Human CD66b+ Cell Isolation Kit (positive selection)

1. Mouse Cells

2. Human Cells

1.High Purity: Efficiently captures CD66b+ cells to obtain highly enriched target cells.
2.High Viability: The gentle isolation process helps maintain high cell viability and preserves cells for downstream applications.
3.Easy Operation: Column-free separation using a simple magnetic rack.

Suitable for isolating CD66b+ cells from human peripheral blood.

1.Cell Sample Preparation: Collect fresh anticoagulated whole blood into a centrifuge tube and perform red blood cell (RBC) lysis.
Note: EDTA-anticoagulated whole blood is recommended. The volume of lysis buffer and lysis time can be adjusted as needed. If initial lysis is incomplete, a second round of lysis may be performed. A minor residual amount of RBCs will not significantly affect isolation purity.
2.Cell Washing: Following RBC lysis, resuspend the cell pellet in PBS and wash by centrifuging at 500 x g for 5 min.
3.Resuspension: Discard the supernatant, resuspend cells in Isolation Buffer, and adjust cell concentration to 1 x 10^8 cells/mL.
Note: Recommended Isolation Buffer: PBS containing 2 mM EDTA and 2% FBS. The buffer must be sterile-filtered through a 0.22 μm membrane filter prior to use.
4.Antibody Labeling: Add 500 μL cell suspension (5 x 10^7 cells) directly to the bottom of a sterile flow cytometry tube, add 10 μL CD66b Capture Antibody, mix thoroughly, and incubate at 4°C for 15 min.
Note: Avoid running cell suspensions down the tube wall when adding cells. Scale up the CD66b Capture Antibody volume proportionally when isolating larger cell numbers. If isolating ＜ 1 x 10^7 cells, top up the cell suspension volume to 100 μL and add 2 μL CD66b Capture Antibody. Centrifuge tubes may also be used depending on the compatibility of the magnetic rack.
5.Bead Pretreatment: Resuspend magnetic beads by vortexing. Transfer the required amount of beads into a 1.5 mL tube, add 1 mL Isolation Buffer, centrifuge at 10,000 x g for 1 min, and discard the supernatant. Repeat the wash step once. Resuspend beads in Isolation Buffer back to their original volume. (e.g., if 20 μL beads were washed, resuspend in 20 μL Isolation Buffer after washing).
6.Bead Binding: After antibody incubation, add 100 μL pretreated Releasable Magnetic Beads to the flow tube, mix thoroughly, and incubate at 4°C for 15 min.
Note: Scale up Releasable Magnetic Beads volume proportionally for larger cell numbers. If isolating ＜ 1 x 10^7 cells, add 20 μL Releasable Magnetic Beads.
7.Magnetic Separation: After incubation, add 2.5 mL Isolation Buffer to the flow tube and gently pipette up and down to mix. Avoid vigorous vortexing or tube inversion. Place the flow tube on a magnetic rack and let it stand for 5 min.
8.Bead Washing: Discard the supernatant. Remove the flow tube from the magnetic rack, immediately add 2 mL Isolation Buffer, and gently pipette up and down several times to disperse the beads. Place the tube back onto the magnetic rack and let it stand for 5 min.
9.Repeat Washing: Repeat Step 8 two additional times.
Note: Thorough washing is critical to achieving high purity of target cells during downstream elution.
10.Bead Dissociation (First Elution): After magnetic separation, discard the supernatant. Remove the tube, immediately add 1 mL Magnetic Beads Release Buffer to resuspend the beads and prevent them from drying out. Transfer the bead slurry to a 1.5 mL microcentrifuge tube and incubate on a rotary mixer at room temperature for 10 min.
Note: The volume of Magnetic Beads Release Buffer can be scaled according to cell input. If isolating ＜ 1 x 10^7 cells, use 200 μL Magnetic Beads Release Buffer for elution.
11.Cell Collection (First Elution): After incubation, pipette up and down at least 10 times, transfer the slurry to a new flow tube, top up Isolation Buffer to 1.5 mL, and mix gently by pipetting. Place the flow tube on the magnetic rack and let it stand for 5 min.
12.Bead Dissociation (Second Elution): The supernatant contains the target cells; transfer this supernatant to a 15 mL centrifuge tube for collection. Immediately add 1 mL Magnetic Beads Release Buffer to the remaining magnetic beads to keep them moist, transfer the slurry to a 1.5 mL tube, and incubate on a rotary mixer at room temperature for 10 min.
13.Cell Collection (Second Elution): Repeat the cell elution steps (Step 11) once more and retain the supernatant.
14.Final Cell Pooling: Combine the second eluate supernatant with the first eluate, centrifuge at 500 x g for 5 min, and discard the supernatant to yield bead-free CD66b+ cells.
15.Downstream Preparation: Wash cells as required by downstream protocols, then resuspend in the desired buffer or culture medium for subsequent molecular or cellular experiments.

4°C, 2 years.

1.Do not freeze any kit components. Magnetic beads should be stored in their storage solution to prevent drying.
2.Resuspend the beads thoroughly by vortexing before pipetting from the stock tube, taking care to avoid air bubbles.
3.High-quality low-binding pipette tips and reaction tubes are recommended to prevent bead and solution loss due to wall adhesion.
4.This product is intended for scientific research purposes by professionals only and must not be used for clinical diagnosis or treatment, food, or medicine, nor stored in residential areas.
5.For your safety and health, please wear a lab coat and disposable gloves during operation.