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Human NK Cell Isolation Kit (Column-Based)

Human NK Cell Isolation Kit (Column-Based)

Catalog Number: S0K1009 Reactivity: Human Conjugation: Brand: Starter
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$1,000 USD
$1,000 USD
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Product Details

Product Specification


Format

1 mL Human NK Cell Biotin-Antibody Cocktail

2mL Streptavidin Beads
Capacity

Suitable for a total cell count of 1×10⁹, and can perform up to 100 sorting runs.
Principle of Separation

In this negative selection process, non-NK cells are labeled with biotin-conjugated antibodies, and the cells are then incubated with streptavidin‑nanomagnetic beads. As a result, the non‑target cells are marked by the antibody‑bead complex, while the target NK cells remain unlabeled. Subsequently, the cell suspension is transferred to a separation column; under the influence of a magnetic field, the magnetically labeled non‑target cells adhere to the column, whereas the unlabeled NK cells elute into a collection tube under gravity. The resulting supernatant constitutes the NK‑cell‑enriched fraction after removal of the non‑target cells.
Reactivity Human
Magnetic Bead Size 20 nm
Separation Method Column-Based
Selection Strategy Negative Selection
Stability & Storage

Store protected from light at 2–8°C; do not freeze.

Background

Human NK cells (Natural Killer Cells) are key effector cells of the innate immune system, accounting for 5%–20% of peripheral blood lymphocytes. They can directly kill virus-infected and tumor cells without prior antigen sensitization and secrete large amounts of cytokines to regulate immune responses, making them a critical focus in cancer immunology, anti-infectious immunity, and immunotherapy research. NK cells isolated using an NK cell negative selection kit are suitable for research scenarios where direct surface molecule labeling might interfere with downstream applications; the purified NK cells can be immediately used for flow cytometry, cell culture, or DNA/RNA extraction, among other downstream assays.

Protocol

Note: This kit does not include separation buffer; please prepare it yourself. The corresponding Stater catalog number isS0D3018

TakingL Separation Columnas an example

Steps

Operating Instructions

Dosage and Time

Cell Processing and Labeling

1

Isolate PBMCs from peripheral blood using density gradient centrifugation with lymphocyte separation medium, or directly thaw cryopreserved PBMCs, and perform cell counting.

Note: It is recommended to count cells after staining with AO/PI; PBMCs can be centrifuged at 400 g for 7 minutes.

Prepare the sample

2

Resuspend 1×10^7 cells in 100 μL of MagSep Separation Buffer.Note:Adjust reagent volumes proportionally based on sample volume; set aside a pre-sorting sample to determine the proportion of human NK cells.

1×10^77

cells/100 μL7cells/100 μL

3

Add 10 μL of Human NK cell Biotin Antibody Cocktail to the sample and gently mix.

Note:In this step, gently pipette the mixture 2–3 times to mix evenly;

10 μL/100 μL

4

Incubate the antibodies in the cocktail with the cells at room temperature for 10 min;

Incubate at room temperature for 10 min

5

Mix wellStreptavidin Beads, add 20 μLStreptavidin Beadsinto the sample;

20 μL/100 μL

6

Gently mix the magnetic beads and cells, and incubate at room temperature for 5 minutes;

Incubate at room temperature for 5 minutes

Cell sorting

7

Place it in the column-type magnetic separator,L Separation Column, add 3 mL of MagSep Separation Buffer to rinse the separation column;

Rinse the separation column

8

Add the cells to the separation column and collect the target cells using a collection tube;

Note:The cell volume should be at least 500 μL; if insufficient, add MagSep Separation Buffer to make up the volume. Cells that drip down under gravity are the target cells.

Collect the target cells

9

Add 3 mL of MagSep Separation Buffer and collect the target cells.

Note:This step elutes the unlabeled target cells from the separation column, increasing the yield.

Collect the target cells

Notes: Pay attention to the maximum capacity of the separation column.The S Separation Column can hold a maximum of1×107;The L Separation Column can hold a maximum of1×108;


Picture

Validation Data

Human NK cells were enriched from peripheral blood mononuclear cells (PBMCs). After fluorescent staining with CD56-Alexa Fluor® 647 (SDT Cat: S0B8402) and CD3-Alexa Fluor® 488 (SDT Cat: S0B1636), live cells (Live+) were gated for analysis. Flow cytometric results showed that the purity of the sorted human NK cells was 91.6%.