CD Marker Recognition of Human Cytokine-Induced Killer Cells by Flow Cytometry

Published: August 31, 2023

Abstract

Source: Hsiao, C. H., et al. Isolation and Expansion of Cytotoxic Cytokine-induced Killer T Cells for Cancer Treatment. J. Vis. Exp. (2020)

In this video, we demonstrate a flow cytometry-based technique to determine the proportion of cytokine-induced killer T-cells, CIK cells, expanded from a peripheral blood mononuclear cell culture.

Protocol

All procedures involving human participants have been performed in compliance with the institutional, national, and international guidelines for human welfare and have been reviewed by the local institutional review board. 1. CIK induction and expansion On Day 0, culture the PBMCs (1 x 106) in fresh basal medium containing 1,000 IU/mL of IFN-γ for 24 h in a humidified cell culture incubator at 37 °C and 5% CO2. On Day 1, re…

Representative Results

Figure 1: The proportion of CD3+CD56+ T cells from a representative PBMC sample. (A) Lymphocytes were recognized by specific size and granularity. Selected single cell population for analysis by flow cytometry. (B) Statistical analysis of CIK expansion efficacy from three healthy donors was conducted using a t-test (*, p < 0.01).

Divulgaciones

The authors have nothing to disclose.

Materials

APC Mouse Anti-Human CD56 antibody BD 555518 B159
APC Mouse IgG1, κ Isotype Control BD 555751 MOPC-21
BD FACSCanto II Flow Cytometer BD 338962 SN: R33896202856
Dulbecco's Modified Eagle Medium/F12 HyClone SH30023.02 Basal medium for OC-3 cell culture
Fetal bovine serum HyClone SH30084.03 For K562 and OC-3 cell culture. Complete medium contains 10% of FBS
Ficoll-Paque Plus GE Healthcare Life Sciences 71101700-EK Density gradient solution
FITC Mouse Anti-Human CD3 antibody BD 555332 UCHT1
FITC Mouse IgG1, κ Isotype Control BD 555748 MOPC-21
Human anti-CD3 mAb TaKaRa T210 OKT3
Add 2.5 mL of stock (1 mg/1 mL) to 50 mL of Induction medium. Storage stock at -80 °C
RPMI1640 medium Gibco 11875-085 Basal medium for K562 cell culture. Storage stock at 4 °C

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CD Marker Recognition of Human Cytokine-Induced Killer Cells by Flow Cytometry. J. Vis. Exp. (Pending Publication), e21559, doi: (2023).

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