CellXVivo Human Th1 Cell Differentiation Kit

For the differentiation of Th1 cells from a preparation of naïve CD4+ T cells
Catalog # Availability Size / Price Qty
CDK001
CellXVivo Human Th1 Cell Differentiation Kit
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Product Details
Procedure
Citations (4)
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CellXVivo Human Th1 Cell Differentiation Kit Summary

Kit Summary

For the differentiation of Th1 cells from a preparation of naïve CD4+ T cells.

Key Benefits

  • Yields a highly enriched population of Th1 polarized T cells
  • Provides optimized reagents needed to induce Th1 differentiation
  • Involves validated and straightforward procedures
  • Does not require specialized instrumentation
 

Why Differentiate Th1 Cells In Vitro?

T helper type 1 (Th1) cells are a subset of CD4+ effector T cells that promote cell-mediated immune responses against intracellular viral and bacterial pathogens. Differentiation into the Th1 lineage is promoted by cytokines such as IL-12 and IFN-gamma. Th1 cells are characterized by the secretion of IFN-gamma, IL-10, and TNF-alpha. Although Th1 cells are critical for the clearance of intracellular pathogens, exaggerated Th1 responses are associated with autoimmune diseases including rheumatoid arthritis, multiple sclerosis, inflammatory bowel disease, and type 1 diabetes. Th1 polarized cells are present in low abundance in normal human peripheral blood. In vitro differentiation of Th1 cells from the larger naïve CD4+ T cell population provides increased numbers of Th1 cells to facilitate downstream research.

Kit Contents

This kit contains the following optimized reagents to drive efficient differentiation of naïve CD4+ T cells into Th1 polarized cells.

  • Mouse Anti-Human CD3 Antibody
  • Human Th1 Differentiation Supplements
  • Reconstitution Buffers
  • Wash Buffer (20X)

Provides sufficient reagents for the differentiation of one 24-well plate.

 

Stability and Storage

Store the unopened kit at 2 to 8 °C. After opening the kit, Mouse Anti-Human CD3 Antibody, Human Th1 Reagent 1, and Human Th1 Reagent 2 may be stored at 2-8 °C under sterile conditions for up to 30 days or at -20 °C to -70 °C in a manual defrost freezer for up to 3 months. Reconstitution Buffer 1, Reconstitution Buffer 2, and 20X Wash Buffer may be stored under sterile conditions for up to 12 months at 2 to 8 °C. Do not use beyond the expiration date of the kit.

 

Data Examples
Detection of IFN-gamma, IL-4, and IL-17 in Human Th1 Cells
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Verification of Th1 Cell Identity using Flow Cytometry. Human peripheral blood naïve CD4+ T cells without (A, C) and with (B, D) a 5 day differentiation using the reagents included in the Human Th1 Cell Differentiation Kit. (A, B) The cells were stained with an APC-conjugated Mouse Anti-Human IFN-gamma Monoclonal Antibody (Catalog # IC285A) and a PE-conjugated Mouse Anti-Human IL-4 Monoclonal Antibody (Catalog # IC204P). (C, D) The cells were stained with an APC-conjugated Mouse Anti-Human IFN-gamma Monoclonal Antibody (Catalog #IC285A) and a PerCP-conjugated Mouse Anti-Human IL-17 Monoclonal Antibody (Catalog #IC3171C). Control cultures were used to place the quadrants.

Detection of IFN-gamma, IL-4, and IL-17 in Human Th1 Cells
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Detection of IFN-gamma, IL-4, and IL-17 in Human Th1 Cells. Human peripheral blood naïve CD4+ T cells were cultured for 5 days using reagents included in the Human Th1 Cell Differentiation Kit. After fixation of cells in the wells, cytokine expression was determined using the Human IFN-gamma Quantikine ELISA Kit (Catalog #DIF50), the Human IL-4 Quantikine ELISA Kit (Catalog #D4050), and the Human IL-17 Quantikine ELISA Kit (Catalog #D1700).

Specifications

Shipping Conditions
The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.
Storage
Store the unopened product at 2 - 8 °C. Do not use past expiration date.
Species
Human

Product Datasheets

Assay Procedure

Refer to the product datasheet for complete product details.

Briefly, human CD4+ T cells can be differentiated into Th1 cells using the following procedure:

  • Coat a plate with Mouse Anti-Human CD3 Antibody
  • Isolate human naïve CD4+ T cells from a PBMC preparation
  • Culture naïve CD4+ T cells in Human Th1 Differentiation Media for 5 days
  • Stimulate cells with monensin
  • Analyze culture media and cells for a Th1-specific cytokine production profile
 

 

Reagents Provided

Reagents Supplied in the CellXVivo™ Human Th1 Cell Differentiation Kit (Catalog # CDK001):

  • Mouse Anti-Human CD3 Antibody
  • Human Th1 Reagents
  • Reconstitution Buffers
  • Wash Buffer (20X)

 

Other Supplies Required

Reagents

  • MagCellect™ Human Naïve CD4+ T Cell Isolation Kit (Catalog # MAGH115 or equivalent)
  • Monensin
  • PMA
  • Ficoll-Hypaque™
  • RPMI 1640
  • L-Glutamine
  • Ionomycin
  • Penicillin
  • Streptomycin
  • Fetal Bovine Serum (FBS)
  • beta-Mercaptoethanol

Equipment

  • Tissue culture flasks and/or plates
  • Hemocytometer
  • 37 °C and 5% CO2 humidified incubator
  • Centrifuge
  • Pipettes and pipette tips

 

Procedure Overview

Coat wells of a 24-well plate with Mouse Anti-Human CD3 Antibody.

Coat wells of a 24-well plate

Isolate PBMCs from human blood (e.g., using Ficoll-Hypaque™ density centrifugation).

Isolate PBMCs from human blood

Isolate human naïve CD4+ T cells from PBMCs (e.g., using R&D Systems MagCellect™ Human Naïve CD4+ T Cell Isolation Kit).

Isolate T Cells from PBMCs

Perform a cell count.

Perform a cell count

Suspend 1-2 x 105 naïve CD4+ T cells/mL in media supplemented with Human Th1 Reagent 1 and Human Th1 Reagent 2.
Culture the cells on plates pre-coated with Mouse Anti-Human CD3 Antibody for 5 days.

Refresh the Diferentiation Media every 3-4 days.

Suspend cells in media

Stimulate the cells with mitogens.

Stimulate the cells with mitogens

Verify Th1 cell differentiaiton by analying cytokine expression via flow cytometry.

Verify Th1 cell differentiation

Citations for CellXVivo Human Th1 Cell Differentiation Kit

R&D Systems personnel manually curate a database that contains references using R&D Systems products. The data collected includes not only links to publications in PubMed, but also provides information about sample types, species, and experimental conditions.

4 Citations: Showing 1 - 4
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  1. Identification of Novel Molecular Markers of Human Th17 Cells
    Authors: A Sa?kowska, K Kara?, I Karwaciak, A Walczak-Dr, M Krawczyk, M Sobalska-K, J Dastych, M Ratajewski
    Cells, 2020;9(7):.  2020
  2. Intracellular Delivery of Anti-pPKCtheta (Thr538) via Protein Transduction Domain Mimics for Immunomodulation.
    Authors: Ozay E, Gonzalez-Perez G, Torres J, Vijayaraghavan J, Lawlor R, Sherman H, Garrigan D, Burnside A, Osborne B, Tew G, Minter L
    Mol Ther, 2016;24(12):2118-2130.  2016
  3. Biotin Deficiency Induces Th1- and Th17-Mediated Proinflammatory Responses in Human CD4(+) T Lymphocytes via Activation of the mTOR Signaling Pathway.
    Authors: Elahi A, Sabui S, Narasappa N, Agrawal S, Lambrecht N, Agrawal A, Said H
    J Immunol, 0;200(8):2563-2570.  0
  4. Naive CD4(+) T Cells Carrying a TLR2 Agonist Overcome TGF-beta-Mediated Tumor Immune Evasion.
    Authors: Ibrahim M, Scozzi D, Toth K, Ponti D, Kreisel D, Menna C, De Falco E, D'Andrilli A, Rendina E, Calogero A, Krupnick A, Gelman A
    J Immunol, 0;200(2):847-856.  0

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