PubMed
Jerman S, Ward HH, Lee R, et al. (2014). OFD1 and flotillins are integral components of a ciliary signaling protein complex organized by polycystins in renal epithelia and odontoblasts. PLoS One.
Applications
WB, IB, PCA PubMed
Baetz NW, Goldenring JR. (2014). Distinct patterns of phosphatidylserine localization within the Rab11a-containing recycling system. Cell Logist.
Applications
WB, IB, PCA PubMed
Dannenberger D, Nuernberg K, Nuernberg G, Priepke A. (2014). Impact of dietary protein level and source of polyunsaturated fatty acids on lipid metabolism-related protein expression and fatty acid concentrations in porcine tissues. J Agric Food Chem.
Applications
WB, IB, PCA PubMed
Gong H et al. (2013). Lipoprotein lipase (LPL) is associated with neurite pathology and its levels are markedly reduced in the dentate gyrus of Alzheimer's disease brains. The Journal of Histochemistry and Cytochemistry : Official Journal of the Histochemistry Society
Applications
WB, IB, PCA PubMed
Baetz NW, Goldenring JR. (2013). Rab11-family interacting proteins define spatially and temporally distinct regions within the dynamic Rab11a-dependent recycling system. Mol Biol Cell.
Applications
WB, IB, PCA PubMed
Demir K, Kirsch N, Beretta CA, et al. (2013). RAB8B is required for activity and caveolar endocytosis of LRP6. Cell Rep.
Applications
WB, IB, PCA PubMed
Gao Y, Koppen A, Rakhshandehroo M, et al. (2013). Early adipogenesis is regulated through USP7-mediated deubiquitination of the histone acetyltransferase TIP60. Nat Commun.
Applications
IP, Co-IP PubMed
Nelersa CM et al. (2012). High-content analysis of proapoptotic EphA4 dependence receptor functions using small-molecule libraries. J Biolmol Screen
Applications
WB, IB, PCA PubMed
Martínez FP, Tang Q. (2012). Leucine zipper domain is required for Kaposi sarcoma-associated herpesvirus (KSHV) K-bZIP protein to interact with histone deacetylase and is important for KSHV replication. J Biol Chem.
Applications
WB, IB, PCA PubMed
Albrecht E, Komolka K, Kuzinski J, Maak S. (2012). Agouti revisited: transcript quantification of the ASIP gene in bovine tissues related to protein expression and localization. PLoS One.
Applications
WB, IB, PCA PubMed
Medici D, Shore EM, Lounev VY, Kaplan FS, Kalluri R, Olsen BR. (2011). Conversion of vascular endothelial cells into multipotent stem-like cells. Nat Med.
Applications
IP, Co-IP PubMed
Vuletic S, Dong W, Wolfbauer G, Tang C, Albers JJ. (2011). PLTP regulates STAT3 and NFκB in differentiated THP1 cells and human monocyte-derived macrophages. Biochim Biophys Acta.
Applications
WB, IB, PCA PubMed
Cosme-Cruz R, Martínez FP, Perez KJ, Tang Q. (2011). H2B homology region of major immediate-early protein 1 is essential for murine cytomegalovirus to disrupt nuclear domain 10, but is not important for viral replication in cell culture. J Gen Virol.
Applications
WB, IB, PCA PubMed
Rzeczkowski K, Beuerlein K, Müller H, et al. (2011). c-Jun N-terminal kinase phosphorylates DCP1a to control formation of P bodies. J Cell Biol.
Applications
WB, IB, PCA PubMed
Chanda D, Xie YB, Choi HS. (2010). Transcriptional corepressor SHP recruits SIRT1 histone deacetylase to inhibit LRH-1 transactivation. Nucleic Acids Res.
Applications
WB, IB, PCA PubMed
Xie YB, Nedumaran B, Choi HS. (2009). Molecular characterization of SMILE as a novel corepressor of nuclear receptors. Nucleic Acids Res.
Applications
WB, IB, PCA PubMed
Xie YB, Park JH, Kim DK, et al. (2009). Transcriptional corepressor SMILE recruits SIRT1 to inhibit nuclear receptor estrogen receptor-related receptor gamma transactivation. J Biol Chem.
Applications
WB, IB, PCA; IP, Co-IP PubMed
Mitsuhashi S et al. (2008). Tautomycetin suppresses the TNFα/NF-κB pathway via inhibition of IKK activation. Int J Oncol
Applications
IP, Co-IP PubMed
Wolter S, Doerrie A, Weber A, et al. (2008). c-Jun controls histone modifications, NF-kappaB recruitment, and RNA polymerase II function to activate the ccl2 gene. Mol Cell Biol. 2008;28(13):4407-4423. doi:10.1128/MCB.00535-07 Mol Cell Biol.
Applications
WB, IB, PCA PubMed
Cozzone D, Fröjdö S, Disse E, et al. (2008). Isoform-specific defects of insulin stimulation of Akt/protein kinase B (PKB) in skeletal muscle cells from type 2 diabetic patients. Diabetologia.
Applications
WB, IB, PCA PubMed
Li H, Xu LL, Masuda K, et al. (2008). A feedback loop between the androgen receptor and a NEDD4-binding protein, PMEPA1, in prostate cancer cells. J Biol Chem.
Applications
WB, IB, PCA PubMed
Sun J, Lin X. (2008). Beta-arrestin 2 is required for lysophosphatidic acid-induced NF-kappaB activation. Proc Natl Acad Sci U S A.
Applications
WB, IB, PCA PubMed
Nelson RF, Glenn KA, Zhang Y, et al. (2007). Selective cochlear degeneration in mice lacking the F-box protein, Fbx2, a glycoprotein-specific ubiquitin ligase subunit. J Neurosci.
Applications
WB, IB, PCA PubMed
Woo SY, Kim DH, Jun CB, et al. (2007). PRR5, a novel component of mTOR complex 2, regulates platelet-derived growth factor receptor beta expression and signaling. J Biol Chem.
Applications
IP, Co-IP PubMed
Zhang AS, West AP Jr, Wyman AE, Bjorkman PJ, Enns CA. (2005). Interaction of hemojuvelin with neogenin results in iron accumulation in human embryonic kidney 293 cells. J Biol Chem.
Applications
WB, IB, PCA PubMed
Frequently Asked Questions (13)
What is Rockland’s TrueBlot® product line?
Rockland’s TrueBlot® product line is designed to solve common experimental problems when performing immunoprecipitation/co-immunoprecipitation experiments and Western blots of immunoprecipitated samples (IP–Western blot). The product line consists of TrueBlot® monoclonal secondary antibodies and TrueBlot® IP beads.
What is the advantage of TrueBlot® secondaries over regular secondary antibodies?
TrueBlot® antibodies are specific to the whole IgG molecule and will not bind heavy or light chains. This is useful for binding target proteins with an expected MW near 25 kDa (light chains) and 50 kDa (heavy chains).
Why does it appear that the TrueBlot® antibody is binding heavy/light chains in my sample?
The sample must be fully reduced to eliminate cross-reactivity with heavy and light chains. Any reactivity to heavy and light chains could be due to incomplete reduction of your sample. Please be sure to optimize reducing conditions for your sample type.
How can I ensure my samples are fully reduced?
Samples can be fully reduced by heating at 90-100°C for 10 minutes in sample buffer containing reducing agent (for example, DTT to a final concentration of 50 mM or add β-Mercaptoethanol to a final concentration of 2% (v/v)).
With which species/isotypes are TrueBlot® secondary antibodies reactive?
TrueBlot® secondary antibodies are reactive with the IgG of their respective species. For example, if using a primary antibody raised in mouse (mouse host), use TrueBlot® anti-mouse Ig secondary antibodies for detection.
Do TrueBlot® secondary antibodies detect IgM?
TrueBlot® secondary antibodies have not been shown to detect IgM.
Do I need to preclear my lysate before the immunoprecipitation step?
Samples that have many other proteins present (such as lysates) may require preclearing to prevent interference in later IP and WB procedures. Recombinant protein samples require pre-clearing more often than serum samples.
How can I enrich/concentrate my lysate for a low expression of a protein of interest?
The immunoprecipitation procedure can be repeated several times to yield a more concentrated protein solution.
What is the recommended lysis buffer for TrueBlot® products?
The optimal lysis buffer will depend on your sample type. See protocol for buffer recommendations. Generally, 1X RIPA is used to lyse samples.
My target protein has the same MW as a heavy/light chain. How can I be sure that the band is the target protein and not a heavy/light chain?
Be sure to include a positive control for the primary antibody in your experiment and check that the sample is fully reduced.
What is the average size of TrueBlot® magnetic/agarose beads?
The beads are roughly 0.5 μm in diameter.
Why can’t I use Protein A or Protein G beads for the IP step in IP–Western blot when using rabbit species?
Using Protein A or Protein G beads with rabbit species results in contaminated bands. For best results when using rabbit species, use Rockland’s TrueBlot® Rabbit Ig IP beads, which are available in either a magnetic or agarose format. Please see individual TrueBlot® Rabbit product pages for additional details.
Can I use Protein A or Protein G beads for the IP step in IP–Western blot when using mouse, goat, or sheep species?
Yes. If you are using Protein A or Protein G beads with mouse, goat, or sheep species, we recommend the following: Determine the compatibility of Protein A or Protein G with your species and subtype, do not use excessive amounts of slurry, and choose the appropriate elution conditions. In some instances, harsher elution conditions can cause stripping of the subunits of Protein A or Protein G from the bead support and result in non-specific bands. For best results, we recommend using TrueBlot® Ig IP beads, which are available in either agarose or magnetic format.
Related Protocols
Adherent Cell Lysis Protocol
Bifunctional Cross-Linking Protocol
Chromatin Immunoprecipitation (ChIP) Protocol
ELISA Protocol
Fluorescent Western Blotting Protocol
Histone Immunoblotting Protocol
Immunocytochemistry (ICC) Protocol
Immunofluorescence (IF) Protocol
Immunoprecipitation (IP) Protocol
In-Cell Western (ICW) Protocol
IP-WB with TrueBlot® Protocol
Multi-Lysate Western Blotting Protocol
Nuclear & Cytoplasmic Extract Protocol
Nucleosome ChIP Protocol
Re-ChIP Protocol
Sandwich ELISA Protocol for Collagen
Suspension Cultured Cell Lysis Protocol
Western Blotting (WB) Protocol
Certificate of Analysis Lookup
Disclaimer
This product is for research use only and is not intended for therapeutic or diagnostic applications. Please contact a technical service representative for more information. All products of animal origin manufactured by Rockland Immunochemicals are derived from starting materials of North American origin. Collection was performed in United States Department of Agriculture (USDA) inspected facilities and all materials have been inspected and certified to be free of disease and suitable for exportation. All properties listed are typical characteristics and are not specifications. All suggestions and data are offered in good faith but without guarantee as conditions and methods of use of our products are beyond our control. All claims must be made within 30 days following the date of delivery. The prospective user must determine the suitability of our materials before adopting them on a commercial scale. Suggested uses of our products are not recommendations to use our products in violation of any patent or as a license under any patent of Rockland Immunochemicals, Inc. If you require a commercial license to use this material and do not have one, then return this material, unopened to: Rockland Inc., P.O. BOX 5199, Limerick, Pennsylvania, USA.
Related products to: Rabbit TrueBlot®: Anti-Rabbit IgG HRP
Fluorescent TrueBlot®: Anti-Rabbit IgG DyLight™ 800 18-4516-32 Fluorescent TrueBlot®: Anti-Mouse Ig DyLight™ 680 18-4417-32 Fluorescent TrueBlot®: Anti-Rabbit IgG DyLight™ 680 18-4416-32 Fluorescent TrueBlot®: Anti-Rabbit IgG Fluorescein 18-0216-32 Fluorescent TrueBlot®: Anti-Mouse Ig DyLight™ 800 18-4517-32 Related Resources
Selecting TrueBlot® Products
Literature
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IP-WB with TrueBlot® Protocol
Protocols
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TrueBlot®: Publication Quality IP and Western Blot Images
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