Anti-Mouse CD8a (Ly 2) [Clone 53-6.7] — Purified in vivo GOLD™ Functional Grade

Anti-Mouse CD8a (Ly 2) [Clone 53-6.7] — Purified in vivo GOLD™ Functional Grade

Product No.: C375

[product_table name="All Top" skus="C375"]

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Clone
53-6.7
Target
CD8a
Formats AvailableView All
Product Type
Monoclonal Antibody
Alternate Names
T8, Lyt2, Ly-2
Isotype
Rat IgG2a κ
Applications
B
,
CyTOF®
,
Depletion
,
FC
,
IHC FF
,
in vivo
,
IP
,
PhenoCycler®
,
WB

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Data

Ly 2 Clone 53-6.7 Data ImageLy 2 Clone 53-6.7 Data Image
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Antibody Details

Product Details

Reactive Species
Mouse
Host Species
Rat
Recommended Isotype Controls
Recommended Dilution Buffer
Immunogen
Mouse thymus or spleen
Product Concentration
≥ 5.0 mg/ml
Endotoxin Level
< 1.0 EU/mg as determined by the LAL method
Purity
≥95% monomer by analytical SEC
>95% by SDS Page
Formulation
This monoclonal antibody is aseptically packaged and formulated in 0.01 M phosphate buffered saline (150 mM NaCl) PBS pH 7.2 - 7.4 with no carrier protein, potassium, calcium or preservatives added. Due to inherent biochemical properties of antibodies, certain products may be prone to precipitation over time. Precipitation may be removed by aseptic centrifugation and/or filtration.
Product Preparation
Functional grade preclinical antibodies are manufactured in an animal free facility using in vitro cell culture techniques and are purified by a multi-step process including the use of protein A or G to assure extremely low levels of endotoxins, leachable protein A or aggregates.
Storage and Handling
Functional grade preclinical antibodies may be stored sterile as received at 2-8°C for up to one month. For longer term storage, aseptically aliquot in working volumes without diluting and store at ≤ -70°C. Avoid Repeated Freeze Thaw Cycles.
Country of Origin
USA
Shipping
Next Day 2-8°C
Applications and Recommended Usage?
Quality Tested by Leinco
FC The suggested concentration for this 53-6.7 antibody for staining cells in flow cytometry is ≤ 0.25 μg per 106 cells in a volume of 100 μl. Titration of the reagent is recommended for optimal performance for each application.
WB The suggested concentration for this 53-6.7 antibody for use in western blotting is 1-10 μg/ml.
Additional Applications Reported In Literature ?
CyTOF®
CODEX®
IHC (Frozen)
IHC (Paraffin) Clone 53-6.7 has been reported for use in zinc-fixed paraffin-embedded sections and is NOT recommended for immunohistochemistry of formalin-fixed paraffin sections.
IP
B
Depletion
Each investigator should determine their own optimal working dilution for specific applications. See directions on lot specific datasheets, as information may periodically change.

Description

Description

Specificity
Clone 53-6.7 recognizes Lyt 2. Clone 53-6.7 competes with clone 5H10-1 for binding to thymocytes.
Background
CD8 is made up of disulfide-linked α and β chains that form the α(CD8a)/β(CD8b) heterodimer and α/α homodimer. CD8 is part of the Ig superfamily that expresses primarily as CD8a homodimers. CD8a is a 32-34 kD type I glycoprotein that can also form heterodimers with CD8b. CD8 is an antigen co-receptor on T cells that mediates efficient cell to cell interactions within the immune system. CD8 coupled with the T cell receptor on the T lymphocyte recognizes an antigen displayed by an antigen presenting cell (APC) in the context of class I MHC molecules. The CD8 co-receptor also plays a role in T cell signaling by interacting with Lck (lymphocyte-specific protein tyrosine kinase) which leads to the activation of transcription factors that affect the expression of certain genes.
Antigen Distribution
Lyt 2 is present on the surface of most thymocytes and a subpopulation of mature T-lymphocytes which include most T suppressor/cytotoxic cells.
Ligand/Receptor
MHC class I molecule
Function
Co-receptor for TCR
PubMed
NCBI Gene Bank ID

Leinco Antibody Advisor

Powered by AI: AI is experimental and still learning how to provide the best assistance. It may occasionally generate incorrect or incomplete responses. Please do not rely solely on its recommendations when making purchasing decisions or designing experiments.

Clone 53-6.7 is most commonly used in vivo in mice for depletion of CD8α+ cells (cytotoxic T cells), functional blocking of CD8+ T cell activity, and blocking antigen presentation via MHC class I.

Essential applications and supporting details:

  • CD8+ T cell depletion: 53-6.7 induces rapid and sustained depletion of CD8α-expressing cells, enabling experimental assessment of the role of cytotoxic T cells in immune responses, tumor rejection, transplantation models, and infection.
  • Blocking antigen presentation via MHC class I: The antibody binds mouse CD8α and blocks its interaction with MHC class I, interfering with antigen recognition and T cell activation.
  • Inhibition of CD8+ T cell responses: By blocking CD8α, clone 53-6.7 inhibits CD8+ T cell effector functions, such as proliferation and IL-2 production, and can suppress cytotoxic activity in vivo.
  • Functional studies in vivo: Used to dissect the contribution of CD8+ T cells to disease models, immune tolerance, and pathogen defense.
  • Other reported uses:
    • Blocking cytotoxicity in immune assays.
    • Immunoprecipitation and functional assays (when purified for these applications).
    • Quality control as a depletion agent in various immunological experiments.

Additional relevant information:

  • 53-6.7 is not recommended for immunohistochemistry on formalin-fixed paraffin sections but is validated for acetone-fixed frozen and zinc-fixed paraffin-embedded tissues.
  • The antibody is available in functional-grade formulations optimized for in vivo use and tested to ensure low endotoxin and aggregation for reliable T cell depletion/blocking.

In summary, in vivo applications of clone 53-6.7 in mice center on CD8+ T cell depletion, functional blocking, and studies of T cell–dependent immunity.

Commonly Co-Used Antibodies/Proteins with 53-6.7

The 53-6.7 antibody recognizes mouse CD8α, a coreceptor expressed on cytotoxic T cells and a subset of thymocytes. In experimental immunology, it is often used in combination with other antibodies and proteins to characterize immune cell populations or to facilitate functional studies. Here are the most commonly co-used reagents in the literature:

Frequently Co-Used Antibodies

  • Anti-CD4: Used alongside 53-6.7 to distinguish helper (CD4⁺) and cytotoxic (CD8⁺) T cell subsets within the broader T lymphocyte population. This is crucial for immunophenotyping in flow cytometry and immunohistochemistry.
  • Anti-CD3: A pan-T cell marker that identifies all mature T cells, often paired with 53-6.7 and anti-CD4 to further delineate T cell subsets.
  • Anti-CD45: The leukocyte common antigen, used to gate on all leukocytes or to distinguish T cells from other immune cell types.
  • Anti-CD8b: Used to distinguish between CD8αα homodimers and CD8αβ heterodimers, providing additional resolution in studies of T cell development and function.

Additional Markers and Controls

  • Isotype controls: Essential for establishing staining specificity, especially in flow cytometry and functional assays involving 53-6.7.
  • Other leukocyte markers (e.g., CD19 for B cells, CD11b for myeloid cells): Occasionally co-used to exclude non-T cell populations in complex immunophenotyping panels.

Co-Used Proteins and Functional Ligands

  • MHC class I molecules: The natural ligand for CD8, MHC class I is directly relevant in experiments where 53-6.7 is used to block or modulate antigen presentation and T cell activation.
  • IL-2: Since 53-6.7 can inhibit T cell responses to IL-2, studies involving T cell proliferation or cytokine responses often co-assay IL-2 or use anti-IL-2 antibodies.

Summary Table

Antibody/ProteinPurpose/Application
CD4Helper T cell identification
CD3Pan-T cell marker
CD45Leukocyte common antigen
CD8bDistinguish CD8αα vs. CD8αβ
Isotype controlSpecificity control for staining
MHC class ILigand for CD8; used in blocking/functional assays
IL-2T cell activation/proliferation assays

These combinations are standard in mouse immunology to characterize, isolate, and functionally manipulate CD8⁺ T cells in both in vitro and in vivo contexts. The exact panel will vary depending on the experimental question, but CD4, CD3, and CD45 are nearly universal partners for 53-6.7 in multiparameter flow cytometry and related techniques.

Clone 53-6.7 is a widely used monoclonal antibody specific for mouse CD8α, and key findings in the literature center on its diverse experimental uses and mechanistic properties.

  • Functional Blocking and Depletion: Clone 53-6.7 can block antigen presentation via MHC class I and inhibit T cell responses to interleukin-2 (IL-2). It is frequently used to functionally deplete or block CD8α⁺ T cells in vivo and in vitro, thereby allowing studies of CD8⁺ T cell roles in immunity or disease models.
  • Detection and Sorting: The antibody is a primary tool in flow cytometry, immunoprecipitation, and immunohistology to identify, quantify, or isolate CD8α⁺ T cell populations in mouse tissues.
  • Mechanistic Insights: CD8, recognized by 53-6.7, acts as a co-receptor binding to MHC class I and is essential for T cell activation and antigen-specific responses. Blocking CD8 can directly impact cytotoxicity and T cell proliferation.
  • Experimental Notes:
    • The antibody is not recommended for formalin-fixed paraffin sections but is suitable for frozen or zinc-fixed tissues.
    • Functional grade preparations are advised for in vivo experiments to minimize confounding effects of preservatives or endotoxin.
    • The antibody recognizes both CD8αα and CD8αβ forms, thus labeling a variety of immune cells beyond conventional CD8αβ cytotoxic T cells, including specific dendritic cells and intraepithelial lymphocytes.

Summary Table: Main Uses and Effects

ApplicationExperimental Effect/UseReference
CD8α⁺ cell depletion (in vivo/in vitro)Dissects role of CD8⁺ T cells in models
Blocking MHC class I/CD8 interactionsInhibits T cell activation/cytotoxicity, IL-2 response
Flow cytometry and immunostainingIdentifies and quantifies CD8α⁺ populations
ImmunoprecipitationStudies protein complexes involving CD8
Cell isolation/sortingEnables enrichment or depletion of CD8α⁺ cells

Overall, clone 53-6.7 is considered a key reagent for mouse immunology research, essential for studying CD8 T cell biology, immune responses, and experimental immunodepletion or blockade protocols.

The dosing regimens for clone 53-6.7, a rat anti-mouse CD8α monoclonal antibody, can vary significantly across different mouse models. This variation is influenced by several factors, including the specific experimental objective, mouse strain, route of administration, and whether the goal is acute depletion or chronic suppression of CD8+ T cells.

Factors Influencing Dosing Regimens

  1. Experimental Objective: The desired outcome, such as depletion of CD8+ T cells or inhibition of T cell responses, affects the dosing schedule.
  2. Mouse Strain: Different strains may require adjusted dosages due to varying immune responses.
  3. Route of Administration: Intraperitoneal (i.p.) or intravenous (i.v.) routes are commonly used, with i.p. being more frequent for in vivo studies.

Typical Dosing Ranges

While specific dosing ranges for clone 53-6.7 are not universally standardized, typical applications often involve administering doses in the range of 100 μg to several hundred micrograms per mouse. However, precise dosages can depend on the specific requirements of the study and are often determined experimentally.

Applications and Considerations

  • Applications: Clone 53-6.7 is used for blocking antigen presentation via MHC class I, inhibiting T cell responses to IL-2, and depleting CD8a+ cells.
  • Considerations: The efficacy and potential side effects of depleting CD8+ T cells must be considered, as these cells play a crucial role in immune responses, particularly against viral infections and tumors.

In summary, the dosing regimen for clone 53-6.7 in mouse models is highly dependent on the specific experimental context, and dosages should be optimized based on preliminary studies or established protocols for similar applications.

References & Citations

1.) Sarmiento, M. et al. (1980) Journal of Immunology 125(6):2665
2.) Gubin, M. et al. (2018) Cell. 175(4):1014–1030 Journal Link
3.) Sharma S. et al. (2020) Human Vaccines & Immunotherapeutics 16(9):2196-2203 Journal Link
B
CyTOF®
Depletion
Flow Cytometry
IHC FF
in vivo Protocol
Immunoprecipitation Protocol
PhenoCycler®
General Western Blot Protocol

Certificate of Analysis

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Disclaimer AlertProducts are for research use only. Not for use in diagnostic or therapeutic procedures.