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Polymer-based delivery

Polymer-based delivery

Cationic polymers condense nucleic acids into polyplexes through electrostatic interaction with the phosphate backbone. They are cheap, stable, easy to scale and often serum-tolerant — which makes them the workhorse of bioproduction and the entry point for therapeutic non-viral delivery.

7
Candidates profiled with manufacturer protocols
5
Manufacturers whose documentation was used
12
References listed at the end of this page
How it works

From the tube to the target

  1. 1

    Condensation

    Branched or linear polycations wrap DNA into compact 50–200 nm polyplexes, protecting it from nucleases and creating a high local charge density.

  2. 2

    Proton sponge effect

    Polyethylenimine's buffering amine groups resist endosomal acidification, drawing in chloride and water until the endosome ruptures. The mechanism is debated but empirically robust — and it depends on molecular weight.

  3. 3

    Molecular weight trade-off

    High-molecular-weight PEI (25 kDa) transfects efficiently but is cytotoxic; 10 kDa and biodegradable variants trade a little efficiency for much better viability.

  4. 4

    Cargo flexibility

    Polyplexes carry DNA, siRNA, mRNA and Cas9 RNP. Increasingly they are the core of lipid–polymer hybrid particles that add a fusogenic lipid shell for better escape.

FamilyRepresentative productsFormat bias
Linear and branched PEIPEI 25 kDa, PEIpro, jetPEILarge-scale and bioproduction, DNA
Biodegradable polycationsTurboFect, DreamFect, PBAEDifficult lines with a viability constraint
Multi-component polymer blendsjetPRIME, jetOPTIMUS, FuGENEDNA plus siRNA co-delivery, reproducibility
Peptide and protein polymersProtamine, histidine-rich peptides, RNP carriersRNP and mRNA in sensitive cells
7 candidates

Reagents and their manufacturer protocols

Each entry below gives the chemistry, the applications the manufacturer validates it for, the dosing basis and the step-by-step protocol. Scale factors follow the manufacturer’s own tables wherever they publish them.

Candidate 01

PEI 25 kDa (branched)

Sigma-Aldrich / Polysciences — widely prepared in-house

408727
Chemistry
Branched polyethylenimine, 25 kDa, proton-sponge polycation
Best for
Cost-sensitive high-volume plasmid work and lentiviral/AAV vector production in HEK293T; the historical benchmark polymer

Dosing basis: Nitrogen-to-phosphate (N/P) ratio 6–10 for DNA; 3 µg PEI per µg DNA is a common in-house starting point.

Manufacturer protocol — step by step

  1. 1

    Prepare plasmid DNA at ≥1 µg/µL in water or 150 mM NaCl, free of endotoxin.

  2. 2

    Dilute DNA in serum-free DMEM to 1/10 of the final culture volume.

  3. 3

    Add PEI at the chosen N/P ratio, vortex for 10 s immediately.

  4. 4

    Incubate 15–20 min at room temperature to allow polyplex formation.

  5. 5

    Add drop-wise to cells in serum-free medium; replace with complete medium after 4–6 h.

  6. 6

    For vector production, harvest supernatant at 48–72 h and clarify before titration.

Protocol notes

  • Branched 25 kDa PEI is effective but strongly cytotoxic at excess; always titrate N/P before scaling.
  • Linear PEI (jetPEI) generally shows a better toxicity profile at comparable efficiency and is preferred for process work.
  • Endotoxin-contaminated plasmid DNA is the most common cause of PEI-associated death in production cultures.

Source: Sigma-Aldrich — Polyethylenimine (branched, 25 kDa) product information

Candidate 02

TurboFect

Thermo Scientific / Fermentas

R0531
Chemistry
Cationic polymer transfection reagent, serum-compatible
Best for
Plasmid DNA transfection in HEK293, HeLa, COS-7 and CHO with excellent reproducibility; also used for larger-scale protein expression

Dosing basis: Fixed 2:1 ratio (µL reagent per µg DNA) across formats — the vendor's protocol deliberately avoids optimisation titrations.

Manufacturer protocol — step by step

  1. 1

    Dilute 1 µg plasmid DNA in 100 µL serum-free DMEM per 24-well.

  2. 2

    Add 2 µL TurboFect and mix immediately by vortexing briefly or pipetting.

  3. 3

    Incubate 15–20 min at room temperature.

  4. 4

    Add the complex drop-wise to cells — do not remove or replace the culture medium.

  5. 5

    Rock the plate gently to distribute evenly; incubate at 37 °C.

  6. 6

    Analyse expression at 24–48 h. For stable lines, replace medium after 24 h and add selective medium for 10–15 days.

Culture vesselDNASerum-free diluentTurboFect
24-well1 µg100 µL2 µL
6-well4 µg400 µL8 µL
60 mm dish6 µg600 µL12 µL
100 mm dish12 µg1.2 mL24 µL

Protocol notes

  • Quality-control reference from the manufacturer: 0.5 µg eGFP plasmid with 2 µL TurboFect delivered to 5 × 10⁴ HeLa cells gives approximately 90 ± 10% eGFP-positive cells by flow cytometry.
  • Antibiotics in the medium do not interfere, and serum is fully compatible during both complexation and exposure.
  • Use DNA with A260/A280 ≥ 1.8; endotoxin-contaminated preparations cause marked toxicity.
  • The 2:1 ratio is fixed by design — if efficiency is low, increase DNA rather than reagent.

Source: Thermo Scientific — TurboFect transfection reagent user guide (MAN0013147)

Candidate 03

jetPRIME

Polyplus-transfection (Sartorius)

114-15
Chemistry
Proprietary polymer in a ready-to-use buffer with a DNA-condensation additive
Best for
DNA and siRNA transfection with a single protocol; the buffered format makes it markedly more reproducible than in-house PEI

Dosing basis: 2 µL jetPRIME reagent per µg DNA, with 4 µL jetPRIME buffer per µg DNA per 24-well.

Manufacturer protocol — step by step

  1. 1

    Dilute 500 ng DNA in 50 µL jetPRIME buffer per 24-well.

  2. 2

    Vortex 10–15 s, then add 1 µL jetPRIME reagent and vortex again for 10 s.

  3. 3

    Incubate 10 min at room temperature.

  4. 4

    Add 50 µL complex to cells in 500 µL complete medium; serum and antibiotics are compatible.

  5. 5

    For siRNA, use the same scheme with 20 pmol siRNA and 1 µL reagent; no buffer step is needed.

  6. 6

    Analyse expression or knockdown at 24–48 h.

Protocol notes

  • The buffer pre-condenses DNA and is the reason jetPRIME tolerates slightly impure preparations better than raw PEI.
  • For DNA plus siRNA co-transfection, combine the two in one tube before adding the reagent.

Source: Polyplus — jetPRIME transfection reagent protocol

Candidate 04

jetOPTIMUS

Polyplus-transfection (Sartorius)

117-15
Chemistry
Next-generation polymer designed for low-dose, low-toxicity delivery
Best for
Adherent cells where the goal is high efficiency at a fraction of the usual reagent dose — including many hard-to-transfect lines

Dosing basis: Effectively half the dose of earlier polymers: 0.5–1 µL reagent per µg DNA.

Manufacturer protocol — step by step

  1. 1

    Dilute 1 µg DNA in 100 µL jetOPTIMUS buffer per 24-well.

  2. 2

    Add 1 µL jetOPTIMUS reagent and vortex 10 s.

  3. 3

    Incubate 10 min at room temperature.

  4. 4

    Add drop-wise to cells; a medium change 24 h later improves long-term viability.

  5. 5

    Assay at 24–48 h.

Protocol notes

  • Vendor-reported performance is 1.5–3× the efficiency of earlier polymer reagents in HEK293 and HeLa at the same DNA dose.
  • Best suited to adherent formats; suspension cells still favour dedicated electroporation kits.

Source: Polyplus — jetOPTIMUS transfection reagent protocol

Candidate 05

FuGENE 6 / FuGENE HD (polymer components)

Promega

E2691 (FuGENE 6)
Chemistry
Non-liposomal multi-component blend with polymer character
Best for
Difficult-to-transfect and serum-sensitive cells; FuGENE 6 remains widely valid for CHO and primary cultures

Dosing basis: 3:1 reagent-to-DNA for FuGENE 6; range 2:1 to 6:1 depending on cell line.

Manufacturer protocol — step by step

  1. 1

    Dilute DNA in 100 µL serum-free medium per 24-well.

  2. 2

    Add 3 µL FuGENE 6 directly into the medium; do not allow contact with the tube wall.

  3. 3

    Mix and incubate 10–15 min at room temperature.

  4. 4

    Add drop-wise to cells; serum may be present.

  5. 5

    Assay at 24–48 h.

Protocol notes

  • Although marketed as non-liposomal, the chemistry is a blend that behaves more like a polymer in serum — useful when lipoplex approaches fail.
  • FuGENE HD has largely superseded FuGENE 6 for most lines, but 6 remains in the methods sections of many legacy protocols.

Source: Promega — FuGENE 6 tranfection reagent protocol

Candidate 06

DreamFect Gold

OZ Biosciences

DF41000
Chemistry
Biodegradable cationic polymer
Best for
Primary cells, stem cells and cultures where long-term viability matters more than peak day-2 expression

Dosing basis: 2 µL reagent per µg DNA as an initial ratio; the biodegradable backbone tolerates a wider window than PEI.

Manufacturer protocol — step by step

  1. 1

    Dilute DNA in 100 µL serum-free medium per 24-well.

  2. 2

    Add 2 µL DreamFect Gold, mix and incubate 15–20 min at room temperature.

  3. 3

    Add drop-wise to cells in complete medium; serum is compatible.

  4. 4

    Assay at 24–72 h; medium may be left unchanged for long-term cultures.

Protocol notes

  • The biodegradable backbone reduces lysosomal accumulation, which is the main driver of late toxicity with PEI.

Source: OZ Biosciences — DreamFect Gold transfection reagent protocol

Candidate 07

PEIpro

Polyplus-transfection (Sartorius)

115-100
Chemistry
Linear PEI manufactured under cGMP conditions
Best for
Large-scale and clinical-grade vector production in HEK293 or HEK293T suspension cultures

Dosing basis: Scale per litre of culture, typically 1 mg DNA and 2–4 mg PEIpro per litre.

Manufacturer protocol — step by step

  1. 1

    Dilute DNA in the vendor's complexation buffer at the process-defined concentration.

  2. 2

    Add PEIpro at the optimised DNA:PEI ratio and mix immediately.

  3. 3

    Incubate 15–20 min at room temperature to form polyplexes.

  4. 4

    Add to the production culture; no medium change is required at this scale.

  5. 5

    Harvest vector at 48–72 h depending on the vector system.

Protocol notes

  • Provides a documented regulatory chain of custody — the usual reason to choose it over in-house PEI.
  • Process optimisation targets peak titre, not peak reporter expression; the ratio that maximises both can differ.

Source: Polyplus — PEIpro transfection reagent for bioproduction

Cross-cutting rules

What applies to every candidate on this page

  • Polyplexes are mechanically robust compared with lipoplexes, but N/P ratio and mixing order must still be fixed by SOP.
  • Serum compatibility varies by product — TurboFect and jetPRIME tolerate serum during exposure, raw PEI does not.
  • Higher polymer dose buys efficiency up to a cliff; sample the cliff deliberately during optimisation and stay below it.

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