Biolegend, 304202, Purified anti-human CD45RO Antibody, 100μg

CD45RO is a 180 kD single chain membrane glycoprotein. It is a splice variant of tyrosine phosphatase CD45, lacking the A, B, and C determinants. The CD45RO isoform is expressed on activated and memory T cells, some B cell subsets, activated monocytes/macrophages, and granulocytes. CD45RO enhances both T cell receptor and B cell receptor signaling mediated activation. CD45 and its isoforms non-covalently associate with lymphocyte phosphatase-associated phosphoprotein (LPAP) on T and B lymphocytes. CD45 has been reported to be associated with several other cell surface antigens including CD1, CD2, CD3, and CD4. CD45 has also been reported to bind galectin-1 and CD22. CD45 isoform expression can change in response to cytokines.
100μg
Verified Reactivity: Human
Reported Reactivity: Chimpanzee, Cynomolgus, Common Marmoset
Antibody Type: Monoclonal
Host Species: Mouse
Immunogen: IL-2 dependent T cell line, CA1
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide.
Preparation: The antibody was purified by affinity chromatography.
Concentration: 0.5 mg/ml
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C.
Application: FC - Quality tested WB, IP, IHC-P - Reported in the literature, not verified in house SB - Community verified
Recommended Usage: Each lot of this antibody is quality control tested by immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is ≤ 2.0 µg per million cells in 100 µl volume. It is recommended that the reagent be titrated for optimal performance for each application.
Application Notes: The UCHL1 antibody is commonly used in combination with antibodies against CD45RA to discern memory and naïve T cells. Additional reported applications (for the relevant formats) include: immunohistochemical staining of acetone-fixed frozen tissue sections5 and formalin-fixed paraffin-embedded tissue sections4, Western blotting2, and immunoprecipitation3. UCHL1 signal intensity may be reduced in the presence of True-Stain Monocyte Blocker™ (Cat. No. 426102). Clone S19021B is not impacted by Monocyte Blocker and is recommended as an alternative to UCHL1.
Additional Product Notes: For the use of this antibody in spatial biology applications, we have partnered with Lunaphore Technologies for demonstration of our antibodies on the COMET™. The COMET™ platform is an automated, end-to-end spatial biology solution developed for rapid and flexible multiplex tissue profiling. More information on the COMET™ and a complete list of our antibodies that have been demonstrated on the COMET™ can be found here.
Application References(PubMed link indicates BioLegend citation): Knapp W, et al. Eds. 1989. Leucocyte Typing IV. Oxford University Press. New York. (FC) Ishii T, et al. 2001. P. Natl. Acad. Sci. USA 98:12138. (WB) Ponsford M, et al. 2001. Clin. Exp. Immunol. 124:315. (IP) Yamada M, et al. 1996. Stroke 27:1155. (IHC) Sakkas LI, et al. 1998. Clin. Diagn. Lab. Immunol. 5:430. (IHC) Baba N, et al. 2010. Int. Immunol. 22:237. PubMed Thakral D, et al. 2008. J. Immunol. 180:7431. (FC) PubMed Weiss L, et al. 2010. P. Natl. Acad. Sci. USA 107:10632. PubMed Wu YY, et al. 2007. Infect. Immun. 75:4357. PubMed Mozaffarian N, et al. 2008. Rheumatology 47:1335. PubMed Roque S, et al. 2007. J. Immunol. 178:8028. PubMed Yoshino N, et al. 2000. Exp. Anim. (Tokyo) 49:97. (FC) Smith SH, et al. 1986. Immunology 58:63. (Immunogen) Peterson VM, et al. 2017. Nat. Biotechnol. 35:936. (PG)
Product Citations: Fischer JR, et al. 2023. Cell Rep Med. 4:100977. PubMed Rouers A, et al. 2021. Cell Rep Med. 2:100278. PubMed McIlwain DR, et al. 2021. Cell Host Microbe. 29:1828. PubMed Jaiswal A, et al. 2022. Cancer Cell. 40:524. PubMed Friebel E, et al. 2020. Cell. 181(7):1626-1642.e20. PubMed Kaufmann M, et al. 2021. Med. 2(3):296-312.e8. PubMed Delgobo M, et al. 2021. Front Immunol. 12:584538. PubMed Liu F, et al. 2020. J Immunother Cancer. 8:. PubMed Ferrian S, et al. 2021. Cell Rep Med. 2:100419. PubMed Guo T, et al. 2018. J Immunol. 200:500. PubMed Cobb DA, et al. 2022. J Immunother Cancer. 10:. PubMed Cosgrove PR, et al. 2021. Pediatr Res. . PubMed Wu Y, et al. 2007. Infect Immun. 75:4357. PubMed Xu-Monette ZY, et al. 2019. Cancer Immunol Res. 7:644. PubMed Hazenberg MD, et al. 2019. Blood Adv. 2.659722222. PubMed Del Alcazar D, et al. 2019. Cell Rep. 28:3047. PubMed Gee MH, et al. 2018. Cell. 172:549. PubMed Mozaffarian N, et al. 2008. Rheumatology. 47:1335. PubMed Wu Y, et al. 2007. Infect Immun . 178:2802. PubMed Chng MHY, et al. 2020. Immunity. 51(6):1119-1135.e5.. PubMed Singh AK, et al. 2017. Front Immunol. 0.513888889. PubMed Sergio Gonzalez‐Duque et al. 2018. Cell metabolism. 28(6):946-960 . PubMed White M, et al. 2015. J Immunol. 195: 1858-1867. PubMed Mishra A, et al. 2021. Cell. 184(13):3394-3409.e20. PubMed NULL, et al. 2022. Cell. 185:916. PubMed Bender C, et al. 2020. Sci Adv. :6. PubMed Martin E, et al. 2020. JCI Insight. :5. PubMed Li Z, et al. 2020. J Clin Lab Anal. 34:e23155. PubMed Chang MH, et al. 2021. Cell Rep. 37:109902. PubMed
RRID: AB_314418 (BioLegend Cat. No. 304202)
Structure: Tyrosine phosphatases, type I transmembrane, 180 kD (isoform of CD45 containing none of the A, B, or C determinants)
Distribution: Activated and memory T cells, B cell subsets, monocytes, macrophages, granulocytes
Function: Enhances TCR and BCR signaling
Ligand/Receptor: CD22
Cell Type: B cells, Granulocytes, Macrophages, Mesenchymal Stem Cells, Monocytes, T cells, Tregs
Biology Area: Cell Biology, Immunology, Inhibitory Molecules, Neuroscience, Neuroscience Cell Markers, Stem Cells
Molecular Family: CD Molecules
Antigen References: 1. Thomas M. 1989. Annu. Rev. Immunol. 7:339. 2. Trowbridge I, et al. 1994. Annu. Rev. Immunol. 12:85.
Gene ID: 5788
UniProt: View information about CD45RO on UniProt.org
Clone: UCHL1
Regulatory Status: RUO
Workshop: IV N31
Other Names: CD45RO
Isotype: Mouse IgG2a, κ
Q: If an antibody clone has been previously successfully used in IBEX in one fluorescent format, will other antibody formats work as well?
A: It’s likely that other fluorophore conjugates to the same antibody clone will also be compatible with IBEX using the same sample fixation procedure. Ultimately a directly conjugated antibody’s utility in fluorescent imaging and IBEX may be specific to the sample and microscope being used in the experiment. Some antibody clone conjugates may perform better than others due to performance differences in non-specific binding, fluorophore brightness, and other biochemical properties unique to that conjugate.
Q: Will antibodies my lab is already using for fluorescent or chromogenic IHC work in IBEX?
A: Fundamentally, IBEX as a technique that works much in the same way as single antibody panels or single marker IF/IHC. If you’re already successfully using an antibody clone on a sample of interest, it is likely that clone will have utility in IBEX. It is expected some optimization and testing of different antibody fluorophore conjugates will be required to find a suitable format; however, legacy microscopy techniques like chromogenic IHC on fixed or frozen tissue is an excellent place to start looking for useful antibodies.
Q: Are other fluorophores compatible with IBEX?
A: Over 18 fluorescent formats have been screened for use in IBEX, however, it is likely that other fluorophores are able to be rapidly bleached in IBEX. If a fluorophore format is already suitable for your imaging platform it can be tested for compatibility in IBEX.
Q: The same antibody works in one tissue type but not another. What is happening?
A: Differences in tissue properties may impact both the ability of an antibody to bind its target specifically and impact the ability of a specific fluorophore conjugate to overcome the background fluorescent signal in a given tissue. Secondary stains, as well as testing multiple fluorescent conjugates of the same clone, may help to troubleshoot challenging targets or tissues. Using a reference control tissue may also give confidence in the specificity of your staining.
Q: How can I be sure the staining I’m seeing in my tissue is real?
A: In general, best practices for validating an antibody in traditional chromogenic or fluorescent IHC are applicable to IBEX. Please reference the Nature Methods review on antibody based multiplexed imaging for resources on validating antibodies for IBEX.