Rat IgG2b kappa Isotype Control (eB149/10H5), Super Bright™ 780, eBioscience™, Invitrogen™

Description

Description: The rat IgG2b monoclonal antibody is useful as an isotype control immunoglobulin. Applications Reported: This eB149/10H5 antibody has been reported for use in flow cytometric analysis. Applications Tested: This eB149/10H5 antibody has been tested by flow cytometric analysis of normal human peripheral blood cells and mouse splenocytes. Use at the same concentration as the experimental antibody. Super Bright 780 is a tandem dye that can be excited with the violet laser line (405nm) and emits at 780nm. We recommend using a 780/60 bandpass filter. Please make sure that your instrument is capable of detecting this fluorochrome. When using two or more Super Bright dye-conjugated antibodies in a staining panel, it is recommended to use Super Bright Staining Buffer (cat. SB-4400) to minimize any non-specific polymer interactions. Fixation: Samples can be stored in IC Fixation Buffer (Product # 00-8222) (100μL of cell sample + 100μL of IC Fixation Buffer) or 1-step Fix/Lyse Solution (Product # 00-5333) for up to 3 days in the dark at 4°C with minimal impact on brightness and FRET efficiency/compensation. Some generalizations regarding fluorophore performance after fixation can be made, but clone specific performance should be determined empirically. Excitation: 405nm; Emission: 780nm; Laser: Violet Laser

The isotype of a primary antibody and the application it is being used in can result in background staining. Primary antibody background noise can be caused by binding to Fc receptors on target cells; by non-specific interactions with cellular proteins, carbohydrates, and lipids; or by cell autofluorescence. Isotype control antibodies can act as negative controls to help differentiate non-specific background signal from specific antibody signal because they have no relevant specificity to a target antigen. While isotype controls are most commonly used in flow cytometry, they are useful in other applications such as chromatin immunoprecipitation (ChIP), immunohistochemistry, and gel shifts. Isotype controls should match with the primary antibody species and isotype so that the level of specific staining by the primary antibody may be accurately determined. If using directly labeled primary antibodies, the isotype control works best if conjugated with the same label as the test antibody.