Human Exhausted CD8⁺ T Cell IHC Antibody Sampler Kit #43049

Kit Usage Information

Protocols

• 2203: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Paraffin), Immunofluorescence, Flow, ChIP Magnetic
• 7074: Western Blotting
• 15372: Western Blotting, Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin)
• 45208: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin), Flow Cytometry Live Cell Unconjugated Rabbit
• 46890: Western Blotting, Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin)
• 73758: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin)
• 85061: Western Blotting, Immunoprecipitation (Agarose), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin)
• 85336: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Paraffin), Flow
• 86163: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Leica® Bond™), Immunohistochemistry (Paraffin), Flow
• 99567: Western Blotting, Immunoprecipitation (Magnetic), Immunohistochemistry (Paraffin), Immunofluorescence, Flow Triton Permeabilization (Rabbit)

Product Description

The Human Exhausted CD8⁺ T Cell IHC Antibody Sampler Kit provides an economical means of characterizing the extent of exhaustion in T cells in formalin-fixed, paraffin-embedded tissue samples.

Specificity / Sensitivity

Each antibody in the Human Exhausted CD8⁺ T Cell IHC Antibody Sampler Kit detects endogenous levels of its target human protein. Tox/Tox2 (E6I3Q) Rabbit mAb does not cross-react with Tox3 or Tox4 proteins. TCF1/TCF7 (C63D9) Rabbit mAb does not recognize the dominant negative isoforms of TCF1/TCF7 lacking the amino-terminal β-catenin binding domain and does not cross-react with LEF1. Granzyme B (D6E9W) Rabbit mAb recognizes human Granzyme B protein and is also reactive with mouse Granzyme B; however, this antibody is not suggested for immunohistochemical analysis of mouse tissues. Instead, Granzyme B (E5V2L) Rabbit mAb (Mouse Specific) #44153 is recommended for IHC analysis of mouse tissue samples. Non-specific staining using Granzyme B (D6E9W) Rabbit mAb in the sweat glands has been observed. TIGIT (E5Y1W) XP^® Rabbit mAb cross-reacts with an unidentified protein of 42 kDa in some cell extracts.

Source / Purification

Monoclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding the carboxy terminus of human CD8α protein, Glu178 of human CD3ε protein, Ala522 of human Tox protein, Pro96 of human TCF1/TCF7 protein, and Ala274 of human PD-1 protein. Monoclonal antibodies are produced by immunizing animals with a recombinant protein specific to human Granzyme B protein, the carboxy terminus of human TIGIT protein, the extracellular domain of human TIM-3 protein, and the amino terminus of human LAG3 protein.

Background

Cluster of Differentiation 3 (CD3) is a multiunit protein complex expressed on the surface of T cells that directly associates with the T cell receptor (TCR). CD3 is composed of four polypeptides: ζ, γ, ε, and δ. Engagement of the TCR complex with antigens presented in major histocompatibility complexes induces tyrosine phosphorylation in the immunoreceptor tyrosine-based activation motif (ITAM) of CD3 proteins. CD3 phosphorylation is required for downstream signaling through ZAP-70 and p85 subunit of PI-3 kinase, leading to T cell activation, proliferation, and effector functions (1). CD8 is a transmembrane glycoprotein expressed primarily on cytotoxic T cells, but has also been described on a subset of dendritic cells in mice (2,3). On T cells, CD8 is a co-receptor for the TCR, and these two distinct structures are required to recognize antigen bound to MHC Class I. CD8 ensures specificity of the TCR–antigen interaction, prolongs the contact between the T cell and the antigen presenting cell, and recruits the tyrosine kinase Lck, which is essential for T cell activation (2). Tox, Tox2, and TCF1/TCF7 play key roles in T cell development. Tox is also induced by high antigen stimulation during chronic viral infection or cancer, regulating T cell persistence and exhaustion. TCF1/TCF7 preserves the effector function of exhausted T cells during viral infection or cancer. EOMES is a key transcription factor for memory T cells and for full effector differentiation of CD8⁺ T cells. The dynamic expression of these transcription factors help characterize the extent to which a T cell is exhausted and will respond to antigen stimulation (4-8). Granzyme B is a serine protease expressed by cytotoxic T lymphocytes and natural killer (NK) cells and is a key component of immune responses to pathogens and transformed cells (9). PD-1 (PDCD1, CD279), TIGIT (VSIG9, VSTM3), TIM-3 (HAVCR2), and LAG3 (CD223) are immune cell co-inhibitory receptors (also known as immune checkpoints) that negatively regulate T cell function and dampen the immune response to pathogens and cancer (10-15). In addition to activated T cells, PD-1 is expressed by activated B cells and monocytes. Following interaction with its ligands, PD-L1 and PD-L2, PD-1 is phosphorylated at ITIM and ITSM motifs leading to recruitment of protein tyrosine phosphatases SHP-1 and SHP-2 and suppression of TCR signaling. TIGIT is expressed at low levels on subsets of T cells and NK cells, and is upregulated at the protein level following activation of these cells. TIGIT marks exhausted T cells in the tumor microenvironment and during human immunodeficiency virus (HIV) infection. TIM-3 is expressed by exhausted T cells in the settings of chronic infection and cancer. Tumor-infiltrating macrophages and dendritic cells also express TIM-3. LAG3 is primarily expressed by activated CD4⁺ T cells, CD8⁺ T cells, FoxP3⁺ T regulatory cells (Tregs), and natural killer (NK) cells. Co-expression of multiple immune checkpoints help characterize the extent to which a T cell is exhausted and will respond to antigen stimulation. Therapeutic blockade of several of these immune checkpoint receptors is a promising strategy for neoplastic intervention by enabling anti-tumor immune responses (10-15).

1. Kuhns, M.S. et al. (2006) Immunity 24, 133-9.
2. Zamoyska, R. (1994) Immunity 1, 243-6.
3. Shortman, K. and Heath, W.R. (2010) Immunol Rev 234, 18-31.
4. Aliahmad, P. et al. (2012) Curr Opin Immunol 24, 173-7.
5. Yao, C. et al. (2019) Nat Immunol 20, 890-901.
6. Alfei, F. et al. (2019) Nature 571, 265-269.
7. Seo, H. et al. (2019) Proc Natl Acad Sci U S A 116, 12410-12415.
8. Wang, Y. et al. (2019) Front Immunol 10, 169.
9. Trapani, J.A. (2001) Genome Biol 2, REVIEWS3014.
10. Schildberg, F.A. et al. (2016) Immunity 44, 955-72.
11. Anderson, A.C. et al. (2016) Immunity 44, 989-1004.
12. Callahan, M.K. et al. (2016) Immunity 44, 1069-78.
13. Chen, L. and Flies, D.B. (2013) Nat Rev Immunol 13, 227-42.
14. Chauvin, J.M. et al. (2015) J Clin Invest 125, 2046-58.
15. Chew, G.M. et al. (2016) PLoS Pathog 12, e1005349.

Pathways & Proteins

Explore pathways + proteins related to this product.

Select Your Pathway/Protein Adherens Junction Dynamics T Cell Receptor Signaling Wnt/ß-Catenin Signaling Protein: O94900 Protein: P01732 Protein: P07766 Protein: P10144 Protein: P18627 Protein: P36402 Protein: Q15116 Protein: Q495A1 Protein: Q8TDQ0 Protein: Q96NM4

Database Links

UniProt ID: P07766 , P36402 , Q96NM4 , O94900 , P10144 , Q495A1 , P01732 , P18627 , Q15116 , Q8TDQ0

Entrez-Gene Id: 916 , 6932 , 84969 , 9760 , 3002 , 201633 , 925 , 3902 , 5133 , 84868

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