The EasySep™ Direct Human Pan-Granulocyte Isolation Kit isolates functional, highly purified neutrophils, eosinophils, and basophils directly from human whole blood by immunomagnetic negative selection. No lysis, density gradient centrifugation, or other processing steps are required and isolated cells are immediately available for flow cytometry, functional assays, culture, and other downstream applications.
概述
EasySep™ Direct 人Pan-Granulocyt分选试剂盒通过免疫磁珠负选，直接从人全血中分离具有功能的、高纯度的中性粒细胞、嗜酸性粒细胞和嗜碱性粒细胞。无需裂解、密度梯度离心或其他处理步骤，分选后的细胞可直接用于流式细胞术、功能分析、细胞培养及其他下游应用。
MAGNET COMPATIBILITY • EasySep™ Magnet (Catalog #18000), or • “The Big Easy” EasySep™ Magnet (Catalog #18001), or • Easy 50 EasySep™ Magnet (Catalog #18002), or • EasyEights™ EasySep™ Magnet (Catalog #18103)
SUBTYPE Cell Isolation Kits
CELL TYPE Granulocytes and Subsets
SPECIES Human
SAMPLE SOURCE Whole Blood
SELECTION METHOD Negative
APPLICATION Cell Isolation
BRAND EasySep
AREA OF INTEREST Immunology
实验数据
Figure 1. Typical EasySep™ Direct Human Pan-Granulocyte Isolation Profile
Starting with human whole blood from normal healthy donors, the typical pan-granulocyte (neutrophil [CD66b+CD16+], eosinophil [CD66b+CD16-] and basophil [CD66b-CD123+]) content of the non-lysed final isolated fraction is 98.4 ± 1.5% (gated on CD45). In the above example, the pan-granulocyte content of the lysed whole blood start sample and the non-lysed final isolated fraction is 61.8% and 99.6% (gated on CD45), respectively.
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.
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.
Nuclear positioning facilitates amoeboid migration along the path of least resistance. J. Renkawitz et al. Nature 2019 apr
Abstract
During metazoan development, immune surveillance and cancer dissemination, cells migrate in complex three-dimensional microenvironments1-3. These spaces are crowded by cells and extracellular matrix, generating mazes with differently sized gaps that are typically smaller than the diameter of the migrating cell4,5. Most mesenchymal and epithelial cells and some-but not all-cancer cells actively generate their migratory path using pericellular tissue proteolysis6. By contrast, amoeboid cells such as leukocytes use non-destructive strategies of locomotion7, raising the question how these extremely fast cells navigate through dense tissues. Here we reveal that leukocytes sample their immediate vicinity for large pore sizes, and are thereby able to choose the path of least resistance. This allows them to circumnavigate local obstacles while effectively following global directional cues such as chemotactic gradients. Pore-size discrimination is facilitated by frontward positioning of the nucleus, which enables the cells to use their bulkiest compartment as a mechanical gauge. Once the nucleus and the closely associated microtubule organizing centre pass the largest pore, cytoplasmic protrusions still lingering in smaller pores are retracted. These retractions are coordinated by dynamic microtubules; when microtubules are disrupted, migrating cells lose coherence and frequently fragment into migratory cytoplasmic pieces. As nuclear positioning in front of the microtubule organizing centre is a typical feature of amoeboid migration, our findings link the fundamental organization of cellular polarity to the strategy of locomotion.
High-throughput single-cell rheology in complex samples by dynamic real-time deformability cytometry. B. Fregin et al. Nature communications 2019
Abstract
In life sciences, the material properties of suspended cells have attained significance close to that of fluorescent markers but with the advantage of label-free and unbiased sample characterization. Until recently, cell rheological measurements were either limited by acquisition throughput, excessive post processing, or low-throughput real-time analysis. Real-time deformability cytometry expanded the application of mechanical cell assays to fast on-the-fly phenotyping of large sample sizes, but has been restricted to single material parameters as the Young's modulus. Here, we introduce dynamic real-time deformability cytometry for comprehensive cell rheological measurements at up to 100 cells per second. Utilizing Fourier decomposition, our microfluidic method is able to disentangle cell response to complex hydrodynamic stress distributions and to determine viscoelastic parameters independent of cell shape. We demonstrate the application of our technology for peripheral blood cells in whole blood samples including the discrimination of B- and CD4+ T-lymphocytes by cell rheological properties.
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