Isolation of Microparticles Derived from Apoptotic T Lymphocytes In Vitro
Published: August 31, 2023
Abstract
Source: Yang, C. et al., Generation of Lymphocytic Microparticles and Detection of their Proapoptotic Effect on Airway Epithelial Cells. J. Vis. Exp. (2015)
This video demonstrates the generation and isolation of T lymphocyte-derived microparticles, LMPs, from T lymphocytes following treatment with actinomycin-D.
Protocol
1. LMPs Production and Characterization NOTE: To prevent contamination, ensure that all materials used in this experiment are sterile or autoclaved. Perform all steps at RT in a biological safety cabinet under sterile conditions, unless otherwise indicated. Stimulation and Collection of MPs Thaw an aliquot of 10 million CEM T cells in a 37 °C water bath. Dilute in 10 ml pre-warmed serum-free hematopoietic medium such as X-VIVO…
Declarações
The authors have nothing to disclose.
Materials
| CEM T cells | ATCC | CCL-119 | |
| X-VIVO 15 medium | Cambrex, Walkersville | 04-744Q | |
| Flask T75 | Sarstedt | 83.1813.502 | |
| Flask T175 | Sarstedt | 83.1812.502 | |
| Actinomycin D | Sigma Chemical Co. | A9415-2mg | |
| PBS | Lifetechnologies | 14190-144 | |
| 0.22µm filter | Sarstedt | 83.1826.001 | |
| Cell incubator | Mandel | Heracell 150 | |
| Low speed centrifuge | IEC | Centra8R | |
| High speed centrifuge | Beckman | Avanti J8 | |
| High speed rotor for 250ml bottle | Beckman | JLA16.250 | |
| High speed rotor for 50ml tube | Beckman | JA30.50 |
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Isolation of Microparticles Derived from Apoptotic T Lymphocytes In Vitro. J. Vis. Exp. (Pending Publication), e21570, doi: (2023).