Biolegend, 137017, Brilliant Violet 421™ anti-mouse CD68 Antibody, 50μg

Mouse CD68, also known as macrosialin, is an 85-115 kD member of the lysosomal-associated membrane protein (LAMP) family. It is a heavily glycosylated and predominantly intracellular protein, mainly in late endosomes. Macrosialin is the murine homolog to the human macrophage glycoprotein CD68. It is expressed on tissue macrophages, Langerhans cells and at low levels on dendritic cells. Lamp proteins may have functions relating to cell-cell interaction or cell-ligand interaction. The biological function of CD68 is not completely understood.
50μg
Verified Reactivity: Mouse
Antibody Type: Monoclonal
Host Species: Rat
Immunogen: Purified Con A receptor glycoproteins from the P815 cell line
Formulation: Phosphate-buffered solution, pH 7.2, containing 0.09% sodium azide and BSA (origin USA).
Preparation: The antibody was purified by affinity chromatography and conjugated with Brilliant Violet 421™ under optimal conditions.
Concentration: 0.2 mg/ml
Storage & Handling: The antibody solution should be stored undiluted between 2°C and 8°C, and protected from prolonged exposure to light. Do not freeze.
Application: ICFC - Quality tested FC - Verified SB - Reported in the literature, not verified in house
Recommended Usage: Each lot of this antibody is quality control tested by intracellular immunofluorescent staining with flow cytometric analysis. For flow cytometric staining, the suggested use of this reagent is ≤0.5 µg per million cells in 100 µl volume. It is recommended that the reagent be titrated for optimal performance for each application. Brilliant Violet 421™ excites at 405 nm and emits at 421 nm. The standard bandpass filter 450/50 nm is recommended for detection. Brilliant Violet 421™ is a trademark of Sirigen Group Ltd. Learn more about Brilliant Violet™. This product is subject to proprietary rights of Sirigen Inc. and is made and sold under license from Sirigen Inc. The purchase of this product conveys to the buyer a non-transferable right to use the purchased product for research purposes only. This product may not be resold or incorporated in any manner into another product for resale. Any use for therapeutics or diagnostics is strictly prohibited. This product is covered by U.S. Patent(s), pending patent applications and foreign equivalents.
Excitation Laser: Violet Laser (405 nm)
Application Notes: Additional reported (for relevant formats) applications include: immunoprecipitation1, 2, Western Blot1, 2, immunohistochemical staining of frozen sections2 and paraformaldehyde-fixed paraffin-embedded sections3, and spatial biology (IBEX)9,10.
Additional Product Notes: Iterative Bleaching Extended multi-pleXity (IBEX) is a fluorescent imaging technique capable of highly-multiplexed spatial analysis. The method relies on cyclical bleaching of panels of fluorescent antibodies in order to image and analyze many markers over multiple cycles of staining, imaging, and, bleaching. It is a community-developed open-access method developed by the Center for Advanced Tissue Imaging (CAT-I) in the National Institute of Allergy and Infectious Diseases (NIAID, NIH).
Application References(PubMed link indicates BioLegend citation): Silva RP, et al. 1999. Biochem. J. 338:687. (IP, WB) Rabinowitz SS, et al. 1991. J. Exp. Med. 174:827. (IP, WB, IHC) Wu J, et al. 2008. P. Natl. Acad. Sci. USA 105:16934. (IHC) Kayama H, et al. 2012. PNAS. 109:5010. PubMed Park S, et al. 2013. Biomaterials. 34:598. PubMed Guiducci C, et al. 2013. J Exp Med. 210:2903. PubMed McKinstry SU, et al. 2014. J Neurosci. 34:9455. PubMed Li X, et al. 2015. J Am Heart Assoc. 6:4. PubMed Radtke AJ, et al. 2020. Proc Natl Acad Sci U S A. 117:33455-65. (SB) PubMed Radtke AJ, et al. 2022. Nat Protoc. 17:378-401. (SB) PubMed
Product Citations: Bonifacio JPPL, et al. 2022. J Virol. 96:e0087122. PubMed Chen J, et al. 2022. Adv Sci (Weinh). 9:e2200668. PubMed Gomez-Salinero JM, et al. 2022. Cell Stem Cell. 29:593. PubMed Cai B, et al. 2021. Mol Cancer. 20:165. PubMed Socodato R, et al. 2020. Sci Signal. 13: . PubMed Gangoso E, et al. 2021. Cell. 184:2454. PubMed
RRID: AB_2562949 (BioLegend Cat. No. 137017)
Structure: A member of the lysosomal-associated membrane protein (lamp) family.
Distribution: Expressed on tissue macrophages, Langerhans cells, and at low levels on dendritic cells.
Function: Involved in cell-cell interaction or cell-ligand interaction, still not completely understood.
Cell Type: Antigen-presenting cells, Dendritic cells, Langerhans cells, Leukocytes, Macrophages
Biology Area: Cell Biology, Immunology, Innate Immunity, Neuroscience, Neuroscience Cell Markers
Molecular Family: Adhesion Molecules, CD Molecules, Innate Immune Signaling
Antigen References: 1. Ramprasad MP, et al. 1996. Proc. Natl. Acad. Sci. USA 93:14833. 2. Smith MJ, et al. 1987. J. Cell. Sci. 87:113.
Gene ID: 12514
UniProt: View information about CD68 on UniProt.org
Clone: FA-11
Regulatory Status: RUO
Other Names: Macrosialin
Isotype: Rat IgG2a
Q: What is the F/P ratio range of our BV421™ format antibody reagents?
A: It is lot-specific. On average it ranges between 2-4.
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.