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ProteanFect™ Max Transfection Kit
ProteanFect™ Max Transfection Kit is designed to efficiently transfect a wide range of primary cells and hard-to-transfect cell lines with high efficiency, while maintaining an excellent safety profile. It facilitates easy scale-up and meets the demands of high-throughput experiments. Additionally, it is user-friendly, making transfection of primary cells as straightforward as transfecting HEK293T cells. For research use only.
| Number | Standard | Price(USD) | Stock |
|---|---|---|---|
| PT02-0002B | 0.02 mL | inquire | inquire |
| PT02-0010B | 0.1 mL | Inquire | Inquire |
| PT02-0100B | 1.0 mL | Inquire | Inquire |
Transfection Performance
Publication
Zhu, P. et al. Efficient Generation of CAR-T By the First Coacervate-Based Gene Delivery System. ASH Annual Meeting & Exposition, (2024).
Gao, X. et al. The First Coacervate-based Delivery System for Advanced Gene Therapy. ASGCT 28th Annual Meeting Abstracts, (2025).
Gao, X. et al. The First Coacervate-Based Delivery System for CAR-T Cell Engineering and Manufacturing. Presented at ASGCT 28th Annual Meeting Abstracts.
Documents
Product Information
Components & Storage
The kit is shipped on dry ice. Once received, store the components as indicated below. The kit includes positive control samples with EGFP-encoding mRNA and plasmid DNA (pDNA) to verify transfection efficiency.
| Components | Storage |
|---|---|
| Reagent A | 4°C |
| Reagent B | -20°C |
| Reagent C | 4°C |
| EGFP mRNA (1 µg/µL) | -20°C |
| EGFP pDNA (0.5 µg/µL) | -20°C |
Shipping Information
Shipped on Dry Ice
Compatible Cell Lines
Primary Cells and Cell Lines Successfully Transfected Using ProteanFect™ Max Transfection Kit
| Cell lines | Tested Nucleic Acid Types for Transfection |
|---|---|
| Human primary T cells | mRNA, siRNA |
| Human primary NK cells | mRNA, siRNA |
| Human primary monocytes | mRNA, siRNA |
| Human primary CD34+ hematopoietic stem cells | mRNA, siRNA |
| Mouse primary cardiomyocytes | mRNA, siRNA |
| Mouse primary neurons | pDNA, mRNA, siRNA |
| Mouse primary glial cells | pDNA, mRNA, siRNA |
| Mouse primary NK cells | mRNA, siRNA |
| Large yellow croaker primary mesenchymal stem cells | mRNA, siRNA |
| Jurkat T (human T lymphoblastic leukemia cells) | pDNA, mRNA, siRNA |
| LX-2 (human hepatic stellate cells) | pDNA, mRNA, siRNA |
| HepG2 (human liver tumor cells) | pDNA, mRNA, siRNA |
| THP-1 (human acute monocytic leukemia cell line) | mRNA, siRNA |
| Raji (human Burkitt lymphoma cells) | mRNA, siRNA |
| K562 (human chronic myeloid leukemia cells) | pDNA, mRNA, siRNA |
| MOLT-16 (human T lymphoblastic leukemia cells) | mRNA, siRNA |
| SH-SY5Y (human neuroblastoma cells) | pDNA, mRNA, siRNA |
| U2OS (human osteosarcoma cells) | pDNA, mRNA, siRNA |
| HFF (human foreskin fibroblasts) | mRNA, siRNA |
| HEK-293 (human embryonic kidney cell line) | pDNA, mRNA, siRNA |
| MC38 (mouse colon cancer cells) | mRNA, siRNA |
| RAW264.7 (mouse mononuclear macrophage leukemia cells) | mRNA, siRNA |
| LLC (mouse Lewis lung cancer cells) | pDNA, mRNA, siRNA |
| C2C12 (mouse myoblasts) | pDNA, mRNA, siRNA |
| COS7 (African green monkey kidney fibroblast-like cells) | pDNA, mRNA, siRNA |
| Human primary B cells | mRNA, siRNA |
| iPSCs ( Induced Pluripotent Stem Cells) | pDNA, mRNA, siRNA |
| UC-MSCs (Umbilical Cord Mesenchymal Stem Cells) | mRNA, siRNA |
| KSL (Mouse Hematopoietic Stem Cells) | mRNA, siRNA |
| Porcine Macrophages | mRNA, siRNA |
| Bovine Fibroblasts | mRNA, siRNA |
| M. rosenbergii Hemocytes | mRNA, siRNA |
| Chicken Primordial Germ Cells | mRNA, siRNA |
| L. crocea Mesenchymal Stem Cells | mRNA, siRNA |
| MEC-1 (Human B Cells) | mRNA, siRNA |
| TMD8 (Human Diffuse Large B-Cell Lymphoma Cells) | mRNA, siRNA |
| NK-92 (Human Natural Killer Cells) | mRNA, siRNA |
| Kasumi-1 (Human Acute Myeloid Leukemia) | mRNA, siRNA |
| U937 (Human Myeloid Leukemia) | mRNA, siRNA |
| HL-60 (Human Promyelocytic Leukemia) | mRNA, siRNA |
| KYSE-510 (Human Esophageal Squamous Cell Carcinoma) | mRNA, siRNA |
| Mum2B (Human Melanoma) | mRNA, siRNA |
| H1-hESC (Human Embryonic Stem Cells) | pDNA, mRNA, siRNA |
| HT-29 (Human Colorectal Adenocarcinoma Cells) | pDNA, mRNA, siRNA |
| 5637 (Human Bladder Carcinoma Cells) | pDNA, mRNA, siRNA |
| HGC-27 (Human Gastric Carcinoma Cells) | mRNA, siRNA |
| U251MG Cells (Human Glioblastoma Cells) | mRNA, siRNA |
| Neuro 2A (Mouse Neuroblastoma Cells) | pDNA, mRNA, siRNA |
| CH12 (Mouse Lymphoma Cells) | mRNA, siRNA |
| BV-2 (Mouse Microglial Cells) | mRNA, siRNA |
| B16 (Mouse Melanoma Cells) | mRNA, siRNA |
| MH-S (Mouse Macrophages) | mRNA, siRNA |
| MODE-K (Mouse Intestinal Epithelial Cells) | mRNA, siRNA |
| MEF (Mouse Embryonic Fibroblasts) | pDNA, mRNA, siRNA |
| 3T3 (Mouse Embryonic Fibroblasts) | mRNA, siRNA |
| OLN93 (Rat Oligodendrocyte Cells) | mRNA, siRNA |
| MDBK (Bovine Kidney Epithelial Cells) | mRNA, siRNA |
| MAC-T (Bovine Mammary Epithelial Cells) | mRNA, siRNA |
Product Data
Transfection Protocol for mRNA per Well of a 96-Well Plate
| Scheme | Step | Suspension Transfection |
|---|---|---|
 |
1. Transfection Complex Preparation |
1.1 Mix nucleic acids with Reagent A. 1.2 Add Reagent B, and mix gently by pipetting up and down 20-30 times or vortexing for 10 seconds. 1.3 (Optional) Add Reagent C. Mix gently by pipetting up and down 2-3 times or vortexing for 2-3 seconds. |
 |
2. Cell Preparation |
2.1 Harvest the cells by centrifugation at 300 g for 5 minutes. 2.2 Discard the supernatant and wash once with Opti-MEM. 2.3 Resuspend in Opti-MEM and adjust the concentration to 5×10⁶-1×10⁷ cells/mL. |
 |
3. Transfection |
3.1 Mix the transfection complex with the cell suspension and gently pipet up and down 2-3 times. 3.2 Incubate for 15-30 minutes (typically 15-30 minutes for primary cells and 30-60 minutes for cell lines) in a cell culture incubator. 3.3 Add 10× volume of culture medium relative to the cell suspension. |
 |
4. Cell Culture and Test |
4.1 Transfer the cells from the tube to the culture plate. 4.2 Incubate transfected cells and assess efficiency after 5 to 48 hours, or at an appropriate time. |
| Scheme | Step | Adherent Transfection |
|---|---|---|
 |
1. Cell Preparation |
1.1 Seed cells one day before transfection. |
 |
2. Transfection Complex Preparation |
2.1cMix nucleic acids with Reagent A. 2.2 Add Reagent B, and mix gently by pipetting up and down 20-30 times or vortexing for 10 seconds. 2.3 (Optional) Add Reagent C. Mix gently by pipetting up and down 2-3 times or vortexing for 2-3 seconds. |
 |
3. Transfection |
3.1 Maintain 50%-80% cell confluence before transfection. 3.2 Remove medium, wash cells once with Opti-MEM, then add Opti-MEM. 3.3 Apply the transfection complex directly to the seeded cells. 3.4 Incubate for 15-30 minutes (typically 15-30 minutes for primary cells and 30-60 minutes for cell lines) in a cell culture incubator. |
 |
4. Cell Culture and Test |
4.1 Replenish with the cell culture medium. 4.2 Incubate transfected cells assess efficiency after 5 to 48 hours, or at an appropriate time. |
Technical FAQ
1.1 Optimize Transfection Parameters
Optimize transfection parameters for each cell type. Extended incubation time: Adjust the incubation time of the transfection complex with cells. The maximum incubation time is 2 hours for cell lines, and 30 minutes for primary cells. Increase transfection complex: Consider increasing the amount of transfection complex to improve transfection efficiency.
1.2 Severe Cytotoxicity Caused by Plasmid DNA
The transfection of pDNA into primary cells, such as primary T cells, can induce cytotoxicity and inflammatory responses. Due to the risk of significant toxicity, pDNA transfection is generally not recommended for primary T cells.
1.3 Improve Plasmid DNA Quality
Transfection efficiency is highly dependent on the quality of the plasmid DNA. Endotoxin-free: Use an endotoxin-free kit to prepare plasmid DNA. Optimal concentration: Ensure an OD260/280 ratio between 1.7 and 1.9, and dilute DNA to 0.5–2 µg/µL using nuclease-free water.
1.4 Improve Cell Condition
For cell lines, transfect cells with >90% viability, confirmed by trypan blue exclusion. Avoid using cells beyond 15 passages, and allow 2-3 passages for recently thawed cells to stabilize before transfection.
For primary cells, proper activation is crucial for optimal transfection efficiency. For example, human primary T cells generally achieve the best transfection results after stimulation with anti-CD3/CD28 activation beads or antibodies for 2-10 days. Do not remove the activation beads until transfection.
1.5 Use Positive Control
We recommend using a 96-well plate format to optimize transfection conditions for a specific cell type, with EGFP mRNA as the positive control.
Optimize transfection parameters for each cell type. Extended incubation time: Adjust the incubation time of the transfection complex with cells. The maximum incubation time is 2 hours for cell lines, and 30 minutes for primary cells. Increase transfection complex: Consider increasing the amount of transfection complex to improve transfection efficiency.
1.2 Severe Cytotoxicity Caused by Plasmid DNA
The transfection of pDNA into primary cells, such as primary T cells, can induce cytotoxicity and inflammatory responses. Due to the risk of significant toxicity, pDNA transfection is generally not recommended for primary T cells.
1.3 Improve Plasmid DNA Quality
Transfection efficiency is highly dependent on the quality of the plasmid DNA. Endotoxin-free: Use an endotoxin-free kit to prepare plasmid DNA. Optimal concentration: Ensure an OD260/280 ratio between 1.7 and 1.9, and dilute DNA to 0.5–2 µg/µL using nuclease-free water.
1.4 Improve Cell Condition
For cell lines, transfect cells with >90% viability, confirmed by trypan blue exclusion. Avoid using cells beyond 15 passages, and allow 2-3 passages for recently thawed cells to stabilize before transfection.
For primary cells, proper activation is crucial for optimal transfection efficiency. For example, human primary T cells generally achieve the best transfection results after stimulation with anti-CD3/CD28 activation beads or antibodies for 2-10 days. Do not remove the activation beads until transfection.
1.5 Use Positive Control
We recommend using a 96-well plate format to optimize transfection conditions for a specific cell type, with EGFP mRNA as the positive control.
Transfected cells may exhibit transient changes in behavior, but typically, viability will be restored by the second day post-transfection.
Suspension transfection may yield higher efficiency in certain adherent cell lines. We recommend testing both methods to determine the optimal condition.
Reagent C is not required for most cell lines but may boost efficiency in hard-to-transfect ones. We suggest testing its impact in untested cell lines, both with and without Reagent C.
For further questions, please contact us at: tech@nanoportalbio.com.