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EasySep™人Naïve CD4+ T细胞分选试剂盒

人Naïve CD4+ T细胞的免疫磁珠负选
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产品号 #19555_C

人Naïve CD4+ T细胞的免疫磁珠负选

产品优势

  • 操作简单、快捷，且无需分离柱
  • 纯度高达96%
  • 分选得到的细胞无标记

产品组分包括

  • EasySep™ 人 Naïve CD4+ T细胞分选试剂盒（产品号 #19555）
    • EasySep™ 人Naïve CD4+ T细胞分选抗体混合物，1 mL
    • EasySep™ 生物素标记的抗CD45RO抗体，1 mL
    • EasySep™ Dextran RapidSpheres™磁珠，1 mL
  • RoboSep™人Naïve CD4+ T细胞分选试剂盒（产品号 #19555RF）
    • EasySep™人Naïve CD4+ T细胞分选抗体混合物，1 mL
    • EasySep™ 生物素标记的抗CD45RO抗体，1 mL
    • EasySep™ Dextran RapidSpheres™磁珠，1 mL
    • RoboSep™ 缓冲液（产品号 #20104）
    • RoboSep™ 过滤吸头（产品号 #20125）
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    string(7) "18000_C" NULL
    2. string(7) "20144_C" NULL
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应用领域
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总览

使用EasySep™人Naïve CD4+ T细胞分选试剂盒，可轻松高效地通过免疫磁珠负选从新鲜或冻存的人外周血单个核细胞(PBMC)样本中分离高纯度的初始CD4+ T细胞。EasySep™结合了单克隆抗体的特异性和无柱磁性系统的简便性，迄今已广泛应用于发表的研究中超过20年。

在此EasySep™负选过程中，表达以下标记物的非目标细胞通过抗体复合物和磁珠标记被靶向去除：CD8、CD14、CD16、CD19、CD20、CD25、CD36、CD56、TCRγδ、GlyA、CD61、CD66b、CD123、HLA-DR及CD45RO。通过EasySep™磁极将被磁珠标记的细胞与未被标记的目的初始CD4+ T细胞分离，接着简单地将目的细胞倾倒或吸取至一个新的试管中即可。经磁珠分选获得的初始CD4+ T细胞可直接用于流式细胞术、细胞培养或DNA/RNA提取等下游实验。

如需更快速的细胞分选方案，我们推荐EasySep™人     初始CD4+ T细胞分选试剂盒 II（产品号 #17555），仅需11分钟即可完成细胞分离。

了解更多关于免疫磁珠EasySep™技术的工作原理，或如何通过RoboSep™实现全自动化免疫磁珠细胞分选 。探索更多优化您实验流程的产品，包括培养基、添加剂、抗体等。

磁体兼容性
• EasySep™ Magnet (Catalog #18000) • “The Big Easy” EasySep™ Magnet (Catalog #18001) • Easy 50 EasySep™ Magnet (Catalog #18002) • EasyEights™ EasySep™ Magnet (Catalog #18103) • RoboSep™-S (Catalog #21000)
 
亚型
细胞分选试剂盒
 
细胞类型
T 细胞，T 细胞，CD4+
 
种属
人
 
样本来源
PBMC
 
筛选方法
Negative
 
应用
细胞分选
 
品牌
EasySep，RoboSep
 
研究领域
免疫
 

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Data Figures

Typical EasySep™ Human Naïve CD4+ T Cell Isolation Profile

Figure 1. Typical EasySep™ Human Naïve CD4+ T Cell Isolation Profile

Starting with a single-cell suspension of PBMCs, the naïve CD4+ T cell (CD3+CD4+CD45RA+CD45RO-) content of the isolated fraction typically ranges from 91.3% - 96.9%. In the example above, the purities of the start and isolated fraction are 11.1% and 93.2%, respectively.

Protocols and Documentation

Find supporting information and directions for use in the Product Information Sheet or explore additional protocols below.

Document Type
Product Name
Catalog #
Lot #
Language
Document Type
Product Information Sheet
Product Name
RoboSep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555RF
Lot #
All
Language
English
Document Type
Product Information Sheet
Product Name
EasySep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
RoboSep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555RF
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
RoboSep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555RF
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
RoboSep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555RF
Lot #
All
Language
English
Document Type
Safety Data Sheet 4
Product Name
RoboSep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555RF
Lot #
All
Language
English
Document Type
Safety Data Sheet 1
Product Name
EasySep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Product Name
EasySep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555
Lot #
All
Language
English
Document Type
Safety Data Sheet 3
Product Name
EasySep™ Human Naïve CD4+ T Cell Isolation Kit
Catalog #
19555
Lot #
All
Language
English

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

Resources and Publications

Educational Materials (10)

Tools for Your Immunology Research
Brochure
Tools for Your Immunology Research
EasySep™ Cell Separation Technology
Brochure
EasySep™ Cell Separation Technology
Human T Cell Research Product Workflow
Brochure
Human T Cell Research Product Workflow
Frequencies of Human Cell Types in Blood-Related Sources
Wallchart
Frequencies of Human Cell Types in Blood-Related Sources
Antigen Processing and Presentation
Wallchart
Antigen Processing and Presentation
Human Immune Cytokines
Wallchart
Human Immune Cytokines
How to Isolate PBMCs from Whole Blood Using Density Gradient Centrifugation (Ficoll™ or Lymphoprep™)
1:37
Video
How to Isolate PBMCs from Whole Blood Using Density Gradient Centrifugation (Ficoll™ or Lymphoprep...
How EasySep™ Magnetic Cell Separation Technology Works: Fast and Easy Cell Isolation
1:57
Video
How EasySep™ Magnetic Cell Separation Technology Works: Fast and Easy Cell Isolation
Isolate Cells with a Simple Pour-Off: EasySep™ Cell Separation Technology
1:13
Video
Isolate Cells with a Simple Pour-Off: EasySep™ Cell Separation Technology
Simultaneous Cell Isolation from Multiple Samples Using the EasyEights™ EasySep™ Magnet
0:57
Video
Simultaneous Cell Isolation from Multiple Samples Using the EasyEights™ EasySep™ Magnet

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.

Publications (11)

Exosomal B7-H4 from irradiated glioblastoma cells contributes to increase FoxP3 expression of differentiating Th1 cells and promotes tumor growth. Y. Tian et al. Redox biology 2022 oct

Abstract

BACKGROUND Glioblastoma (GBM) is the most common and aggressive form of primary brain tumor. Although numerous postoperative therapeutic strategies have already been developed, including radiotherapy, tumors inevitably recur after several years of treatment. The coinhibitory molecule B7-H4 negatively regulates T cell immune responses and promotes immune escape. Exosomes mediate intercellular communication and initiate immune evasion in the tumor microenvironment (TME). OBJECTIVE This study aimed to determine whether B7-H4 is upregulated by radiation and loaded into exosomes, thus contributing to immunosuppression and enhancing tumor growth. METHODS Iodixanol density-gradient centrifugation and flow cytometry were used to verify exosomal B7-H4. Na{\{i}}ve T cells were differentiated into Th1 cells with or without exosomes. T cell-secreted cytokines and markers of T cell subsets were measured. Mechanistically the roles of B7-H4 and ALIX in GBM were analyzed using databases and tissue samples. Co-immunoprecipitation and pull-down assays were used to tested the direct interactions between ATM and ALIX or STAT3. In vitro ATM kinase assays western blotting and site-directed mutation were used to assess ATM-mediated STAT3 phosphorylation. Finally the contribution of exosomal B7-H4 to immunosuppression and tumor growth was investigated in vivo. RESULTS Exosomes from irradiated GBM cells decreased the anti-tumor immune response of T cell in vitro and in vivo via delivered B7-H4. Mechanistically irradiation promoted exosome biogenesis by increasing the ATM-ALIX interaction. Furthermore the ATM-phosphorylated STAT3 was found to directly binds to the B7-H4 promoter to increase its expression. Finally the radiation-induced increase in exosomal B7-H4 induced FoxP3 expression during Th1 cell differentiation via the activated STAT1 pathway. In vivo exosomal B7-H4 decreased the radiation sensitivity of GBM cells and reduced the survival of GBM mice model. CONCLUSION This study showed that radiation-enhanced exosomal B7-H4 promoted immunosuppression and tumor growth hence defining a direct link between irradiation and anti-tumor immune responses. Our results suggest that co-administration of radiotherapy with anti-B7-H4 therapy could improve local tumor control and identify exosomal B7-H4 as a potential tumor biomarker."
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NOTCH1 signaling during CD4+ T-cell activation alters transcription factor networks and enhances antigen responsiveness. A. B. Wilkens et al. Blood 2022 nov

Abstract

Adoptive transfer of T cells expressing chimeric antigen receptors (CAR-T) effectively treats refractory hematologic malignancies in a subset of patients but can be limited by poor T-cell expansion and persistence in vivo. Less differentiated T-cell states correlate with the capacity of CAR-T to proliferate and mediate antitumor responses, and interventions that limit tumor-specific T-cell differentiation during ex vivo manufacturing enhance efficacy. NOTCH signaling is involved in fate decisions across diverse cell lineages and in memory CD8+ T cells was reported to upregulate the transcription factor FOXM1, attenuate differentiation, and enhance proliferation and antitumor efficacy in vivo. Here, we used a cell-free culture system to provide an agonistic NOTCH1 signal during na{\{i}}ve CD4+ T-cell activation and CAR-T production and studied the effects on differentiation transcription factor expression cytokine production and responses to tumor. NOTCH1 agonism efficiently induced a stem cell memory phenotype in CAR-T derived from na{\"{i}}ve but not memory CD4+ T cells and upregulated expression of AhR and c-MAF driving heightened production of interleukin-22 interleukin-10 and granzyme B. NOTCH1-agonized CD4+ CAR-T demonstrated enhanced antigen responsiveness and proliferated to strikingly higher frequencies in mice bearing human lymphoma xenografts. NOTCH1-agonized CD4+ CAR-T also provided superior help to cotransferred CD8+ CAR-T driving improved expansion and curative antitumor responses in vivo at low CAR-T doses. Our data expand the mechanisms by which NOTCH can shape CD4+ T-cell behavior and demonstrate that activating NOTCH1 signaling during genetic modification ex vivo is a potential strategy for enhancing the function of T cells engineered with tumor-targeting receptors."
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The N6-methyladenosine writer WTAP contributes to the induction of immune tolerance post kidney transplantation by targeting regulatory T cells. Z. Wang et al. Laboratory investigation; a journal of technical methods and pathology 2022 nov

Abstract

N6-methyladenosine (m6A) modification is involved in diverse immunoregulation, while the relationship between m6A modification and immune tolerance post kidney transplantation remains unclear. Expression of Wilms tumor 1-associating protein (WTAP), an m6A writer, was firstly detected in tolerant kidney transplant recipients (TOL). Then the role of WTAP on regulatory T (Treg) cell differentiation and function in CD4+ T cells from kidney transplant recipients with immune rejection (IR) was investigated. The potential target of WTAP and effect of WTAP on immune tolerance in vivo were subsequently verified. WTAP was upregulated in CD4+ T cells of TOL and positively correlated with Treg cell proportion. In vitro, WTAP overexpression promoted Treg cell differentiation and enhanced Treg cell-mediated suppression toward na?ve T cells. Forkhead box other 1 (Foxo1) was identified as a target of WTAP. WTAP enhanced m6A modification of Foxo1 mRNA in coding sequence (CDS) region, leading to up-regulation of Foxo1. Overexpression of m6A demethylase removed the effect of WTAP overexpression, while Foxo1 overexpression reversed these effects. WTAP overexpression alleviated allograft rejection in model mice, as evidenced by reduced inflammatory response and increased Treg population. Our study suggests that WTAP plays a positive role in induction of immune tolerance post kidney transplant by promoting Treg cell differentiation and function. leading to up-regulation of Foxo1. Overexpression of m6A demethylase removed the effect of WTAP overexpression while Foxo1 overexpression reversed these effects. WTAP overexpression alleviated allograft rejection in model mice as evidenced by reduced inflammatory response and increased Treg population. Our study suggests that WTAP plays a positive role in induction of immune tolerance post kidney transplant by promoting Treg cell differentiation and function."
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更多信息

更多信息
Species Human
Magnet Compatibility • EasySep™ Magnet (Catalog #18000) • “The Big Easy” EasySep™ Magnet (Catalog #18001) • Easy 50 EasySep™ Magnet (Catalog #18002) • EasyEights™ EasySep™ Magnet (Catalog #18103) • RoboSep™-S (Catalog #21000)
Sample Source PBMC
Selection Method Negative
标记抗体
  1. 抗人CD3抗体，克隆UCHT1
    抗人CD3抗体，克隆UCHT1

    小鼠（BALB/c）单克隆IgG1抗体，抗人、黑猩猩CD3

  2. 抗人CD45抗体，克隆HI30
    抗人CD45抗体，克隆HI30

    小鼠单克隆IgG1抗体，抗人、黑猩猩CD45

  3. 抗人CD45RO抗体，克隆UCHL1
    抗人CD45RO抗体，克隆UCHL1

    小鼠单克隆IgG2a抗体，抗人、黑猩猩、普通狨猴CD45RO

  4. 抗人CD4抗体，克隆OKT4
    抗人CD4抗体，克隆OKT4

    小鼠单克隆IgG2b抗体，抗人、恒河猴、食蟹猴CD4

  5. 抗人CD45RA，克隆HI100
    抗人CD45RA，克隆HI100

    抗人、黑猩猩CD45RA的小鼠单克隆IgG2b抗体

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