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Human CD4 T Cell Isolation Kit (Column-Based)

Human CD4 T Cell Isolation Kit (Column-Based)

Catalog Number: S0K1003 Reactivity: Human Conjugation: Brand: Starter
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Product Details

Product Specification


Format

1 mL Human CD4 T Cell Biotin-Antibody Cocktail

1.5 mL 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-CD4 T cells are labeled with biotin-conjugated antibodies, and the cells are then incubated with streptavidin‑nanomagnetic beads. As a result, the non‑target cells become bound to the magnetic beads via the antibody complexes, while the target CD4 T cells remain unlabeled. Subsequently, the cell suspension is loaded onto a separation column; under the influence of a magnetic field, the magnetically labeled non‑target cells are retained within the column, whereas the unlabeled CD4 T cells elute by gravity into a collection tube. The resulting supernatant represents the CD4 T 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 CD4 T cells are key immune cells in the immune system, playing a central role in immune responses, immune regulation, and defense against infection and tumors. This kit employs a column-based negative selection approach: non-CD4 T cells are labeled with biotin-conjugated antibodies, and unwanted cells are removed using magnetic bead separation, thereby enriching highly pure, high‑activity CD4 T cells. This method avoids the potential impact of direct labeling on CD4 T cell function and is suitable for downstream experimental studies such as cell culture and functional assays.

Protocol

Note: This kit does not include the 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 then 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.

Sample Preparation

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 CD4 T cells.

1×10^7cells/100 μL

37cells/100 μL

3

Add 10 μL of Human CD4 T cell Biotin Antibody Cocktail to the sample and gently mix;

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

10 μL/100 μL

4

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

Incubate at room temperature for 10 minutes

5

Mix wellStreptavidin Beads, add 15 μLStreptavidin Beadsinto the sample;

15 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

In the column-based magnetic separator, installL 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; the 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 maximum number of labeled cells in an S Separation Column is1×107;The maximum number of labeled cells in an L Separation Column is1×108;



Picture

Validation Data

Human CD4 T cells were enriched from peripheral blood mononuclear cells (PBMCs) and, after sorting, stained with CD4-FITC (SDT Cat: S0B8380). Analysis was performed by gating on live cells (Live+). Flow cytometric results showed that the purity of the sorted Human CD4 T cells was 93.8%.