总览

使用EasySep™ Direct 人中性粒细胞分选试剂盒，可轻松高效地从人全血样本中通过免疫磁珠负选获得高纯度人中性粒细胞。EasySep™技术结合单克隆抗体的特异性和免磁柱系统的简便性，已在发表的研究中广泛应用超过20年。

在该EasySep™负选流程中，非目的细胞会被抗体复合物和EasySep™ Direct RapidSpheres™ 磁珠标记。以下非目的细胞会被特异性去除：嗜碱性粒细胞、嗜酸性粒细胞、T细胞、B细胞、单核细胞、NK细胞和红系细胞。通过EasySep™磁极将被磁珠标记的细胞与未被标记的目的细胞分离，目的细胞可通过倾倒或者移液吸取到新试管中，分选后的细胞可直接用于流式细胞术、细胞培养或DNA/RNA提取等下游应用。

该产品可替代EasySep™人中性粒细胞分选试剂盒 (产品号 #19257) 以实现更快地细胞分选。

了解更多EasySep™免疫磁珠技术的工作原理，或者如何通过RoboSep™实现全自动化免疫磁珠细胞分选。探索更多为您的实验流程优化的产品，包括细胞鉴定、冷冻保存等相关试剂。

在该EasySep™负选流程中，非目的细胞会被抗体复合物和EasySep™ Direct RapidSpheres™ 磁珠标记。以下非目的细胞会被特异性去除：嗜碱性粒细胞、嗜酸性粒细胞、T细胞、B细胞、单核细胞、NK细胞和红系细胞。通过EasySep™磁极将被磁珠标记的细胞与未被标记的目的细胞分离，目的细胞可通过倾倒或者移液吸取到新试管中，分选后的细胞可直接用于流式细胞术、细胞培养或DNA/RNA提取等下游应用。

该产品可替代EasySep™人中性粒细胞分选试剂盒 (产品号 #19257) 以实现更快地细胞分选。

了解更多EasySep™免疫磁珠技术的工作原理，或者如何通过RoboSep™实现全自动化免疫磁珠细胞分选。探索更多为您的实验流程优化的产品，包括细胞鉴定、冷冻保存等相关试剂。

在该EasySep™负选流程中，非目的细胞会被抗体复合物和EasySep™ Direct RapidSpheres™ 磁珠标记。以下非目的细胞会被特异性去除：嗜碱性粒细胞、嗜酸性粒细胞、T细胞、B细胞、单核细胞、NK细胞和红系细胞。通过EasySep™磁极将被磁珠标记的细胞与未被标记的目的细胞分离，目的细胞可通过倾倒或者移液吸取到新试管中，分选后的细胞可直接用于流式细胞术、细胞培养或DNA/RNA提取等下游应用。

该产品可替代EasySep™人中性粒细胞分选试剂盒 (产品号 #19257) 以实现更快地细胞分选。

了解更多EasySep™免疫磁珠技术的工作原理，或者如何通过RoboSep™实现全自动化免疫磁珠细胞分选。探索更多为您的实验流程优化的产品，包括细胞鉴定、冷冻保存等相关试剂。

在该EasySep™负选流程中，非目的细胞会被抗体复合物和EasySep™ Direct RapidSpheres™ 磁珠标记。以下非目的细胞会被特异性去除：嗜碱性粒细胞、嗜酸性粒细胞、T细胞、B细胞、单核细胞、NK细胞和红系细胞。通过EasySep™磁极将被磁珠标记的细胞与未被标记的目的细胞分离，目的细胞可通过倾倒或者移液吸取到新试管中，分选后的细胞可直接用于流式细胞术、细胞培养或DNA/RNA提取等下游应用。

该产品可替代EasySep™人中性粒细胞分选试剂盒 (产品号 #19257) 以实现更快地细胞分选。

了解更多EasySep™免疫磁珠技术的工作原理，或者如何通过RoboSep™实现全自动化免疫磁珠细胞分选。探索更多为您的实验流程优化的产品，包括细胞鉴定、冷冻保存等相关试剂。

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
粒细胞及其亚群

种属
人

样本来源
Whole Blood

筛选方法
Negative

应用
细胞分选

品牌
EasySep，RoboSep

研究领域
免疫，感染性疾病(传染病)

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实验数据

Figure 1. Typical EasySep™ Direct Human Neutrophil Isolation Profile

Starting with human whole blood from normal healthy donors, the typical neutrophil (CD66b+CD16+) content of the non-lysed final isolated fraction Is 97.3 ± 1.4% (gated on CD45) or 94.0 ± 3.7% (not gated on CD45). In the example above, the neutrophil (CD66b+CD16+) content of the lysed whole blood start sample and the non-lysed final isolated fraction is 50.6% and 97.2% (gated on CD45), respectively, or 43.6% and 95.9% (not gated on CD45), respectively. The starting frequency of neutrophils in the non-lysed whole blood start sample above is 0.04% (data not shown).

Figure 2. Typical EasySep™ Direct Protocol

Use of Highly Enriched Lymphocytes Isolated with EasySep™ Direct Improves DSA Detection Compared to Whole Leukocyte Cell Preparations

Figure 3. Use of Highly Enriched Lymphocytes Isolated with EasySep™ Direct Improves DSA Detection Compared to Whole Leukocyte Cell Preparations

Lymphocytes (Ly), neutrophils (Nu), and monocytes (Mo) were isolated from volunteer donors (n = 5) using EasySep™ Direct Catalog #19655, #19666, and #19669, respectively. Whole leukocyte (WL) preparations were obtained by adding Ly, Nu, and Mo cells in equal propotions. WL (low lymphocyte purity) and Ly (high ymphocyte purity) preparations were treated with pronase and then used to perform the flow cytometry cross-match (FCXM) assay against negative control sera or several dilutions of positive control sera. The median channel fluorescence shifts (MCFS) were generated by using the negative control sera samples as a baseline. The MCF shifts between WL and Ly were then compared. Each column with error bars represents the mean ± SEM (n = 5 donors). Data kindly provided by Dr. Robert Liwski.

产品说明书及文档

请在《产品说明书》中查找相关支持信息和使用说明，或浏览下方更多实验方案。

Document Type

Product Name

Catalog #

Lot #

Language

Document Type

Product Information Sheet

Product Name

RoboSep™ Human Neutrophil Isolation Kit

Catalog #

100-0404

Lot #

All

Language

English

Document Type

Product Information Sheet

Product Name

EasySep™ Direct Human Neutrophil Isolation Kit

Catalog #

19666

Lot #

All

Language

English

Document Type

Safety Data Sheet 1

Product Name

RoboSep™ Human Neutrophil Isolation Kit

Catalog #

100-0404

Lot #

All

Language

English

Document Type

Safety Data Sheet 2

Product Name

RoboSep™ Human Neutrophil Isolation Kit

Catalog #

100-0404

Lot #

All

Language

English

Document Type

Safety Data Sheet 3

Product Name

RoboSep™ Human Neutrophil Isolation Kit

Catalog #

100-0404

Lot #

All

Language

English

Document Type

Safety Data Sheet 1

Product Name

EasySep™ Direct Human Neutrophil Isolation Kit

Catalog #

19666

Lot #

All

Language

English

Document Type

Safety Data Sheet 2

Product Name

EasySep™ Direct Human Neutrophil Isolation Kit

Catalog #

19666

Lot #

All

Language

English

应用领域

本产品专为以下研究领域设计，适用于工作流程中的高亮阶段。探索这些工作流程，了解更多我们为各研究领域提供的其他配套产品。

Research Area

Workflow Stages

Workflow Stages for Granulocyte Research

Cell Sourcing

Cell Isolation

Cell Characterization

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.

NETosis and thrombosis in vaccine-induced immune thrombotic thrombocytopenia. H. H. L. Leung et al. Nature communications 2022 sep

Abstract

Vaccine-induced immune thrombotic thrombocytopenia (VITT) is a rare yet serious adverse effect of the adenoviral vector vaccines ChAdOx1 nCoV-19 (AstraZeneca) and Ad26.COV2.S (Janssen) against COVID-19. The mechanisms involved in clot formation and thrombocytopenia in VITT are yet to be fully determined. Here we show neutrophils undergoing NETosis and confirm expression markers of NETs in VITT patients. VITT antibodies directly stimulate neutrophils to release NETs and induce thrombus formation containing abundant platelets, neutrophils, fibrin, extracellular DNA and citrullinated histone H3 in a flow microfluidics system and in vivo. Inhibition of NETosis prevents VITT-induced thrombosis in mice but not thrombocytopenia. In contrast, in vivo blockage of Fc$\gamma$RIIa abrogates both thrombosis and thrombocytopenia suggesting these are distinct processes. Our findings indicate that anti-PF4 antibodies activate blood cells via Fc$\gamma$RIIa and are responsible for thrombosis and thrombocytopenia in VITT. Future development of NETosis and Fc$\gamma$RIIa inhibitors are needed to treat VITT and similar immune thrombotic thrombocytopenia conditions more effectively, leading to better patient outcomes.

View publication

Neutrophil degranulation and severely impaired extracellular trap formation at the basis of susceptibility to infections of hemodialysis patients. S. Talal et al. BMC medicine 2022 oct

Abstract

BACKGROUND Chronic kidney disease patients are at increased risk of mortality with cardiovascular diseases and infections as the two leading causes of death for end-stage kidney disease treated with hemodialysis (HD). Mortality from bacterial infections in HD patients is estimated to be 100-1000 times higher than in the healthy population. METHODS We comprehensively characterized highly pure circulating neutrophils from HD and healthy donors. RESULTS Protein levels and transcriptome of HD patients' neutrophils indicated massive neutrophil degranulation with a dramatic reduction in reactive oxygen species (ROS) production during an oxidative burst and defective oxidative cellular signaling. Moreover, HD neutrophils exhibit severely impaired ability to generate extracellular NET formation (NETosis) in NADPH oxidase-dependent or independent pathways, reflecting their loss of capacity to kill extracellular bacteria. Ectopic hydrogen peroxidase (H2O2) or recombinant human SOD-1 (rSOD-1) partly restores and improves the extent of HD dysfunctional neutrophil NET formation. CONCLUSIONS Our report is one of the first singular examples of severe and chronic impairment of NET formation leading to substantial clinical susceptibility to bacteremia that most likely results from the metabolic and environmental milieu typical to HD patients and not by common human genetic deficiencies. In this manner, aberrant gene expression and differential exocytosis of distinct granule populations could reflect the chronic defect in neutrophil functionality and their diminished ability to induce NETosis. Therefore, our findings suggest that targeting NETosis in HD patients may reduce infections, minimize their severity, and decrease the mortality rate from infections in this patient population.

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Peripheral blood mononuclear cell phenotype and function are maintained after overnight shipping of whole blood. R. K. Johnson et al. Scientific reports 2022 nov

Abstract

Same day processing of biospecimens such as blood is not always feasible, which presents a challenge for research programs seeking to study a broad population or to characterize patients with rare diseases. Recruiting sites may not be equipped to process blood samples and variability in timing and technique employed to isolate peripheral blood mononuclear cells (PBMCs) at local sites may compromise reproducibility across patients. One solution is to send whole blood collected by routine phlebotomy via overnight courier to the testing site under ambient conditions. Determining the impact of shipping on subsequent leukocyte responses is a necessary prerequisite to any experimental analysis derived from transported samples. To this end, whole blood was collected from healthy control subjects and processed fresh or at 6, 24 and 48 h after collection and handling under modeled shipping conditions. At endpoint, whole blood was assessed via a complete blood count with differential and immunophenotyped using a standardized panel of antibodies [HLADR, CD66b, CD3, CD14, CD16]. PBMCs and neutrophils were isolated from whole blood and subjected to ex vivo stimulation with lipopolysaccharide and heat-killed Staphylococcus aureus. Stimulated release of cytokines and chemokines was assessed by cytometric bead array. RNA was also isolated from PBMCs to analyze transcriptional changes induced by shipping. The complete blood count with differential revealed that most parameters were maintained in shipped blood held for 24 h at ambient temperature. Immunophenotyping indicated preservation of cellular profiles at 24 h, although with broadening of some populations and a decrease in CD16 intensity on classical monocytes. At the transcriptional level, RNAseq analysis identified upregulation of a transcription factor module associated with inflammation in unstimulated PBMCs derived from whole blood shipped overnight. However, these changes were limited in both scale and number of impacted genes. Ex vivo stimulation of PBMCs further revealed preservation of functional responses in cells isolated from shipped blood held for 24 h at ambient temperature. However, neutrophil responses were largely abrogated by this time. By 48 h neither cell population responded within normal parameters. These findings indicate that robust immunophenotyping and PBMC stimulated response profiles are maintained in whole blood shipped overnight and processed within 24 h of collection, yielding results that are representative of those obtained from the sample immediately following venipuncture. This methodology is feasible for many patient recruitment sites to implement and allows for sophisticated immunological analysis of patient populations derived from large geographic areas. With regard to rare disease research, this meets a universal need to enroll patients in sufficient numbers for immunoprofiling and discovery of underlying pathogenic mechanisms.

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EasySep™ Direct人中性粒细胞分选试剂盒

产品号 #（选择产品）

产品号 #19666_C

产品优势

产品组分包括

What Our Scientist Says

总览

实验数据

产品说明书及文档

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Scientists Helping Scientists™

种属	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)
样本来源	Whole Blood
Selection Method	Negative

EasySep™ Direct人中性粒细胞分选试剂盒

产品号 #（选择产品）

产品号 #19666_C

产品优势

产品组分包括

What Our Scientist Says

总览

实验数据

产品说明书及文档

应用领域

相关材料与文献

技术资料 (10)

常见问题

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

How does the separation work?

Which columns do I use?

How can I analyze the purity of my enriched sample?

Can EasySep™ separations be automated?

Can EasySep™ be used to isolate rare cells?

Are the EasySep™ magnetic particles FACS-compatible?

Can the EasySep™ magnetic particles be removed after enrichment?

Can I alter the separation time in the magnet?

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

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

文献 (24)

Abstract

Abstract

Abstract

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