Figure 1. General Workflow for Cargo Delivery with the CellPore™ Transfection System

The CellPore™ Transfection System enables efficient delivery of various cargo(es) into a broad range of cell types, including immune cells, stem cells, and immortalized cell lines. The simple four-step workflow comprises preparing a single cell suspension in CellPore™ Delivery Medium, followed by direct addition of one or more cargo types (e.g. RNA, CRISPR-Cas9 ribonucleoproteins, antibodies). Then, the mixture is transferred to a single-use CellPore™ Delivery Cartridge to be processed on the CellPore™ instrument. Cells are mechanoporated during the run to enable cargo uptake directly into the cytosol for various applications.

Figure 2. CellPore™ Offers Broad Cell Type & Cargo Delivery Capabilities

The CellPore™ Transfection System is compatible with a wide range of cell types, including primary human and mouse immune cells, stem cells, and immortalized cell lines. The system mechanoporates cells to enable delivery of various cargo types directly to the cytosol where they can execute their functions. Cargos that contain a nuclear localization signal (NLS) can also traffic into the nucleus (e.g. Cas9 nuclease).

Figure 3. CellPore™ Enables Efficient Delivery of Cargo to Primary Immune Cells, Stem Cells, and Cell Lines

Cells were resuspended in appropriate CellPore™ Delivery Media and transfected using the CellPore™ Transfection System. (A) Cas9 RNPs targeting TRAC (TCRɑβ), B2M (MHC-I), CIITA (MHC-II), TIGIT (TIGIT), KLRC1 (NKG2A), or PTPRC (CD45) were successfully delivered to various primary immune cells, stem cells, and immortalized cell lines, resulting in the knockout of their respective surface marker with minimal impact to cell viability (relative to untreated controls). (B) Representative flow plot of co-delivery of mCherry and eGFP mRNA to human unactivated T cells, which resulted in over 90% co-expression of both constructs. (C) Delivery of a self-amplifying RNA construct encoding GFP (1.5 pmol, 5 μg) to human CD34+ hematopoietic stem & progenitor cells resulted in successful GFP expression expansion over 7 days (with 0.8 μg/mL puromycin; n = 2). (D) Delivery of a 3.5 Kb GFP-encoding plasmid DNA to Jurkats resulted in dose-dependent increase in GFP expression and maintenance of high viability (n = 4-6). Results measured via flow cytometry. Untreated refers to unmanipulated cells. No Cargo control refers to cells mechanoporated without cargo. Data presented as mean ± SD.

Figure 4. Mechanoporation Helps Maintain Better Cell Quality Compared to Electroporation

Unactivated human T cells were either mechanoporated or electroporated, and analyzed for their inflammatory responses and transcriptome integrity. (A) Heatmaps display the expression levels of 14 key cytokines and chemokines across 3 donors using a bead-based multiplex cytokine analysis array (24 hours post transfection). Unactivated T cells mechanoporated via CellPore™ maintained baseline cytokine expression (comparable to untreated samples), whereas considerable increases in the levels of several pro-inflammatory cytokines released in the supernatant were detected in electroporated samples (n = 3). (B) Volcano plots illustrate an overview of differential gene expression in unactivated T cells, 6 hours following transfection. Relative to an untreated control, mechanoporation safeguarded baseline gene expression, whereas electroporation resulted in more widespread gene dysregulation, indicated by the highlighted genes (red data points) with greater than two-fold change in expression (p value < 0.05, n = 3).