PBMC Cell Electroporation Systems
A PBMC cell electroporation system includes an electroporation machine and compatible electroporation tubes. A few Celetrix electroporation models can be used for PBMC cell electroporation transfection with high-efficiency and low-toxicity. It can maintain a high level of PBMC cell survival, allowing immunotherapy applications such as CAR and TCR T-cell generation and CRISPR knockdown of T-cell genes. Transposon tools are available to increase transfection efficiency.
The electroporation conditions for PBMC (peripheral blood mononuclear cells), purified unstimulated T cells or NK cells are similar. Unstimulated T cells have a small cell diameter of only 7-8 um and stimulated T cells grow to 10-12 um diameter. Because of the small size, PBMC or unstimulated T cells require a much higher electroporaiton voltage than cell lines or stimulated T cells. The voltage is typically 1200 – 1400 V and only 20 ul and 100 ul tubes can be used under PBMC mode.
PBMCs can also be electroporated in the Legacy mode as we used in the previous Model LE/EX machines. Under Legacy mode, PBMCs need to be cooled down to 4oC by inserting the electroporation tube completely into ice in an upward direction for 4-6 minutes. After cooling, the cells can be electroporated immediately without wiping off the water drops on the tube. The parameter is 800 – 860 V 20 ms. Model EX+ machine can process 600 ul, or up to 200M PBMCs in Legacy mode. The cooling time for 600ul tube is 7-8 minutes and the parameter is 1350 – 1400 V 20ms.
Fresh PBMC cells survive quite well after electroporation. The true survival rate is over 60% calculated by the ratio of survived cells/initial cells. Frozen-thawed PBMC cells need about 30-40V more voltage than fresh PBMC cells.
Electroporation voltage are influenced by cell density and donor difference. For example, increasing from 2 M cells to 4 M cells in 20 ul electroporation requires a voltage increase of about 20 V. There might be +/- 20 V difference for 20 ul tubes due to donor difference.
Transfection efficiency is also heavily influenced by plasmid quality. Good quality GFP plasmid can produce over 80% transfection efficiency. mRNA typically has higher electroporation efficiency and for PBMC, mRNA transfection efficiency is usually >92% with good viability.
Contemporary Transposon Tools: A Review and Guide through Mechanisms and Applications of Sleeping Beauty, piggyBac and Tol2 for Genome Engineering: Transposons enable several avenues for genome manipulations in vertebrates, including transgenesis for the generation of transgenic cells in tissue culture comprising the generation of pluripotent stem cells, the production of germline-transgenic animals for basic and applied research, forward genetic screens for functional gene annotation in model species and therapy of genetic disorders in humans.
Celetrix PBMC cell electroporation machines:
Cell electroporation upgrade model LE+, new cell line mode and PBMC mode. for 20 ul, 100 ul, 120 ul, and 200 ul electroporation tubes.
Cell electroporation upgrade model EX+, new cell line mode and PBMC mode. for 20 ul, 100 ul, 120 ul, 200 ul, 600 ul, and 1 ml electroporation tubes.
Cell electroporation large-scale model SLT, great for large scale T cell electroporation. for 200 ul, 1 ml, 5 ml, and 10 ml electroporation tubes.
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transient gene expression (TGE).
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