Goat IgG TrueBlot™, Mouse IgG TrueBlot™, Rabbit IgG TrueBlot™ & Sheep IgG TrueBlot™
It is easy and seamless to generate publication-quality data with TrueBlot™ - simply substitute the conventional HRPO blotting reagent with Goat TrueBlot™, Mouse TrueBlot™, Rabbit TrueBlot™ or Sheep TrueBlot™. TrueBlot™ is a horseradish peroxidase conjugated immunoblotting detection reagent, which enables unhindered detection of blotted target bands.
TrueBlot™ is ideal for use in protocols involving immunoblotting of immunoprecipitated proteins, because TrueBlot™ preferentially detects the native disulfide form of mouse IgG or rabbit IgG. TrueBlot™ reduces interference by the ~55 kDa heavy and ~23 kDa light chains of the immunoprecipitating antibody in IP/immunoblotting applications.
Applications include studies examining post-translational modification (e.g. phosphorylation or acetylation) or protein-protein interactions.
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Figure 1: Stat1 was immunoprecipitated from 0.5 ml of 1x107 Jurkat cells/ml with 3 µg rabbit anti-human Stat1. Precipitate from 5x105 cells was subjected to electrophoresis, transferred to an Immobilon membrane, and immunoblotted with anti-Stat1 using Rabbit TrueBlot™ and conventional HRP-conjugated detection. Lane 1: Detection with Rabbit IgG TrueBlot™ - note the absence of the anti-Stat1 immunoprecipitating heavy and light chains. Lane 2: re-blot of lane 1 using the conventional HRP-conjugated detection anti-rabbit polyclonal antibody - note the presence of the Stat1 immunoprecipitating heavy and light chains confirming that although the immunoprecipitating heavy and light chains were present in the sample in lane 1, Rabbit IgG TrueBlot™ detected native antibody but not the denatured heavy and light chains.
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Note that there are three key procedural considerations:
- use of anti-Ig beads for precipitation - never use Protein A or Protein G beads when using Rabbit TrueBlot.
Protein A or G should not be used for IP in conjunction with Rabbit TrueBlot because protein A and G contaminants bind to rabbit IgG with high affinity - causing strong contaminating bands. If a rabbit, mouse, rat or goat antibody is used for IP, use anti-rabbit Ig, anti-mouse Ig, anti-rat Ig or anti-goat Ig beads (respectively) for immunoprecipitation.
- complete reduction of immunoprecipitate.
- effective blocking of the immunoblot using milk as blocking protein, rather than BSA. BSA is not an effective blocker.
For more information, please click on the links below:
