Cell Electroporation upgrade Model EX+

Catalog No.	11-0106
Product Name	Cell Electroporation upgrade Model EX+
Supplier Name	CELETRIX LLC
Brand Name	Celetrix
Summary	The PBMC cell electroporation upgrade model EX+ (11-0106) is an upgraded version of model LE+ (11-0101) with larger electroporation volume (600 ul and 1 ml electroporation tubes). If requested, it can be equipped with optional electrofusion accessories for hybridoma generation. Mouse spleen cells can be fused with myeloma cells such as SP2/0 and P3X63Ag8 with high efficiency. Keywords: cell electroporation, CHO electroporation transfection, CHO stable cell line development, PBMC cell electroporation, NK cell electroporation, T cell electroporation, recombinant stable cell line development.
Shipping	The PBMC cell electroporation upgrade model EX+ (11-0106) is shipped at ambient temperature.
Stability & Storage	The PBMC cell electroporation upgrade model EX+ (11-0106) is stable at room temperature.
Note	Products will be shipped from the warehouse in USA. Promotion is running from time to time. Welcome to send a request for quote to message@sydlabs.com.
Order Offline	Syd Labs, Inc. 4 Avenue E, Hopkinton, MA 01748 USA. Phone: 1-617-401-8149 Fax: 1-617-606-5022 Email: message@sydlabs.com.

Description

11-0106: PBMC Cell Electroporation upgrade Model EX+

1、New cell line mode and PBMC mode;
2、Voltage range 300 – 1500 V;
3、Optimized for large plasmid electroporation;
4、For 20 ul, 100 ul, 120 ul, 200 ul, 600 ul, and 1 ml electroporation tubes

Simplified operation with single adjustable parameter
For mammalian cells, with DNA, RNA, siRNA or proteins.
Integrated oscilloscope, electrical current pulse display.

The Celetrix PBMC cell-electroporation system, combining a powerful electroporator, innovative specialized electroporation tubes, and a proprietary electroporation buffer for cell protection, is designed to achieve high-efficiency and low-toxicity electroporation. The PBMC cell electroporation technology can maintain a high level of cell survival, allowing immunotherapy applications such as CAR and TCR T-cell generation and CRISPR knockdown of T-cell genes. Our PBMC cell electroporation machines can electroporate human PBMCs at an efficiency of 60–90%. For other types of eukaryotic cells, such as human iPSCs, primary neurons, and cell lines, the electroporation efficiency can reach more than 90% with very low cytotoxicity. Our PBMC cell electroporation system can also electroporate mRNA or protein (cas9/RNP) at nearly 100% efficiency.

Genome editing with Cell Electroporation Model LE+ and Model LE+
With Celetrix electroporation, we can deliver RNP (ribonucleoprotein complex containing Cas9 protein and gRNA) or Cas9/gRNA plasmid into cells to perform gene editing.

RNP electroporation has very low cell toxicity and the efficiency of gene editing can usually reach over 90%, with no apparent cell type limitation. For primary T cells, NK cells and stem cells, Cas9 plasmids have a high toxicity upon electroporation and we recommend using RNP on these cells.

T cell gene editing can be done with either unstimulated T cells (PBMC) or stimulated T cells, using different voltages. On the LE+/EX+ model machines, we can use the PBMC mode to electroporate RNP into PBMCs with a high voltage. Stimulated T cells are electroporated at 2-3 days of stimulation time and the voltage is similar to common cell lines, much lower than the voltage of PBMCs. For example, we can knock out the TCR using 2-3 day stimulated T cells and the efficiency of knockout is 92% and cell growth is not impaired.

T cell, NK cell and CD34+ HSC cells are important for gene and cell therapy applications. Our technical support can make gene editing in these cells very straightforward for you. Also, we have a large scale single tube electroporation machine that can process a 10 ml sample, or 2x10E9 cells in a single zap.

iPSC cells are also similar to common cell lines with Celetrix electroporation. RNP electroporation can achieve high efficiency gene editing in iPSCs, as shown with AAVS1 and OCT4 genes. One easy way to achieve high gene editing efficiency is to use relatively high concentration of RNP, such as 10 ug Cas9 and 5 ug gRNA in a 20 ul electroporation.

Plasmid based gene editing is suitable for cell lines. The cell toxicity of large plasmids containing Cas9 gene and gRNA gene can be tolerated by cell lines. The gene editing efficiency from plasmid electroporation is usually lower than the efficiency from RNP electroporation. However, we can use drug resistant gene selection to improve the gene editing efficiency. For example, THP-1 cells was electroporated with a plasmid containing Cas9, gRNA, GFP and puromycin-resistance genes and cells were all positive in GFP after puromycin selection, with a high rate of gene editing.

Normally it takes long time to do gene editing with plasmids. We have developed protocols to perform quick gene editing, even with plasmids. Because of the high cell viability and efficiency of Celetrix electroporation, drug selection can be done for a short period at the transient transfection stage and the remaining cells carry gene editing without stable integration of the plasmid. This new method speeds up plasmid based gene editing and allows sequential editing of multiple genes.

T Cell Immunotherapy with Celetrix electroporation
Human T cells have been hard to transfect with other methods, especially for plasmids.

With Celetrix electroporation, T cells can be transfected any time from the fresh PBMC stage to the stimulated or cultured stage. The Celetrix efficiency is highest compared to all other electroporation methods on the market. Our technology is also the only one that maintains high level of cell survival and expansion, allowing immunotherapy applications such as CAR, TCR-T generation and CRISPR knock-down of T cell genes.

Human PBMC transfection with GFP plasmid

Transfection efficiency with GFP plasmid after 24 hours is over 80%

Human PBMC transfection of CAR-T2A-GFP with SB transposon

4 days

14 days

20 days

14 days

Human PBMC CAR expression at 65.9% after 14 days.
Cells form clusters and proliferate similar to un-electroporated cells.

Electroporation—– The Future of CAR-T Therapy

Viral vectors have been the predominant delivery vehicles in immunotherapy, since the other existing electroporation methods fared poorly with immune cells.
The new high-performance Celetrix technology swiftly electroporates unstimulated PBMC cells and supports fast cell expansion, enabling clinical T cell transfection for CAR-T therapy.

References:

Optimized DNA electroporation for primary human T cell engineering: Electroporation of primary T cells. Electroporation was performed using the Celetrix electroporation system (Celetrix, Manassas, VA, USA), according to the following protocol.

The Past, Present, and Future of Non-Viral CAR T Cells: The electroporation efficiency depends on the voltage, the number of pulses, pulse width, and on the cell type and activation state. Furthermore, temperature, electroporation buffer, DNA and cell concentration influence the transduction efficiency. High-intensity pulses generally result in higher transduction efficiency but affect cellular viability. Small-scale electroporation can be achieved using Nucleofector 4D (Lonza, Basel, Switzerland) Neon (Thermo Fisher Scientific, Waltham) which use a cuvette and a pipette tip chamber, respectively. Other commercialized instruments are for example the Celetrix electroporation system (Celetrix, Manassas, VA, USA), and the BTX ECM 830 system (Harvard Bioscience, Hollistone USA).

Abstract A026: Novel electroporation method for quick CAR-T-cell manufacture: With a redesigned electroporation system, we can now achieve very high transfection efficiency for T-cells while maintaining cell survival. For Sleeping Beauty transposon-based CAR expression, we found that over a period of two to three weeks the efficiency can get to 60% to 90% with fasT-cell proliferation. The protein expression time after electroporation is very short. For simple GFP plasmids we can observe GFP expression after only 30 minutes. Unlike viral vectors, electroporation works well on fresh natural T-cells, thereby eliminating the need for expensive cell expansion and virus production altogether and cutting the huge economic burden of CAR-T therapy.

Development of a reporter gene method to measure the bioactivity of anti-CD38 × CD3 bispecific antibody: The plasmid was transfected into 2 × 107 Jurkat-CD3-NFAT-RE-Luc cells using cell electroporation (Celetrix, VI, USA; 1080 V, 30 ms, 1 pulse).

A review of emerging physical transfection methods for CRISPR/Cas9-mediated gene editing

Abstract A44: High-efficiency electroporation of nonexpanded T cells and NK cells for CAR-T and CAR-NK: We developed a novel electroporation technology that corrected many physics mistakes in the field. We can now achieve very high transfection efficiency for fresh T cells while maintaining cell survival. For Sleeping Beauty transposon-based CAR expression, we found that over a period of two to three weeks the efficiency can get to 60% to 90% with fast cell proliferation. The protein expression time after electroporation is very short. For simple GFP plasmids we can observe GFP expression after only 30 minutes. Similarly, for NK cells, we were able to achieve 42% transfection efficiency with fresh nonexpanded NK cells. We further compared post-electroporation cell proliferation potential between nonexpanded fresh T cells and stimulated/expanded T cells. It turned out that although fresh T cells are smaller in volume, they are much better than expanded T cells in survival after electroporation. Expanded NK cells also showed much higher cytotoxicity than fresh NK cells after electroporation. With our new electroporation technology, CAR-T and CAR-NK can now be manufactured quicker and better. Our new protocols focusing on fresh T cells and fresh NK cells would be a great help for the field.

Celetrix electroporation machines:

Cell electroporation basic model SP100, catalog No. 11-0103, simplified single pulse for all cell types; for 20 ul, and 100 ul electroporation tubes

Cell electroporation upgrade model LE+, catalog No. 11-0101, new cell line mode and PBMC mode. for 20 ul, 100 ul, 120 ul, and 200 ul electroporation tubes.

Cell electroporation upgrade model EX+, catalog No. 11-0106, 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, catalog No. 11-0104, for 200 ul, 1 ml, 5 ml, and 10 ml electroporation tubes.

Cell electroporation upgrade model EX+ with the electrofusion function, catalog No. 11-0106F, for 20 ul, 100 ul, 120 ul, 200 ul, 400 ul, 600 ul, and 1 ml electroporation tubes. Model EX+ and Model SLT can be equipped with optional fusion accessories for hybridoma generation. Mouse spleen cells can be fused with myeloma cells such as SP2/0 and P3X63Ag8 with high efficiency.

UHV Transformer (Ultra High Voltage), catalog No. 11-0201, bacteria cells: 20 ul, 100 ul, and 600 ul electroporation tubes; yeast cells: 20 ul, 100 ul, 200 ul, and 1000 ul electroporation tubes.

UHV-Plus (Ultra High Voltage), catalog No. 11-0202, bacteria cells: 20 ul, 100 ul, and 600 ul electroporation tubes; yeast cells: 20 ul, 100 ul, 200 ul, and 1000 ul electroporation tubes; mammalian cells: 20 ul and 200 ul electroporation tubes.

Please remember our product information: Cell electroporation upgrade model EX+ catalog number: 11-0106 Celetrix.

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