EasySep™ Human CD19 Positive Selection Kit II

Immunomagnetic positive selection kit

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Immunomagnetic positive selection kit
From: 667 USD

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Overview

The EasySep™ Human CD19 Positive Selection Kit II is designed to isolate CD19+ cells from fresh or previously frozen peripheral blood mononuclear cells by positive selection. Desired cells are targeted with Tetrameric Antibody Complexes recognizing CD19 and dextran-coated magnetic particles. The cocktail also contains an antibody to human Fc receptor to minimize nonspecific binding. Labeled cells are separated using an EasySep™ magnet without the use of columns. Cells of interest remain in the tube while unwanted cells are poured off. The CD19 antigen is expressed on mature B cells and B cell progenitors but is lost upon maturation to plasma cells. It is also expressed on follicular dendritic cells.

This product replaces the EasySep™ Human CD19 Positive Selection Kit (Catalog #18054) for even faster cell isolations.
Advantages:
• Fast and easy-to-use
• Up to 99% purity
• No columns required
Components:
  • EasySep™ Human CD19 Positive Selection Kit II (Catalog #17854)
    • EasySep™ Human CD19 Positive Selection Cocktail II, 1 mL
    • EasySep™ Dextran RapidSpheres™ 50100, 1 mL
  • RoboSep™ Human CD19 Positive Selection Kit II (Catalog #17854RF)
    • EasySep™ Human CD19 Positive Selection Cocktail II, 1 mL
    • EasySep™ Dextran RapidSpheres™ 50100, 1 mL
    • RoboSep™ Buffer (Catalog #20104)
    • RoboSep™ Filter Tips (Catalog #20125)
Magnet Compatibility:
• EasySep™ Magnet (Catalog #18000)
• “The Big Easy” EasySep™ Magnet (Catalog #18001)
• EasyPlate™ EasySep™ Magnet (Catalog #18102)
• EasyEights™ EasySep™ Magnet (Catalog #18103)
• Easy 50 EasySep™ Magnet (Catalog #18002)
• RoboSep™-S (Catalog #21000)
Subtype:
Cell Isolation Kits
Cell Type:
B Cells
Species:
Human
Sample Source:
PBMC
Selection Method:
Positive
Application:
Cell Isolation
Brand:
EasySep; RoboSep
Area of Interest:
Chimerism; Immunology

Scientific Resources

Educational Materials

(10)

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

Typical EasySep™ Human CD19 Positive Selection Profile

Figure 1. Typical EasySep™ Human CD19 Positive Selection Profile

Starting with a single cell suspension of human PBMCs, the CD19+ cell content of the isolated fraction is typically 98 ± 1% (mean ± SD using the purple EasySep™ Magnet).

Publications

(4)
Nature communications 2018 OCT

Sequencing HIV-neutralizing antibody exons and introns reveals detailed aspects of lineage maturation.

E. L. Johnson et al.

Abstract

The developmental pathways of broadly neutralizing antibodies (bNAbs) against HIV are of great importance for the design of immunogens that can elicit protective responses. Here we show the maturation features of the HIV-neutralizing anti-V1V2 VRC26 lineage by simultaneously sequencing the exon together with the downstream intron of VRC26 members. Using the mutational landscapes of both segments and the selection-free nature of the intron region, we identify multiple events of amino acid mutational convergence in the complementarity-determining region 3 (CDR3) of VRC26 members, and determine potential intermediates with diverse CDR3s to a late stage bNAb from 2 years prior to its isolation. Moreover, we functionally characterize the earliest neutralizing intermediates with critical CDR3 mutations, with some emerging only 14 weeks after initial lineage detection and containing only {\~{}}6{\%} V gene mutations. Our results thus underscore the utility of analyzing exons and introns simultaneously for studying antibody maturation and repertoire selection.
Nature biotechnology 2018 JAN

Multiplexed droplet single-cell RNA-sequencing using natural genetic variation.

Kang HM et al.

Abstract

Droplet single-cell RNA-sequencing (dscRNA-seq) has enabled rapid, massively parallel profiling of transcriptomes. However, assessing differential expression across multiple individuals has been hampered by inefficient sample processing and technical batch effects. Here we describe a computational tool, demuxlet, that harnesses natural genetic variation to determine the sample identity of each droplet containing a single cell (singlet) and detect droplets containing two cells (doublets). These capabilities enable multiplexed dscRNA-seq experiments in which cells from unrelated individuals are pooled and captured at higher throughput than in standard workflows. Using simulated data, we show that 50 single-nucleotide polymorphisms (SNPs) per cell are sufficient to assign 97% of singlets and identify 92% of doublets in pools of up to 64 individuals. Given genotyping data for each of eight pooled samples, demuxlet correctly recovers the sample identity of<99% of singlets and identifies doublets at rates consistent with previous estimates. We apply demuxlet to assess cell-type-specific changes in gene expression in 8 pooled lupus patient samples treated with interferon (IFN)-β and perform eQTL analysis on 23 pooled samples.
Oncoimmunology 2018

Co-stimulatory signaling determines tumor antigen sensitivity and persistence of CAR T cells targeting PSCA+ metastatic prostate cancer.

S. J. Priceman et al.

Abstract

Advancing chimeric antigen receptor (CAR)-engineered adoptive T cells for the treatment of solid cancers is a major focus in the field of immunotherapy, given impressive recent clinical responses in hematological malignancies. Prostate cancer may be amenable to T cell-based immunotherapy since several tumor antigens, including prostate stem-cell antigen (PSCA), are widely over-expressed in metastatic disease. While antigen selectivity of CARs for solid cancers is crucial, it is problematic due to the absence of truly restricted tumor antigen expression and potential safety concerns with on-target off-tumor" activity. Here
Frontiers in immunology 2018

PD-L1+ Regulatory B Cells Are Significantly Decreased in Rheumatoid Arthritis Patients and Increase After Successful Treatment.

E. R. Zacca et al.

Abstract

Background: B cells play an important role in the development and maintenance of rheumatoid arthritis (RA). Although IL-10-producing B cells represent a major subset of regulatory B cells (Bregs) able to suppress autoimmune and inflammatory responses, recent reports showed that B cell-mediated immune suppression may also occur independent of IL-10. For instance, B cells can modulate T cell immune responses through the expression of regulatory molecules such as PD-L1. So far, PD-L1-expressing B cells have not been analyzed in RA patients. Objective: To analyze the frequency of PD-L1-expressing B cells in the peripheral blood of RA patients compared to healthy controls (HC) matched for sex and age, their function on T cell response and their changes in response to therapy. Methods: Fresh peripheral blood B cells from RA patients and HC were characterized by flow cytometry and their functionality assessed in a co-culture system with autologous T cells. Results: The frequencies of CD19+PD-L1+ B cells, CD24hiCD38-PD-L1+ and CD24hiCD38hiPD-L1+ B cells were significantly lower in untreated RA patients than in HC. In a follow-up study, the frequencies of PD-L1+ B cells (CD19+PD-L1+ B cells, CD24hiCD38-PD-L1+ and CD24hiCD38hiPD-L1+ B cells) increased significantly after treatment in good responder patients, although the frequency of total CD24hiCD38hi B cells decreased. CD19+ B cells from untreated RA patients and HC upregulated PD-L1 expression similarly upon stimulation with CpG plus IL-2 and were able to suppress, in vitro, CD8+ T cell proliferation and cytokine production in a PD-L1-dependent manner. Conclusions: Our results show that PD-L1+ B cells exhibiting T cell suppressive capacity are significantly decreased in untreated RA patients but increase in response to successful treatment. PD-L1 expression on B cells from RA patients can be modulated in vitro and PD-L1+ B cells could thus provide new perspectives for future treatment strategies.
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.