OVA257-264 (SIINFEKL) peptide bound to H-2Kb Monoclonal Antibody (eBio25-D1.16 (25-D1.16)), PerCP-eFluor™ 710, eBioscience™

Product Specific Information

Description: The 25-D1.16 monoclonal antibody reacts with the ovalbumin-derived peptide SIINFEKL bound to H-2Kb of MHC class I, but not with unbound H-2Kb, or H-2Kb bound with an irrelevant peptide. This antibody has proven to be very useful tracking the quantity and localization of these specific antigen-presenting cells (APC) in vivo.^M
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Applications Reported: This eBio25-D1.16 (25-D1.16) antibody has been reported for use in flow cytometric analysis.^M
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Applications Tested: This eBio25-D1.16 (25-D1.16) antibody has been tested by flow cytometric analysis of SIINFEKL peptide-pulsed mouse splenocytes. This can be used at less than or equal to 0.125 µg per test. A test is defined as the amount (µg) of antibody that will stain a cell sample in a final volume of 100 µL. Cell number should be determined empirically but can range from 10e5 to 10e8 cells/test. It is recommended that the antibody be carefully titrated for optimal performance in the assay of interest.^M
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PerCP-eFluor® 710 can be used in place of PE-Cy5, PE-Cy5.5 or PerCP-Cy5.5. PerCP-eFluor® 710 emits at 710 nm and is excited with the blue laser (488 nm). Please make sure that your instrument is capable of detecting this fluorochrome. For a filter configuration, we recommend using the 685 LP dichroic mirror and 710/40 band pass filter, however the 695/40 band pass filter is an acceptable alternative.^M
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Our testing indicates that PerCP-eFluor® 710 conjugated antibodies are stable when stained samples are exposed to freshly prepared 2% formaldehyde overnight at 4°C, but please evaluate for alternative fixation protocols.^M
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Cells can be pulsed with the SIINFEKL peptide according to the following protocol:^M
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1. With cells in flow staining buffer, add SIINFEKL peptide to a final concentration of 30 uM.^M
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2. Incubate cells at 37°C for 2 hours.^M
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3. Wash cells with flow staining buffer.^M
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4. Proceed with cell surface staining as normal.^M
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For additional information see the references listed below.^M
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Excitation: 488 nm; Emission: 710 nm; Laser: Blue Laser.^M
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Filtration: 0.2 µm post-manufacturing filtered.