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| Product Name | Human Lymphocyte Subgroup Typing Kit |
| Catalog No. | RESK-0188 |
| Description | Based on biological functions and cell-surface antigen expression, human peripheral blood lymphocytes can be divided into three populations: T lymphocytes, B lymphocytes, and NK cells. T lymphocytes mainly participate in specific antigen-mediated cellular immune responses and regulate the secretion of immunoglobulins by B lymphocytes. T lymphocytes can be further subdivided by function into helper/inducer T cells and suppressor/cytotoxic T cells. NK cells (natural killer cells) mediate cytotoxicity against certain tumor cells and virus-infected cells independently of MHC class I and class II molecules. The Human Lymphocyte Subgroup Typing Kit consists of a series of dual-color fluorescently labeled antibodies. It is mainly used to determine the percentage of each lymphocyte subset in total peripheral blood lymphocytes, namely mature T lymphocytes (CD3+), helper/inducer T cells (Th/i: CD3+CD4+), suppressor/cytotoxic T cells (Ts/c: CD3+CD8+), B lymphocytes (CD19+), and NK cells (CD3-CD16+ and/or CD56+), and can also be used to calculate the Th/Ts (CD3+CD4+/CD3+CD8+) cell ratio. |
| Intended Use | For the flow cytometric determination of the percentages of human peripheral blood lymphocyte subsets (mature T cells, helper/inducer T cells, suppressor/cytotoxic T cells, B cells, and NK cells) within total lymphocytes, and for calculation of the CD3+CD4+/CD3+CD8+ (Th/Ts) ratio. |
| Principle / Technology | Dual-color fluorescently labeled antibodies analyzed by flow cytometry. Compared with traditional lymphocyte subset analysis methods, flow cytometry combines the specificity of antibodies with the sensitivity of single-cell analysis by the flow cytometer, making this analysis method more accurate and more convenient. |
| Detection Method | Flow cytometry. Excitation wavelength Ex = 488 nm. The orange-red fluorescence of PE (Em = 578 nm) is recommended to be detected in the FL2 channel; the maximum excitation wavelength of FITC is Ex = 488 nm and the maximum emission wavelength is Em = 525 nm, and the green fluorescence of FITC is detected in the FL1 channel. |
| Sample Type | EDTA-anticoagulated venous blood (at least 1 mL). Staining and analysis are recommended within 6 h after collection. The leukocyte concentration of the tested peripheral blood should be 3.0 x 10^3 - 10.0 x 10^3 cells/uL. If it is higher than 10.0 x 10^3 cells/uL, dilute the blood sample; if it is lower, increase the sample volume or isolate leukocytes. |
| Reaction Conditions / Protocol | 1. Dilute the red blood cell lysis solution to a 1x working solution with deionized water. The hemolytic efficiency of the lysis solution is affected by temperature, so equilibrate it to room temperature (20-25 °C) before use. Note: after dilution, store protected from light at room temperature, and dilute according to the amount needed each time. 2. For each patient specimen, use six 12 x 75 mm flow cytometry tubes labeled with the sample number (1, 2, 3, ...) and the tube letters (A, B, C, D, E, F). 3. Add 20 uL of Reagent A, B, C, D, E, and F to the corresponding tube of each sample. 4. Add 100 uL of anticoagulated whole blood to each tube, mix thoroughly, and react at room temperature (20-25 °C) protected from light for 20-30 min. 5. Add 2 mL of (1x) red blood cell lysis solution (20-25 °C) to each tube, mix thoroughly, and react at room temperature protected from light for 10-12 min until the liquid becomes clear. 6. Centrifuge at 1,000 rpm for 5 min and discard the supernatant. 7. Add 2 mL of PBS to wash, centrifuge at 1,000 rpm for 5 min, and discard the supernatant. 8. Add 0.5 mL of 2% formaldehyde solution to fix the cells, mix, and analyze on the flow cytometer within 24 h. Note: if the stained cells are analyzed on the flow cytometer immediately, formaldehyde fixation is not required; simply wash and resuspend the cells in 0.5 mL of PBS before analysis. |
| Components / Formulation | A series of dual-color fluorescently labeled antibody reagents (Reagents A-F) and a red blood cell lysis solution. Reagent A is used as the negative control and Reagent B is used for lymphocyte gating. |
| Storage Conditions | Store at 2-8 °C protected from light. Strictly do not freeze. Avoid direct exposure to light. |
| Package Specifications | 50 T |
| Product Form | Liquid reagents |
| Regulatory / Compliance | For research use only. |
| Application Notes / Precautions | Equilibrate the red blood cell lysis solution to room temperature (20-25 °C) before use; store the diluted lysis solution protected from light at room temperature. If cells are analyzed immediately after staining, fixation with formaldehyde is not required; wash and resuspend the cells in 0.5 mL of PBS before analysis. Staining and analysis are recommended within 6 h after blood collection. |
For research use only, not for clinical use.
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