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EasySep™ Human Naïve CD8+ T Cell Enrichment Kit

Immunomagnetic negative selection kit

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From: 793 USD

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Immunomagnetic negative selection kit
From: 793 USD

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Overview

The EasySep™ Human Naïve CD8+ T Cell Enrichment Kit is designed to isolate naïve CD8+ T cells from fresh peripheral blood mononuclear cells by negative selection. Unwanted cells are targeted for removal with antibody complexes recognizing CD4, CD14, CD16, CD19, CD20, CD36, CD45RO, CD56, CD57, CD94, CD123, CD244, TCRγ/δ, glycophorin A and dextran-coated magnetic particles. The labeled cells are separated using an EasySep™ magnet without the use of columns. Desired cells are poured off into a new tube.
For even faster cell isolations, we recommend the new EasySep™ Human Naïve CD8+ T Cell Isolation Kit (19258) which isolates cells in as little as 30 minutes.
Advantages:
• Fast, easy-to-use and column-free
• Up to 92% purity
• Untouched, viable cells
Components:
  • EasySep™ Human Naïve CD8+ T Cell Enrichment Kit (Catalog #19158)
    • EasySep™ Human Naïve CD8+ T Cell Enrichment Cocktail, 1 mL
    • EasySep™ Human CD45RO Depletion Cocktail, 1 mL
    • EasySep™ D2 Magnetic Particles, 2 x 1 mL
  • RoboSep™ Human Naive CD8+ T Cell Enrichment Kit with Filter Tips (Catalog #19158RF)
    • EasySep™ Human Naïve CD8+ T Cell Enrichment Cocktail, 1 mL
    • EasySep™ Human CD45RO Depletion Cocktail, 1 mL
    • EasySep™ D2 Magnetic Particles, 5 x 1 mL
    • RoboSep™ Buffer (Catalog #20104)
    • RoboSep™ Filter Tips (Catalog #20125) x 2
Magnet Compatibility:
• EasySep™ Magnet (Catalog #18000)
• “The Big Easy” EasySep™ Magnet (Catalog #18001)
• Easy 50 EasySep™ Magnet (Catalog #18002)
• RoboSep™-S (Catalog #21000)
Subtype:
Cell Isolation Kits
Cell Type:
T Cells; T Cells, CD8+
Species:
Human
Sample Source:
PBMC
Selection Method:
Negative
Application:
Cell Isolation
Brand:
EasySep; RoboSep
Area of Interest:
Immunology

Scientific Resources

Educational Materials

(8)

Frequently Asked Questions

Can EasySep™ be used for either positive or negative selection?

Yes. The EasySep™ kits use either a negative selection approach by targeting and removing unwanted cells or a positive selection approach targeting desired cells. Depletion kits are also available for the removal of cells with a specific undesired marker (e.g. GlyA).

How does the separation work?

Magnetic particles are crosslinked to cells using Tetrameric Antibody Complexes (TAC). When placed in the EasySep™ Magnet, labeled cells migrate to the wall of the tube. The unlabeled cells are then poured off into a separate fraction.

Which columns do I use?

The EasySep™ procedure is column-free. That's right - no columns!

How can I analyze the purity of my enriched sample?

The Product Information Sheet provided with each EasySep™ kit contains detailed staining information.

Can EasySep™ separations be automated?

Yes. RoboSep™, the fully automated cell separator, automates all EasySep™ labeling and cell separation steps.

Can EasySep™ be used to isolate rare cells?

Yes. We recommend a cell concentration of 2x108 cells/mL and a minimum working volume of 100 µL. Samples containing 2x107 cells or fewer should be suspended in 100 µL of buffer.

Are the EasySep™ magnetic particles FACS-compatible?

Yes, the EasySep™ particles are flow cytometry-compatible, as they are very uniform in size and about 5000X smaller than other commercially available magnetic beads used with column-free systems.

Can the EasySep™ magnetic particles be removed after enrichment?

No, but due to the small size of these particles, they will not interfere with downstream applications.

Can I alter the separation time in the magnet?

Yes; however, this may impact the kit's performance. The provided EasySep™ protocols have already been optimized to balance purity, recovery and time spent on the isolation.

For positive selection, can I perform more than 3 separations to increase purity?

Yes, the purity of targeted cells will increase with additional rounds of separations; however, cell recovery will decrease.

How does the binding of the EasySep™ magnetic particle affect the cells? is the function of positively selected cells altered by the bound particles?

Hundreds of publications have used cells selected with EasySep™ positive selection kits for functional studies. Our in-house experiments also confirm that selected cells are not functionally altered by the EasySep™ magnetic particles.

If particle binding is a key concern, we offer two options for negative selection. The EasySep™ negative selection kits can isolate untouched cells with comparable purities, while RosetteSep™ can isolate untouched cells directly from whole blood without using particles or magnets.
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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.

Data and Publications

Data

Start: 98% CD8+CD45RA+CCR7+CD45RO-CD56-CD57- of total peripheral blood mononuclear cells (PBMC)

Figure 1. Typical EasySep™ Human Naïve CD8+ T Cell Enrichment Profile

Starting with fresh mononuclear cells, the naïve CD8+ T cell (CD8+CD45RA+CCR7+CD45RO-CD56-CD57-) content of the enriched fraction typically ranges from 85 - 92%. In the example above, the purities of the start and enriched fractions are 9.8% and 91.4%, respectively.

Publications

(2)
Cancer research 2017 NOV

CD155T/TIGIT Signaling Regulates CD8+ T-cell Metabolism and Promotes Tumor Progression in Human Gastric Cancer.

He W et al.

Abstract

The T-cell surface molecule TIGIT is an immune checkpoint molecule that inhibits T-cell responses, but its roles in cancer are little understood. In this study, we evaluated the role TIGIT checkpoint plays in the development and progression of gastric cancer. We show that the percentage of CD8 T cells that are TIGIT+ was increased in gastric cancer patients compared with healthy individuals. These cells showed functional exhaustion with impaired activation, proliferation, cytokine production, and metabolism, all of which were rescued by glucose. In addition, gastric cancer tissue and cell lines expressed CD155, which bound TIGIT receptors and inactivated CD8 T cells. In a T cell-gastric cancer cell coculture system, gastric cancer cells deprived CD8 T cells of glucose and impaired CD8 T-cell effector functions; these effects were neutralized by the additional glucose or by TIGIT blockade. In gastric cancer tumor cells, CD155 silencing increased T-cell metabolism and IFNγ production, whereas CD155 overexpression inhibited T-cell metabolism and IFNγ production; this inhibition was neutralized by TIGIT blockade. Targeting CD155/TIGIT enhanced CD8 T-cell reaction and improved survival in tumor-bearing mice. Combined targeting of TIGIT and PD-1 further enhanced CD8 T-cell activation and improved survival in tumor-bearing mice. Our results suggest that gastric cancer cells inhibit CD8 T-cell metabolism through CD155/TIGIT signaling, which inhibits CD8 T-cell effector functions, resulting in hyporesponsive antitumor immunity. These findings support the candidacy of CD155/TIGIT as a potential therapeutic target in gastric cancer. Cancer Res; 77(22); 6375-88. textcopyright2017 AACR.
Journal of immunology (Baltimore, Md. : 1950) 2016 MAR

Enhanced Cytotoxic CD8 T Cell Priming Using Dendritic Cell-Expressing Human Papillomavirus-16 E6/E7-p16INK4 Fusion Protein with Sequenced Anti-Programmed Death-1.

Garcia-Bates TM et al.

Abstract

The incidence of human papillomavirus (HPV)-related head and neck squamous cell carcinoma has increased in recent decades, though HPV prevention vaccines may reduce this rise in the future. HPV-related cancers express the viral oncoproteins E6 and E7. The latter inactivates the tumor suppressor protein retinoblastoma (Rb), which leads to the overexpression of p16(INK4) protein, providing unique Ags for therapeutic HPV-specific cancer vaccination. We developed potential adenoviral vaccines that express a fusion protein of HPV-16 E6 and E7 (Ad.E6E7) alone or fused with p16 (Ad.E6E7p16) and also encoding an anti-programmed death (PD)-1 Ab. Human monocyte-derived dendritic cells (DC) transduced with Ad.E6E7 or Ad.E6E7p16 with or without Ad.αPD1 were used to activate autologous CD8 CTL in vitro. CTL responses were tested against naturally HPV-infected head and neck squamous cell carcinoma cells using IFN-γ ELISPOT and [(51)Cr]release assay. Surprisingly, stimulation and antitumor activity of CTL were increased after incubation with Ad.E6E7p16-transduced DC (DC.E6E7p16) compared with Ad.E6E7 (DC.E6E7), a result that may be due to an effect of p16 on cyclin-dependent kinase 4 levels and IL-12 secretion by DC. Moreover, the beneficial effect was most prominent when anti-PD-1 was introduced during the second round of stimulation (after initial priming). These data suggest that careful sequencing of Ad.E6E7.p16 with Ad.αPD1 could improve antitumor immunity against HPV-related tumors and that p16 may enhance the immunogenicity of DC, through cyclin-dependent pathways, Th1 cytokine secretion, and by adding a nonviral Ag highly overexpressed in HPV-induced cancers.
STEMCELL TECHNOLOGIES INC.’S QUALITY MANAGEMENT SYSTEM IS CERTIFIED TO ISO 13485. PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED.