Optimizing Celetrix electroporation is easy. Start from our recommended parameters, run a quick GFP plasmid test, and fine-tune the voltage for your cell density, temperature and payload.
| Cell health | Culture cells to 50–70% of their maximum density before electroporation, and keep this consistent. |
| Cell density | Cell lines: 0.5–2×106 per 20 µl (typically 1×106) or 2.5–10×106 per 100 µl. PBMC: 1–5×106 per 20 µl. Higher density needs slightly higher voltage. |
| Single cells | Avoid clumps: clumped cells transfect poorly. Do not over-digest adherent cells. |
| Temperature | Room temperature (about 23 °C) works best. At 4 °C, voltage needs to be 10–20% higher. Adjust voltage if your lab is very warm or cold. |
| DNA quality | Use high-purity plasmid at 50–200 µg/ml. More DNA raises efficiency but also toxicity, so lower the voltage slightly. |
| Payload | Large plasmid < small plasmid < Cas9 RNP / siRNA (voltage needed). |
| Recovery | Recover in antibiotic-free medium; antibiotics can be added from day 2. After high voltage, plate cells at higher density. |
Pulse settings stay the same; only the voltage changes. Example based on 500 V in a 20 µl tube:
| 20 µl | 500 V (base) |
| 100 µl | 510 V (20 µl + 10 V) |
| 120 µl | 700 V (20 µl × 1.4) |
| 200 µl | 1200 V (20 µl × 2.4) |
| 600 µl | 800 V (20 µl × 1.6) |
| 1 ml | 1250 V (20 µl × 2.5) |
Easy-to-miss details
Electroporation uses a brief electric pulse to drive DNA, mRNA, siRNA, proteins or Cas9 RNP across the cell membrane. The voltage falls mostly on the cell membrane, so the inside of the cell is protected. Celetrix electroporators suspend cells in a protective Celetrix buffer to keep viability high.
With Celetrix electroporation, start from the recommended voltage for your cell type and test 3 voltages, 20 V apart, with a GFP plasmid. Use healthy, single cells and electroporate each sample right after loading the tube.
Some cell death is normal. At the right voltage, about 10–20% of cells die after Celetrix electroporation; if almost none die, the voltage may be too low. If there is a lot of cell death, the voltage is likely too high: lower it by 20–30 V and test 2–3 voltages. Always recover freshly electroporated cells in antibiotic-free medium.
Electroporation acts at the cell membrane, so its small toxicity stays at the membrane and has little effect inside the cell. Lipid and other chemical transfection reagents affect not only the membrane but also components inside the cell. That is why Celetrix electroporation keeps cell toxicity low and cells healthier than transfection reagents. Learn how the voltage falls on the cell membrane in our electroporation mechanism page.
Start from the Celetrix recommended parameters for your cell type. On Celetrix electroporators, large cells such as cell lines and activated T, B and NK cells usually need 460–600 V in Cell Line mode (20 µl tube), while small cells such as unstimulated PBMC need a higher voltage in PBMC mode. Keep cell density fixed, then fine-tune the voltage in 20 V steps.
The best voltage depends on your sample. Higher cell density needs a slightly higher voltage. With larger plasmids, cells tolerate high voltage less well, so lower the voltage slightly. That is why Celetrix gives a voltage range instead of one fixed voltage: choose a value within the range to match your cell density and plasmid size.
For Celetrix electroporation of cell lines, use 0.5–2×106 cells per 20 µl tube (typically 1×106) or 2.5–10×106 per 100 µl. Celetrix tubes scale from 20 µl to 1 ml with the same protocol.
Use Celetrix electroporation buffer. It protects cells during the pulse and is matched to Celetrix voltage settings, so results stay consistent from small to large volumes.
Celetrix electroporation works with a wide range of cells: cell lines (e.g., HEK293, CHO, Jurkat, K562), primary cells, stem cells such as iPSCs and MSCs, and PBMC, T, B and NK cells. See the Celetrix recommended parameters for more than 60 cell types.
Yes. With Celetrix electroporators, Cas9 RNP and siRNA usually need a slightly higher voltage than small plasmids, while large plasmids need a lower voltage.
