The Cytek^® cFluor^® 21-Color Human T-Cell Kit enables the identification and characterization of major human T cell (CD3+) subpopulations (CD4+, CD8+, and TCRγδ+) in human peripheral mononuclear cells (PBMCs) and whole blood. The reagents in this kit also support detailed analysis of CD3+CD4+ and CD3+CD8+ naive, central memory, effector memory, and effector subsets, as well as specialized CD3+CD4+ helper T cell subsets, including Th1, Th2, Th17, Th22, T follicular helper (Tfh), and T peripheral helper (Tph) cells. Activation and exhaustion markers, including CD38, HLA-DR, CD366 (TIM-3), and TIGIT, are incorporated to assess activation and functional states across all defined T cell subsets.

This panel has been optimized and titrated for robust and reproducible immunophenotyping of T cells in PBMCs and whole blood. It provides a multiparametric approach to characterize T cell subset distribution, functional status, and activation/exhaustion phenotypes.

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PRODUCT DETAILS

Tested Dilution:  Use 5 µL / test for same-day, room-temperature protocols. For staining titers involving overnight incubation at 4°C, refer to the corresponding staining protocols.

Application: Flow Cytometry

Storage: 2-8°C and protected from Light. Do not freeze

Formulation: Phosphate-buffered saline, pH 7.2, containing 0.09% sodium azide and 0.2% BSA (BSA Country of Origin USA)
For nanoparticle dye reagents: Phosphate-buffered saline, pH 7.2, containing 0.1% PEG 3350, 0.1% Pluronic F-68, 0.05% Tween-20, 0.09% sodium azide and 1% BSA (BSA Country of Origin USA)

Data Sheets: TDS | SDS

The panel contains the following single vial reagents.

Table 1: Composition of the Cytek^® cFluor^® 21-Color Human T-Cell Kit

DATA

Figure 1: T cell subpopulation gating hierarchy. Example of gating strategy using a whole blood sample. Lymphocytes were identified from CD45+ single cells. CD3+ and TCRγδ+ T cells were defined. The CD3+ T cells were then classified into CD4+ and CD8+ T cells. CD4+, CD8+, and TCRγδ+ T cells were subsequently partitioned into naive and memory T cells. A) The memory CD4+ T cells were further divided into T helper subsets. B) Memory CD8+ T cells were further divided into cytotoxic T subsets.

RECOMMENDED USAGE

Human fresh whole blood (Heparin) and frozen PBMC samples have been tested to validate the performance of this kit. Please refer to the section below to download this kit's staining protocols, data acquisition protocols, experiment templates, and fluorochrome list.

Use appropriate personal protective equipment per product safety data sheet when using this product.

• Download the Sample Preparation Guidelines for the Cytek^® cFluor^® 21-Color Human T-Cell Kit for
  • Whole Blood, Same-Day Room Temperature Staining
  • Whole Blood, Overnight 4°C Staining
  • PBMCs, Same-Day Room Temperature Staining
  • PBMCs, Overnight 4°C Staining
• Download the Acquisition Guidelines for the Cytek^® cFluor^® 21-Color Human T-Cell Kit for
  • Whole Blood, Same-Day Room Temperature Staining
  • Whole Blood, Overnight 4°C Staining
  • PBMCs, Same-Day Room Temperature and Overnight 4°C Staining
• Download the Experiment Template for the Cytek^® cFluor^® 21-Color Human T-Cell Kit for
  • Whole Blood, Same-Day Room Temperature Staining
  • Whole Blood, Overnight 4°C Staining
  • PBMCs, Same-Day Room Temperature and Overnight 4°C Staining
• Download the Cytek^® cFluor^® 21-Color Human T-Cell Kit Assay Tags

REFERENCES

Mousset C.M., et al. 2019. Cytometry A. 95:647

Yoshitomi H., et al. 2020. Cell. Mol. Immunol. 18:523

Rasouli J., et al. 2015. J of Immunol. 194:5085

Laidlaw B.J., et al. 2016. Nat. Rev. Immunol. 16:102

Golubovskaya V., et al. 2016. Cancers. 8:36

Hu Y., et al. 2023. Signal Transduct. Target.Ther. 8:434

Baessler A., et al. 2024. Annu. Rev. Immunol. 42:179

Mosopust D., et al. 2026. Nat. Rev. Immunol. 26:298

For Research Use Only. Not intended for use in diagnostic procedures.

cFluor^® V547, cFluor^® B515, cFluor^® B548, cFluor^® BYG610, cFluor^® R685, cFluor^® R720, and cFluor^® R840 are equivalent to CF^®405L, CF^®488A, CF^®514, RPE-CF^®596R, CF^®660C, CF^®700, and APC-CF^®790T respectively, manufactured and provided by Biotium, Inc. under an Agreement between Biotium and Cytek (LICENSEE). The manufacture, use, sale, offer for sale, or import of the product is covered by one or more of the patents or pending applications owned or licensed by Biotium. The purchase of this product includes a limited, non-transferable immunity from suit under the foregoing patent claims for using only this amount of product for the purchaser’s own internal research. No right under any other patent claim, no right to perform any patented method, and no right to perform commercial services of any kind, including without limitation reporting the results of purchaser's activities for a fee or other commercial consideration, is conveyed expressly, by implication, or by estoppel.

cFluor^® UV395, cFluor^® V435, cFluor^® V605, cFluor^® V670, cFluor^® V715, and cFluor^® V780 are StarBright™ UltraViolet 400 Dye, StarBright™ Violet 440 Dye, StarBright™ Violet 610 Dye, StarBright™ Violet 670 Dye, StarBright™ Violet 710 Dye, and StarBright™ Violet 790 Dye respectively, manufactured and provided by Bio-Rad, Inc.

StarBright is a trademark of Bio-Rad Laboratories, Inc. in certain jurisdictions. StarBright Dyes are manufactured and provided by Bio-Rad under an Agreement between Bio-Rad supplied to Cytek, being subject to certain conditions.

cFluor^® BYG610, cFluor^® BYG667, cFluor^® BYG710, cFluor^® BYG750, and cFluor^® BYG781 are tandem dyes made with R-PE. cFluor^® R780 and cFluor^® R840 are tandem dyes made with APC. cFluor^® V605, cFluor^® V670, cFluor^® V715, and cFluor^® V780 are tandem dyes. Caution – Tandem dyes may show changes in their emission spectra with prolonged exposure to light or fixatives.