eoe sub articles

EoE Sub Articles

1. Preparation of Jurkat T cell line (Day 2)
NOTE: All procedures should be conducted in a class II biological safety cabinet.
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	<li>Culture Jurkat T cells in 24 mL of basal cell culture medium + 10% FBS + 5% penicillin/streptomycin at 37 &#176;C + 5% CO2 (Table of Material). Jurkat T cells are a suspension cell line that can be maintained through passaging 1:6-1:8 into pre-warmed culturing media every 3 days. Do not shake.</l...

an in vitro technique for uv irradiation-mediated generation of apoptotic t cells

Source: Hodge, M. X., et al. <a href="https://app.jove.com/v/60109">In Vivo Assessment of Alveolar Macrophage Efferocytosis Following Ozone Exposure.</a> J. Vis. Exp. (2019).
This video demonstrates a method to induce apoptosis in the Jurkat cells &#8212; a human T lymphocyte leukemia cell line. The cells are exposed to ultraviolet C (UVC) radiation, which causes extensive DNA damage and leads to apoptotic cell death.

An In Vitro Technique for UV Irradiation-Mediated Generation of Apoptotic T Cells

1. Preparation of Jurkat T cell line (Day 2)
NOTE: All procedures should be conducted in a class II biological safety cabinet.

	Culture Jurkat T cells in ...

Take a suspension of Jurkat cells &#8212; T lymphocyte leukemia cells.
Mix an aliquot with a cell viability dye and ensure an adequate number of living cells.
Plate the cells in a culture dish and apply ultraviolet C or UVC, radiation for the required duration.
UVC penetrates the cells and is absorbed by DNA, forming pyrimidine photoproducts.
Post-irradiation, incubate the cells. Extensive UVC-mediated DNA damage initiates the intrinsic apoptosis pathway, activating pro-apoptotic proteins on the outer mitochondrial membrane.
Activated proteins oligomerize, forming membrane pores and releasing cytochrome&#160; C from the intermembrane space into the cytoplasm.
Cytochrome C complexes with the apoptotic protease activating factor-1, forming an apoptosome complex. The apoptosome activates caspases, causing apoptosis.
Introduce fluorophore-labeled annexin V to label phosphatidylserine on the membrane, an apoptotic cell marker, and add propidium iodide that enters through the damaged membrane to label the DNA.
Using flow cytometry, determine the annexin V-labeled early apoptotic cells and dual-stained late apoptotic cells.