ImmunoCult™ Human CD3/CD28 T Cell Activator
Human T cell activation and expansion reagent
Overview
ImmunoCult™ Human CD3/CD28 T Cell Activator is designed to activate and expand human T cells in the absence of magnetic beads, feeder cells, or antigen. ImmunoCult™ Human CD3/CD28 T Cell Activator consists of soluble tetrameric antibody complexes that bind CD3 and CD28 cell surface ligands. Binding of the tetrameric antibody complexes results in the cross-linking of CD3 and CD28 cell surface ligands, thereby providing the required primary and co-stimulatory signals for T cell activation. Activated T cells can be expanded in ImmunoCult™-XF T Cell Expansion Medium or other media for culturing human T cells supplemented with cytokines.
The ImmunoCult™ Human CD3/CD28 T Cell Activator can be used on the Seahorse XF Analyzer to measure T cell activation response and is also available as part of the Agilent Seahorse XF Hu T Cell Activation Assay Kit.
This product is designed for cell therapy research applications following the recommendations of USP<1043> on Ancillary Materials, and we can currently work with you to qualify this reagent under an approved investigational new drug (IND) or clinical trial application (CTA).
The ImmunoCult™ Human CD3/CD28 T Cell Activator can be used on the Seahorse XF Analyzer to measure T cell activation response and is also available as part of the Agilent Seahorse XF Hu T Cell Activation Assay Kit.
This product is designed for cell therapy research applications following the recommendations of USP<1043> on Ancillary Materials, and we can currently work with you to qualify this reagent under an approved investigational new drug (IND) or clinical trial application (CTA).
Advantages
• Robust activation and expansion of human T cells without the use of magnetic beads, feeder cells, or antigen
• Provides a gentle activation stimulus that maintains high viability of activated and expanded T cells
• Highly stable, filter-sterilized soluble reagent
• Provides a gentle activation stimulus that maintains high viability of activated and expanded T cells
• Highly stable, filter-sterilized soluble reagent
Contains
• Anti-human CD3 monospecific tetrameric antibody complex
• Anti-human CD28 monospecific tetrameric antibody complex
• Anti-human CD28 monospecific tetrameric antibody complex
Subtype
Supplements
Cell Type
T Cells, T Cells, CD4+, T Cells, CD8+
Species
Human
Application
Activation, Cell Culture, Expansion
Brand
ImmunoCult
Area of Interest
Cell Therapy, Immunology
Related Products
Related Products
Scientific Resources
Product Documentation
| Document Type | Product Name | Catalog # | Lot # | Language |
|---|---|---|---|---|
|
Document Type
Product Information Sheet
|
Product Name
ImmunoCult™ Human CD3/CD28 T Cell Activator
|
Catalog # 10971, 10991 |
Lot # All |
Language English |
|
Document Type
Safety Data Sheet
|
Product Name
ImmunoCult™ Human CD3/CD28 T Cell Activator
|
Catalog # 10971, 10991 |
Lot # All |
Language English |
Educational Materials(18)
Brochure
T Cell Reagents for Your Cellular Therapy Research
Brochure
Isolate Human Immune Cells
Technical Bulletin
Optimization of Human T Cell Expansion Protocol: Effects of Early Cell Dilution
Wallchart
Production of Chimeric Antigen Receptor T Cells
Wallchart
Frequencies of Cell Types in Human Peripheral Blood
Wallchart
Human Immune Cytokines
Wallchart
Antigen Processing and Presentation
Wallchart
The Immune Response to HIV Poster
Video
4:30
How to Isolate Cells Directly from Whole Blood Using the EasySep™ Purple/Silver Magnets
Video
1:57
How EasySep™ Magnetic Cell Separation Technology Works: Fast and Easy Cell Isolation
Video
0:59
CAR T Cell Manufacturing Workflow: Isolation, Activation and Expansion
Video
1:37
How to Isolate PBMCs from Whole Blood Using Density Gradient Centrifugation (Ficoll™ or Lymphoprep™)
Webinar
47:14
Online Immunology Journal Club: Expanding the Therapeutic Tool Kit for CAR T Cells
Webinar
54:39
Qualification of Ancillary/Raw Materials for Clinical Use
Webinar
48:32
T Cell Differentiation and Cancer Immunity
Scientific Poster
Workflow Solutions for Human T Cell Isolation and Expansion
Scientific Poster
Rapid Expansion of Functional Human T Cells Using a Novel Serum-Free and Xeno-Free Culture Medium
Scientific Poster
Optimization of Human T Cell Activation and Expansion Protocols Improves Efficiency of Genetic Modification and Overall Cell Yield
Load More Educational Materials
Product Applications
This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.
Research Area
Workflow Stages
Workflow Stages for
T Cell Research
Workflow Stages for
T Cell Engineering
Workflow Stages for
T Cell Therapy Research
Data and Publications
Data
Figure 1. Activated Morphology of Human T Cells Stimulated With ImmunoCult™ Human CD3/CD28 T Cell Activator
Image of human T cells isolated using the EasySep™ Human T Cell Isolation Kit (Catalog #17951), stimulated with ImmunoCult™ Human CD3/CD28 T Cell Activator, and cultured in ImmunoCult™-XF T Cell Expansion Medium (Catalog # 10981).
Figure 2. Activation of EasySep™-isolated Human T Cells stimulated With ImmunoCult™ Human CD3/CD28 T Cell Activator
EasySep™-isolated human T cells were stimulated with ImmunoCult™ Human CD3/CD28 T Cell Activator and cultured in ImmunoCult™-XF T Cell Expansion Medium. Activation of viable CD3+ T cells was assessed by CD25 expression using flow cytometry. On day 0, the frequency of CD25 positive cells was (A) 5.6 ± 2.4% (mean ± SD). Following 3 days of culture, the frequency of CD25 positive cells was (B) 75.4 ± 13.8% (mean ± SD) when stimulated with ImmunoCult™ Human CD3/CD28 T Cell Activator.
Figure 3. Robust Human T Cell Expansion with ImmunoCult™ Human CD3/CD28 T Cell Activator
EasySep™-isolated human T cells were expanded over 12 days with ImmunoCult™ Human CD3/CD28 T Cell Activator in ImmunoCult™-XF T Cell Expansion Medium supplemented with Human Recombinant IL-2. On day 0, 1 x 10^6 EasySep™-isolated human T cells were stimulated with 25 μL of ImmunoCult™ Human CD3/CD28 T Cell Activator in ImmunoCult™-XF T Cell Expansion Medium supplemented with 10 ng/mL Human Recombinant IL-2. On days 3, 5, 7, and 10, viable cells were counted and fresh medium supplemented with IL-2 was added. No additional ImmunoCult™ Human CD3/CD28 T Cell Activator was added during the 12-day culture period (mean ± SD in 6 experiments with 3 donors).
Publications (6)
Science advances 2020 may
Competition between PAF1 and MLL1/COMPASS confers the opposing function of LEDGF/p75 in HIV latency and proviral reactivation.
Abstract
Abstract
Transcriptional status determines the HIV replicative state in infected patients. However, the transcriptional mechanisms for proviral replication control remain unclear. In this study, we show that, apart from its function in HIV integration, LEDGF/p75 differentially regulates HIV transcription in latency and proviral reactivation. During latency, LEDGF/p75 suppresses proviral transcription via promoter-proximal pausing of RNA polymerase II (Pol II) by recruiting PAF1 complex to the provirus. Following latency reversal, MLL1 complex competitively displaces PAF1 from the provirus through casein kinase II (CKII)-dependent association with LEDGF/p75. Depleting or pharmacologically inhibiting CKII prevents PAF1 dissociation and abrogates the recruitment of both MLL1 and Super Elongation Complex (SEC) to the provirus, thereby impairing transcriptional reactivation for latency reversal. These findings, therefore, provide a mechanistic understanding of how LEDGF/p75 coordinates its distinct regulatory functions at different stages of the post-integrated HIV life cycles. Targeting these mechanisms may have a therapeutic potential to eradicate HIV infection.
Cell systems 2020 mar
Gut-Liver Physiomimetics Reveal Paradoxical Modulation of IBD-Related Inflammation by Short-Chain Fatty Acids.
Abstract
Abstract
Although the association between the microbiome and IBD and liver diseases is known, the cause and effect remain elusive. By connecting human microphysiological systems of the gut, liver, and circulating Treg and Th17 cells, we created a multi-organ model of ulcerative colitis (UC) ex vivo. The approach shows microbiome-derived short-chain fatty acids (SCFAs) to either improve or worsen UC severity, depending on the involvement of effector CD4 T cells. Using multiomics, we found SCFAs increased production of ketone bodies, glycolysis, and lipogenesis, while markedly reducing innate immune activation of the UC gut. However, during acute T cell-mediated inflammation, SCFAs exacerbated CD4+ T cell-effector function, partially through metabolic reprograming, leading to gut barrier disruption and hepatic injury. These paradoxical findings underscore the emerging utility of human physiomimetic technology in combination with systems immunology to study causality and the fundamental entanglement of immunity, metabolism, and tissue homeostasis.
Scientific reports 2020 jun
A Simple and Scalable Strategy for Analysis of Endogenous Protein Dynamics.
Abstract
Abstract
The ability to analyze protein function in a native context is central to understanding cellular physiology. This study explores whether tagging endogenous proteins with a reporter is a scalable strategy for generating cell models that accurately quantitate protein dynamics. Specifically, it investigates whether CRISPR-mediated integration of the HiBiT luminescent peptide tag can easily be accomplished on a large-scale and whether integrated reporter faithfully represents target biology. For this purpose, a large set of proteins representing diverse structures and functions, some of which are known or potential drug targets, were targeted for tagging with HiBiT in multiple cell lines. Successful insertion was detected for 86{\%} of the targets, as determined by luminescence-based plate assays, blotting, and imaging. In order to determine whether endogenously tagged proteins yield more representative models, cells expressing HiBiT protein fusions either from endogenous loci or plasmids were directly compared in functional assays. In the tested cases, only the edited lines were capable of accurately reproducing the anticipated biology. This study provides evidence that cell lines expressing HiBiT fusions from endogenous loci can be rapidly generated for many different proteins and that these cellular models provide insight into protein function that may be unobtainable using overexpression-based approaches.
Stem Cell Research 2019 oct
Detection of all adult Tau isoforms in a 3D culture model of iPSC-derived neurons
Abstract
Abstract
Tauopathies are a class of neurodegenerative diseases characterized by the presence of pathological intracellular deposits of Tau proteins. Six isoforms of Tau are expressed in the adult human brain, resulting from alternative splicing of the MAPT gene. Tau splicing is developmentally regulated such that only the smallest Tau isoform is expressed in fetal brain, contrary to the adult brain showing the expression of all 6 isoforms. Induced Pluripotent Stem Cell (iPSC) technology has opened up new perspectives in human disease modeling, including tauopathies. However, a major challenge to in vitro recapitulation of Tau pathology in iPSC-derived neurons is their relative immaturity. In this study, we examined the switch in Tau splicing from fetal-only to all adult Tau isoforms during the differentiation of iPSC-derived neurons in a new 3D culture system. First, we showed that iPSC-induced neurons inside Matrigel-coated alginate capsules were able to differentiate into cortical neurons. Then, using a new assay that allowed both the qualitative and the quantitative analysis of all adult MAPT mRNA isoforms individually, we demonstrated that BrainPhys-maintained neurons expressed the 6 adult MAPT mRNA transcripts from 25 weeks of maturation, making this model highly suitable for modeling Tau pathology and therapeutic purposes.
Nature medicine 2018 OCT
Translational control of tumor immune escape via the eIF4F-STAT1-PD-L1 axis in melanoma.
Abstract
Abstract
Preventing the immune escape of tumor cells by blocking inhibitory checkpoints, such as the interaction between programmed death ligand-1 (PD-L1) and programmed death-1 (PD-1) receptor, is a powerful anticancer approach. However, many patients do not respond to checkpoint blockade. Tumor PD-L1 expression is a potential efficacy biomarker, but the complex mechanisms underlying its regulation are not completely understood. Here, we show that the eukaryotic translation initiation complex, eIF4F, which binds the 5' cap of mRNAs, regulates the surface expression of interferon-$\gamma$-induced PD-L1 on cancer cells by regulating translation of the mRNA encoding the signal transducer and activator of transcription 1 (STAT1) transcription factor. eIF4F complex formation correlates with response to immunotherapy in human melanoma. Pharmacological inhibition of eIF4A, the RNA helicase component of eIF4F, elicits powerful antitumor immune-mediated effects via PD-L1 downregulation. Thus, eIF4A inhibitors, in development as anticancer drugs, may also act as cancer immunotherapies.
Cancer cell 2018 FEB
Eradication of Triple-Negative Breast Cancer Cells by Targeting Glycosylated PD-L1.
Abstract
Abstract
Protein glycosylation provides proteomic diversity in regulating protein localization, stability, and activity; it remains largely unknown whether the sugar moiety contributes to immunosuppression. In the study of immune receptor glycosylation, we showed that EGF induces programmed death ligand 1 (PD-L1) and receptor programmed cell death protein 1 (PD-1) interaction, requiring beta$-1,3-N-acetylglucosaminyl transferase (B3GNT3) expression in triple-negative breast cancer. Downregulation of B3GNT3 enhances cytotoxic T cell-mediated anti-tumor immunity. A monoclonal antibody targeting glycosylated PD-L1 (gPD-L1) blocks PD-L1/PD-1 interaction and promotes PD-L1 internalization and degradation. In addition to immune reactivation, drug-conjugated gPD-L1 antibody induces a potent cell-killing effect as well as a bystander-killing effect on adjacent cancer cells lacking PD-L1 expression without any detectable toxicity. Our work suggests targeting protein glycosylation as a potential strategy to enhance immune checkpoint therapy.
Quality Statement:
PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED. FOR ADDITIONAL INFORMATION ON QUALITY AT STEMCELL, REFER TO WWW.STEMCELL.COM/COMPLIANCE.
