EasySep™ Human B Cell Isolation Kit

9-Minute cell isolation kit using immunomagnetic negative selection

New look, same high quality and support! You may notice that your instrument or reagent packaging looks slightly different from images displayed on the website, or from previous orders. We are updating our look but rest assured, the products themselves and how you should use them have not changed. Learn more

EasySep™ Human B Cell Isolation Kit

9-Minute cell isolation kit using immunomagnetic negative selection

From: 909 USD
Catalog #
17954_C
9-Minute cell isolation kit using immunomagnetic negative selection

Product Advantages


  • Fast, easy-to-use and column-free

  • Up to 96% purity with high recovery

  • Isolated cells are untouched

What's Included

  • EasySep™ Human B Cell Isolation Kit (Catalog #17954)
    • EasySep™ Human B Cell Isolation Cocktail, 1 mL
    • EasySep™ Dextran RapidSpheres™, 1 mL
    • EasySep™ Isolation Cocktail Enhancer, 1 mL
  • EasySep™ Human B Cell Isolation Kit (Catalog #100-0971)
    • EasySep™ Human B Cell Isolation Cocktail, 1 x 10 mL (Catalog #300-0510)
    • EasySep™ Isolation Cocktail Enhancer, 1 x 10 mL (Catalog #300-0511)
    • EasySep™ Dextran RapidSpheres™, 1 x 10 mL (Catalog #300-0380)
  • RoboSep™ Human B Cell Isolation Kit (Catalog #17954RF)
    • EasySep™ Human B Cell Isolation Cocktail, 1 mL
    • EasySep™ Dextran RapidSpheres™, 1 mL
    • EasySep™ Isolation Cocktail Enhancer, 1 mL
    • RoboSep™ Buffer (Catalog #20104)
    • RoboSep™ Filter Tips (Catalog #20125)
New look, same high quality and support! You may notice that your instrument or reagent packaging looks slightly different from images displayed on the website, or from previous orders. We are updating our look but rest assured, the products themselves and how you should use them have not changed. Learn more

Overview

The EasySep™ Human B Cell Isolation Kit is designed to isolate B cells from fresh or previously frozen peripheral blood mononuclear cells or washed leukapheresis samples by immunomagnetic negative selection. The EasySep™ procedure involves labeling unwanted cells with antibody complexes and magnetic particles. The magnetically labeled cells are separated from the untouched, desired cells by using an EasySep™ magnet and simply pouring or pipetting the desired cells into a new tube. This product can be used in place of the EasySep™ Human B Cell Enrichment Kit (Catalog #19054) for even faster cell isolations.
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)
• Easy 250 EasySep™ Magnet (Catalog #100-0821)
Subtype
Cell Isolation Kits
Cell Type
B Cells
Species
Human
Sample Source
Leukapheresis, PBMC
Selection Method
Negative
Application
Cell Isolation
Brand
EasySep, RoboSep
Area of Interest
Chimerism, HLA, Immunology

Data Figures

Figure 1. Typical EasySep™ Human B Cell Isolation Profile

Starting with human PBMCs, the B cell (CD3-CD19+) content of the isolated fraction is typically 95.1 ± 1.4% (mean ± SD). In the example above, the final purities of the start and isolated fractions are 4.5% and 94.9%, 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 #
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Catalog #
17954
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All other lots
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English
Catalog #
17954
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Purchase date 2020-04-07 and later
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English
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17954RF
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All other lots
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English
Catalog #
17954RF
Lot #
Purchase date 2020-04-07 and later
Language
English
Catalog #
100-0971
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All
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English
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Safety Data Sheet 1
Catalog #
17954
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All
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English
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Safety Data Sheet 2
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17954
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English
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Safety Data Sheet 3
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17954
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English
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Safety Data Sheet 1
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17954RF
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All
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English
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Safety Data Sheet 2
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17954RF
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All
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English
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Safety Data Sheet 3
Catalog #
17954RF
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All
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English
Document Type
Safety Data Sheet 1
Catalog #
100-0971
Lot #
All
Language
English
Document Type
Safety Data Sheet 2
Catalog #
100-0971
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.

Resources and Publications

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 (6)

Inflammation-Induced Mucosal KYNU Expression Identifies Human Ileal Crohn's Disease. M. Huhn et al. Journal of clinical medicine 2020 may

Abstract

The widely varying therapeutic response of patients with inflammatory bowel disease (IBD) continues to raise questions regarding the unclarified heterogeneity of pathological mechanisms promoting disease progression. While biomarkers for the differentiation of Crohn's disease (CD) versus ulcerative colitis (UC) have been suggested, specific markers for a CD subclassification in ileal CD versus colonic CD are still rare. Since an altered signature of the tryptophan metabolism is associated with chronic inflammatory disease, we sought to characterize potential biomarkers by focusing on the downstream enzymes and metabolites of kynurenine metabolism. Using immunohistochemical stainings, we analyzed and compared the mucosal tryptophan immune metabolism in bioptic samples from patients with active inflammation due to UC or CD versus healthy controls. Localization-specific quantification of immune cell infiltration, tryptophan-metabolizing enzyme expression and mucosal tryptophan downstream metabolite levels was performed. We found generally increased immune cell infiltrates in the tissue of all patients with IBD. However, in patients with CD, significant differences were found between regulatory T cell and neutrophil granulocyte infiltration in the ileum compared with the colon. Furthermore, we observed decreased kynurenine levels as well as strong kynureninase (KYNU) expression specifically in patients with ileal CD. Correspondingly, significantly elevated levels of the kynurenine metabolite 3-hydroxyanthranilic acid were detected in the ileal CD samples. Highlighting the heterogeneity of the different phenotypes of CD, we identified KYNU as a potential mucosal biomarker allowing the localization-specific differentiation of ileal CD versus colonic CD.
Transitional B cells in quiescent SLE: An early checkpoint imprinted by IFN. Y. Dieudonn\'e et al. Journal of autoimmunity 2019 may

Abstract

Systemic lupus (SLE) is characterized by a break of B cell tolerance that plays a central role in disease pathophysiology. An early checkpoint defect occurs at the transitional stage leading to the survival of autoreactive B cells and consequently the production of pathogenic autoantibodies. The main purpose of our work was to determine whether transitional B cells, as the most immature na{\{i}}ve B cell subset upstream of pathogenic B cells display specific features compared to healthy non SLE subjects. Through extensive analysis of transitional B cells from untreated or low treated mostly Caucasian SLE patients we demonstrated that transitional (T1 and T2) B cell frequencies were increased in SLE and positively correlated with disease activity. SLE transitional B cells displayed defects in two closely inter-related molecules (i.e. TLR9 defective responses and CD19 downregulation). RNA sequencing of sorted transitional B cells from untreated patients revealed a predominant overexpression of interferon stimulated genes (ISGs) even out of flares. In addition early transitional B cells from the bone marrow displayed the highest interferon score reflecting a B cell interferon burden of central origin. Hence the IFN signature in transitional B cells is not confined to African American SLE patients and exists in quiescent disease since the medullary stage. These results suggest that in SLE these 3 factors (i.e. IFN imprintment CD19 downregulation and TLR9 responses impairment) could take part at the early transitional B cell stage in B cell tolerance by-pass ultimately leading in periphery to the expansion of autoantibodies-secreting cells."
Abatacept modulates CD80 and CD86 expression and memory formation in human B-cells. R. Lorenzetti et al. Journal of autoimmunity 2019 jul

Abstract

BACKGROUND Cytotoxic T lymphocyte antigen-4 (CTLA-4) limits T-cell activation and is expressed on T-regulatory cells. Human CTLA-4 deficiency results in severe immune dysregulation. Abatacept (CTLA-4 Ig) is approved for the treatment of rheumatoid arthritis (RA) and its mechanism of action is attributed to effects on T-cells. It is known that CTLA-4 modulates the expression of its ligands CD80 and CD86 on antigen presenting cells (APC) by transendocytosis. As B-cells express CD80/CD86 and function as APC, we hypothesize that B-cells are a direct target of abatacept. OBJECTIVES To investigate direct effects of abatacept on human B-lymphocytes in vitro and in RA patients. METHODS The effect of abatacept on healthy donor B-cells' phenotype, activation and CD80/CD86 expression was studied in vitro. Nine abatacept-treated RA patients were studied. Seven of these were followed up to 24 months, and two up to 12 months only and treatment response, immunoglobulins, ACPA, RF concentrations, B-cell phenotype and ACPA-specific switched memory B-cell frequency were assessed. RESULTS B-cell development was unaffected by abatacept. Abatacept treatment resulted in a dose-dependent decrease of CD80/CD86 expression on B-cells in vitro, which was due to dynamin-dependent internalization. RA patients treated with abatacept showed a progressive decrease in plasmablasts and serum IgG. While ACPA-titers only moderately declined, the frequency of ACPA-specific switched memory B-cells significantly decreased. CONCLUSIONS Abatacept directly targets B-cells by reducing CD80/CD86 expression. Impairment of antigen presentation and T-cell activation may result in altered B-cell selection, providing a new therapeutic mechanism and a base for abatacept use in B-cell mediated autoimmunity.

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