Helmholtz Centre for Environmental Research - UFZ, Department of Environmental Immunology, Leipzig, Germany.
Perinatal Immunology Research Group, Medical Faculty, Saxonian Incubator for Clinical Translation (SIKT), University of Leipzig, Leipzig, Germany.
Front Immunol. 2024 Jan 2;14:1327960. doi: 10.3389/fimmu.2023.1327960. eCollection 2023.
There is a growing need for immunological assays to test toxic and modulatory effects of chemicals. The assays should be easy to use, reproducible and superior to cell line-based assays. We have therefore developed a comprehensive portfolio of assays based on primary human blood cells that are suitable for testing chemical effects.
The flow cytometry-based assays were designed to target a wide range of human peripheral blood mononuclear cells and whole blood, including T cells, NK cells, B cells, basophils and innate-like T cells such as γδT, MAIT and NKT cells. We have selected a set of activation markers for each immune cell, e.g: CD154 (T cells), CD137, CD107a (NK cells), CD63 (basophils), CD69, CD83 (B cells), CD69, IFN-γ (MAIT cells) and we selected cell specific stimuli: aCD3 antibodies (T cells); E. coli and cytokines IL-12/15/18 (MAIT cells); CpG ODN2006, R848 or aCD40 antibodies (B cells), fMLP or aFcϵR1 (basophils) or K562 cells (NK cells).
By selecting immune cell-specific markers and cell-specific stimuli, we were able to induce particular immune responses from the targeted immune cells. For example, the response to stimulation with anti-CD3 antibodies was in 36.8% of CD107a+CD8+ cells. Cytokine stimulation induced the production of IFN-γ in 30% of MAIT cells. After stimulation with , around 50% of MAIT cells produced TNF. About 40% of basophils responded to aFcƐR1 stimulation. Similar activation ranges were achieved in K562-stimulated NK cells.
Our test portfolio covers the most relevant immune cells present in human blood, providing a solid basis for toxicity and immunomodulatory testing of chemicals. By using human blood, the natural composition of cells found in the blood can be determined and the effects of chemicals can be detected at the cellular level.
人们越来越需要免疫学检测来测试化学物质的毒性和调节作用。这些检测方法应该易于使用、可重现，并且优于基于细胞系的检测方法。因此，我们开发了一系列基于原代人外周血单个核细胞的综合检测方法，这些方法适用于测试化学物质的作用。
基于流式细胞术的检测方法旨在针对广泛的人类外周血单个核细胞和全血，包括 T 细胞、NK 细胞、B 细胞、嗜碱性粒细胞和先天样 T 细胞，如 γδT、MAIT 和 NKT 细胞。我们为每种免疫细胞选择了一组激活标志物，例如：CD154（T 细胞）、CD137、CD107a（NK 细胞）、CD63（嗜碱性粒细胞）、CD69、CD83（B 细胞）、CD69、IFN-γ（MAIT 细胞），我们还选择了细胞特异性刺激物：抗 CD3 抗体（T 细胞）；E.coli 和细胞因子 IL-12/15/18（MAIT 细胞）；CpG ODN2006、R848 或抗 CD40 抗体（B 细胞）、fMLP 或抗 FcεR1（嗜碱性粒细胞）或 K562 细胞（NK 细胞）。
通过选择免疫细胞特异性标志物和细胞特异性刺激物，我们能够从靶向免疫细胞中诱导特定的免疫反应。例如，抗 CD3 抗体刺激的反应在 36.8%的 CD107a+CD8+细胞中。细胞因子刺激诱导 MAIT 细胞产生 IFN-γ的比例为 30%。用 R848 刺激后，大约 50%的 MAIT 细胞产生 TNF。约 40%的嗜碱性粒细胞对抗 FcεR1 刺激有反应。在 K562 刺激的 NK 细胞中也达到了类似的激活范围。
我们的测试组合涵盖了人类血液中存在的最相关的免疫细胞，为化学物质的毒性和免疫调节测试提供了坚实的基础。通过使用人血，可以确定血液中天然存在的细胞组成，并在细胞水平上检测化学物质的作用。
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