Mouse TrueBlot® Western Blot Kit
Mouse TrueBlot® IP / Western Blot
Mouse TrueBlot® Western Blot Kit
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Datasheet

Mouse TrueBlot® Western Blot Kit

Rat Monoclonal eB144

88-8887-31
1 Kit
IP, WB
Mouse
Rat
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Description

Background

The Mouse TrueBlot® Western Blot Kit contains the critical supporting reagents, buffers, and substrates for immunoprecipitation and Western blotting of samples using TrueBlot monoclonal secondary antibody in conjunction with your own primary IP antibody and primary (Mouse IgG) Western blotting antibody (see application note for kit components). Mouse IgG TrueBlot® ULTRA is the unique horseradish peroxidase conjugated Anti-Mouse IgG monoclonal secondary antibody which enables detection of immunoblotted target protein bands, without hindrance by interfering immunoglobulin heavy and light chains from your IP antibody. Use it in place of your conventional HRP Anti-Mouse IgG immunoblotting secondary antibody. It is easy to generate publication-quality IP/WB data with Mouse IgG TrueBlot® ULTRA. TrueBlot technology enables unhindered detection of protein bands of interest which would otherwise be obscured by the presence of reduced and denatured heavy and light chain immunoglobulin in the blot (as detected by the conventional immunoblotting HRP anti-mouse IgG reagent). Mouse IgG TrueBlot ULTRA is ideal for use in protocols involving immunoblotting of immunoprecipitated proteins. TrueBlot preferentially detects the non-reduced form of mouse IgG over the reduced, SDS-denatured form of IgG. When the immunoprecipitate is fully reduced immediately prior to SDS-gel electrophoresis, reactivity of Mouse IgG TrueBlot ULTRA 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/immunoblot applications. Applications include studies examining post-translational modification (e.g., phosphorylation or acetylation) or protein-protein interactions.

Application Note

Mouse IgG TrueBlot® ULTRA is provided as 1000X solution. To achieve best results when detecting mouse IgG1 subtypes, we recommend performing a dot blot or titration to determine the optimal dilution factor for your desired application. 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. 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 2.5mls/blot will yield enough reagent for 20 blots. Mouse IgG TrueBlot® ULTRA HRP-conjugated secondary antibody reacting with mouse IgGs for optimal signal detection in immunoprecipitation/immunoblotting experiments. Note that there are three key procedural considerations: 1. Protein A or G beads may be used with the mouse, goat and sheep TrueBlot secondaries, but not with the rabbit TrueBlot secondary. Use of protein A or G beads with the rabbit TrueBlot will result in contaminating bands. 2. Immunoprecipitate should be completely reduced. 3. BLOTTO/Milk should be used as the blocking protein for the immunoblot. A sample of Anti-Alpha-Tubulin (MOUSE) Monoclonal Antibody Cat #(200-301-880S), has been provided to be used as a loading control at the user’s discretion. Dilutions are to be optimized by the user. Mouse TrueBlot® Western Blot Kit Components: 1. Mouse IgG TrueBlot ULTRA: 50 μL 18-8817-31, 2. TrueBlot Enhancer Solution: 25 mL; 3. TrueBlot Blocker: 10 g; 4. TrueBlot Assay Buffer: 30 mL, 20X; 5. TrueBlot Substrate A: 12.5 mL; 6. TrueBlot Substrate B: 12.5 mL; 7. Western Blot Incubation Tray. 8. Anti-Alpha-Tubulin (MOUSE) Monoclonal Antibody - 200-301-880S.

Purity/Specificity

Mouse TrueBlot® ULTRA 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-Mouse Serum. Reactivity is observed against native Mouse IgG by both Western blot and ELISA.

Mouse TrueBlot® Western Blot Kit - 88-8887-31
Anti-Mouse IgG HRP, TrueBlot, HRP TrueBlot ULTRA, Anti-Mouse IgG Peroxidase TrueBlot, TrueBlot for IP/WB, TrueBlot for immunoprecipitation, TrueBlot for western blotting, TrueBlot for chemiluminescent western blotting
IP, WB
Mouse
Rat
Peroxidase (Horseradish) ULTRA
Monoclonal
eB144
IgG
Immunoprecipitation Kit
Liquid (sterile filtered)
0.01 M Sodium Phosphate, 0.15 M Sodium Chloride, pH 7.2
0.1 mg/ml Bovine Serum Albumin (BSA) - IgG and Protease free, 50% (v/v) Glycerol
Wet Ice
Store Kit at 2-8 °C, except Mouse TrueBlot® ULTRA which should be stored 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 opening.

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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.