Isotype Controls:Isotype controls are a type of negative control designed to measure the level of non-specific background signal caused by primary antibodies, based upon the tissue type of the sample. Non-specific binding of primary antibodies are typically caused by the following reasons:
Non-specific Fc receptor binding to cells
Non-specific antibody interactions with cellular proteins or fluorochromes
Non-specific binding of the antibody or fluorochromes to cellular components
Cell autofluorescence
Usually, the background signal is the result of immunoglobulins binding non-specifically to Fc receptors present on the cell surface.
For example, antibodies raised in mice, particularly those of the IgG2a isotype, will bind strongly to some human leukocytes regardless of the test antibody specificity.
In this case you would need a mouse IgG2a isotype control for use with human cells or tissues.
Isotype controls are most commonly used in flow cytometry experiments, but may also be used in immunohistochemistry.
Selecting an Isotype Control:Typically, an isotype control is matched to the host species and isotype of your specific primary antibody, the heavy chain (IgA, IgG, IgD, IgE, or IgM), subclass and light chain (kappa, lambda) class of the primary antibody; the same fluorochrome (PE, APC, etc.), and have the same F:P ratio.
F:P is a measurement of how many fluorescent molecules are present on each antibody.
When using directly labeled primary antibodies, it is also necessary to make sure that the isotype control is conjugated to the same fluorochrome or label as the test antibody.
It is not sufficient to use one with a spectrally similar fluorochrome.
Non-specific Binding:To avoid non-specific binding, you also need to block Fc receptors on cell types, such as spleen cells, with FcR blocking reagents.
Isotype controls should be used at the same concentration/dilution as the primary antibody.
Isotype controls are mainly used to determine if the staining is real.
Apart from isotype controls, unstained cells should always be included in the experimental set-up to monitor autofluorescence.
Applications for Isotype Controls:Although isotype controls are mainly used in flow cytometry, they can be used as standard blocking agents and protein coating agents for other applications including immunofluorescence, immunocytochemistry, western blotting, and ELISA.
Goat IgG (Whole Molecule) Control is highly purified from normal goat serum through Protein G chromatography.
IgG (Whole Molecule) Control is suitable to be used as control, standard, blocking agent, or coating protein in a variety of assays, including ELISA, immunobloting (Dot blot and Western blot), immunoprecipitation, immunodiffusion, and immunoelectrophoresis.
It also may be used as antigen or ligand in immunochemical conjugation reaction.
Applications:Suitable for use in ELISA, Dot Blot, Western Blot, Immunoprecipitation, Immunoelectrophoresis and Immunodiffusion.
Can also be used as a control, standard, blocking agent, or coating protein.
It also may be used as antigen or ligand in immunochemical reactions and conjugations.
Other applications not tested.
Recommended Dilutions:Optimal dilutions to be determined by the researcher.
Storage and Stability:Lyophilized powder may be stored at -20°C.
Stable for 12 months after receipt at -20°C.
Reconstitute with sterile ddH2O.
Aliquot to avoid repeated freezing and thawing.
Store at -20°C.
Reconstituted product is stable for 6 months at -20°C.
For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
Further dilutions can be made in assay buffer.
仕様
Size：4mg
Host：goat
Grade：Affinity Purified
Purity：Purified by Protein G affinity chromatography
Form：Supplied as a lyophilized powder in PBS, pH 7.4, 0.02% sodium azide. Reconstitute with 1ml sterile ddH2O.
