CD45RO Monoclonal Antibody (UCHL1), Super Bright™ 780, eBioscience™

Product Specific Information

Description: The UCHL1 monoclonal antibody reacts with human CD45RO, a 180 kDa isoform of CD45. CD45RO is expressed by most thymocytes, activated memory T cells, granulocytes and monocytes. CD22 is a ligand for CD45RO. Expression of CD45RO and CD45RA is used commonly to discriminate subsets of peripheral T cells.

Applications Reported: This UCHL1 antibody has been reported for use in flow cytometric analysis.

Applications Tested: This UCHL1 antibody has been pre-diluted and tested by flow cytometric analysis of normal human peripheral blood cells. This may be used at 5 µL (1.0 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 10^5 to 10^8 cells/test.

Super Bright 780 is a tandem dye that can be excited with the violet laser line (405 nm) and emits at 780 nm. 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 Complete Staining Buffer (Product # SB-4401) to minimize any non-specific polymer interactions. Please refer to the datasheet for Super Bright Staining Buffer for more information.

In some experiments, we have observed that compensation values for Super Bright 780-conjugated antibodies are higher in the violet 450/50 channel when using UltraComp eBeads microspheres (Product # 01-2222-42) as compared to single-color stained cells. In such circumstances, we would recommend setting compensation with cells. We have also observed this in some experiments using AbC Total Antibody Compensation beads (Product # A10497).

Light sensitivity: This tandem dye is sensitive to photo-induced oxidation. Please protect this vial and stained samples from light.

Fixation: Samples can be stored in IC Fixation Buffer (Product # 00-8222) (100 µL of cell sample + 100 µL 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: 405 nm; Emission: 780 nm; Laser: Violet Laser

Super Bright Polymer Dyes are sold under license from Becton, Dickinson and Company.

Target Information

CD45RO (UCHL1, ubiquitin carboxyl terminal esterase L1) is the shortest isoform of a receptor-type protein tyrosine phosphatase, CD45 glycoprotein. CD45 is crucial in lymphocyte development and antigen signaling, serving as an important regulator of Src-family kinases, promotes cell survival by modulating integrin-mediated signal transduction pathway and is also involved in DNA fragmentation during apoptosis. CD45 isoforms differ in their extracellular domains, whereas they share identical transmembrane and cytoplasmic domains. CD45 isoforms differ in their ability to translocate into the glycosphingolipid-enriched membrane domains and their expression depends on cell type and physiological state of the cell. CD45RO is expressed on macrophages, CD8+ T cells, activated T cells and myeloma cells. Besides the role in immunoreceptor signaling, CD45 is important in promoting cell survival by modulating integrin-mediated signal transduction pathway and is also involved in DNA fragmentation during apoptosis.