Blocking Agents offered by SPI Supplies
For the reduction of background staining using immunogold methods
Introduction:
Non-specific labelling can occur on specimens during immunolabelling
procedures. The source of this background labelling must be determined by
the careful and systematic use of controls and eliminated for a proper
analysis of the specimen. Background labelling can occur from a number of
sources, either in the specimen or in the incubating solutions (see list below).
In either case the background can be substantially reduced by the careful
use of blocking reagents.
- Background labelling from the specimen
- Hydrophobic attraction of proteins or gold particles
- Charge attraction from tissue components to antibodies or gold particles
- Fixing of antibodies through excess aldehyde groups in tissue
- Receptors in tissues for Fc components of antibodies
- Excess sulphur in tissue or support
- Excess lysine in tissue adhesive (for LM sections)
- Background labelling from incubating solutions
- Hydrophobic proteins (e.g. antibodies)
- Charged proteins (e.g. pH, ionic concentration)
- Charged gold particles
- Gelatin in buffer
Most of these sources of background can easily be overcome by the addition
of appropriate blocking reagents to the buffers used for incubating or by a
separate blocking step before the primary antibody.
Available blocking agents:
- Tween® 20
- BSA (fatty acid free)
- Gelatin (fish 45%)
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Monday March 22, 2010
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