Multilayer mediated forward and patterned siRNA transfection using linear-PEI at extended N/P ratios

Sumit Mehrotra, Ilsoon Lee, Christina Chan

Research output: Contribution to journalArticle

  • 40 Citations

Abstract

Gene delivery from a substrate depends, in part, on the vector-nucleic acid complex that is bound to the surface and the cell adhesive properties of the surface. Here, we present a method to deliver patterns of small interfering RNA (siRNA) that capitalize on a forward transfection method (transfection by introducing siRNA transfection reagent complexes onto plated cells); herein denoted as multilayer mediated forward transfection (MFT). This method separates the substrate-mediated delivery from the cell adhesive properties of the surface. pH responsive layer-by-layer (LbL) assembled multilayers were used as the delivery platform and microcontact printing technique (μCP) was used to pattern nanoparticles of transfection reagent-siRNA complexes onto degradable multilayers. Efficient MFT depend on optimal formulation of the nanoparticles. 25 kDa linear polyethylenimine (LPEI) was optimized as the siRNA transfection reagent for normal forward transfection (NFT) of the nanoparticles. A broad range of LPEI-siRNA nitrogen/phosphate (N/P) ratios (ranging from 5 to 90) was evaluated for the relative amounts of siRNA incorporated into the nanoparticles, nanoparticle size and NFT efficiencies. All the siRNA was incorporated into the nanoparticles at N/P ratio near 90. Increasing the amount of siRNA incorporated into the nanoparticles, with increasing N/P ratio correlated with a linear blue shift in the ultraviolet/visible (UV/vis) absorbance spectrum of the LPEI-siRNA nanoparticles. NFT efficiency greater than 80% was achieved with minimal cytotoxicity at N/P ratio of 30 and siRNA concentration of 200 nM. Similarly, MFT efficiency ≥60% was achieved for LPEI-siRNA nanoparticles at N/P ratios greater than 30.

LanguageEnglish (US)
Pages1474-1488
Number of pages15
JournalActa Biomaterialia
Volume5
Issue number5
DOIs
StatePublished - Jun 2009

Profile

Polyetherimides
RNA
Small Interfering RNA
Transfection
Multilayers
Nanoparticles
Polyethyleneimine
Surface Properties
Adhesives
Printing
Nucleic acids
Substrates
Cytotoxicity
Nucleic Acids
Phosphates
Nitrogen
Genes

Keywords

  • Layer-by-layer (LbL) assembly
  • LPEI-siRNA nanoparticles
  • N/P ratio
  • Patterned transfection
  • siRNA transfection

ASJC Scopus subject areas

  • Biomaterials
  • Biomedical Engineering
  • Biotechnology
  • Biochemistry
  • Molecular Biology

Cite this

Multilayer mediated forward and patterned siRNA transfection using linear-PEI at extended N/P ratios. / Mehrotra, Sumit; Lee, Ilsoon; Chan, Christina.

In: Acta Biomaterialia, Vol. 5, No. 5, 06.2009, p. 1474-1488.

Research output: Contribution to journalArticle

@article{9cc375cbd5dd48fea31c7206eaa5f78e,
title = "Multilayer mediated forward and patterned siRNA transfection using linear-PEI at extended N/P ratios",
abstract = "Gene delivery from a substrate depends, in part, on the vector-nucleic acid complex that is bound to the surface and the cell adhesive properties of the surface. Here, we present a method to deliver patterns of small interfering RNA (siRNA) that capitalize on a forward transfection method (transfection by introducing siRNA transfection reagent complexes onto plated cells); herein denoted as multilayer mediated forward transfection (MFT). This method separates the substrate-mediated delivery from the cell adhesive properties of the surface. pH responsive layer-by-layer (LbL) assembled multilayers were used as the delivery platform and microcontact printing technique (μCP) was used to pattern nanoparticles of transfection reagent-siRNA complexes onto degradable multilayers. Efficient MFT depend on optimal formulation of the nanoparticles. 25 kDa linear polyethylenimine (LPEI) was optimized as the siRNA transfection reagent for normal forward transfection (NFT) of the nanoparticles. A broad range of LPEI-siRNA nitrogen/phosphate (N/P) ratios (ranging from 5 to 90) was evaluated for the relative amounts of siRNA incorporated into the nanoparticles, nanoparticle size and NFT efficiencies. All the siRNA was incorporated into the nanoparticles at N/P ratio near 90. Increasing the amount of siRNA incorporated into the nanoparticles, with increasing N/P ratio correlated with a linear blue shift in the ultraviolet/visible (UV/vis) absorbance spectrum of the LPEI-siRNA nanoparticles. NFT efficiency greater than 80{\%} was achieved with minimal cytotoxicity at N/P ratio of 30 and siRNA concentration of 200 nM. Similarly, MFT efficiency ≥60{\%} was achieved for LPEI-siRNA nanoparticles at N/P ratios greater than 30.",
keywords = "Layer-by-layer (LbL) assembly, LPEI-siRNA nanoparticles, N/P ratio, Patterned transfection, siRNA transfection",
author = "Sumit Mehrotra and Ilsoon Lee and Christina Chan",
year = "2009",
month = "6",
doi = "10.1016/j.actbio.2009.01.004",
language = "English (US)",
volume = "5",
pages = "1474--1488",
journal = "Acta Biomaterialia",
issn = "1742-7061",
publisher = "Elsevier BV",
number = "5",

}

TY - JOUR

T1 - Multilayer mediated forward and patterned siRNA transfection using linear-PEI at extended N/P ratios

AU - Mehrotra,Sumit

AU - Lee,Ilsoon

AU - Chan,Christina

PY - 2009/6

Y1 - 2009/6

N2 - Gene delivery from a substrate depends, in part, on the vector-nucleic acid complex that is bound to the surface and the cell adhesive properties of the surface. Here, we present a method to deliver patterns of small interfering RNA (siRNA) that capitalize on a forward transfection method (transfection by introducing siRNA transfection reagent complexes onto plated cells); herein denoted as multilayer mediated forward transfection (MFT). This method separates the substrate-mediated delivery from the cell adhesive properties of the surface. pH responsive layer-by-layer (LbL) assembled multilayers were used as the delivery platform and microcontact printing technique (μCP) was used to pattern nanoparticles of transfection reagent-siRNA complexes onto degradable multilayers. Efficient MFT depend on optimal formulation of the nanoparticles. 25 kDa linear polyethylenimine (LPEI) was optimized as the siRNA transfection reagent for normal forward transfection (NFT) of the nanoparticles. A broad range of LPEI-siRNA nitrogen/phosphate (N/P) ratios (ranging from 5 to 90) was evaluated for the relative amounts of siRNA incorporated into the nanoparticles, nanoparticle size and NFT efficiencies. All the siRNA was incorporated into the nanoparticles at N/P ratio near 90. Increasing the amount of siRNA incorporated into the nanoparticles, with increasing N/P ratio correlated with a linear blue shift in the ultraviolet/visible (UV/vis) absorbance spectrum of the LPEI-siRNA nanoparticles. NFT efficiency greater than 80% was achieved with minimal cytotoxicity at N/P ratio of 30 and siRNA concentration of 200 nM. Similarly, MFT efficiency ≥60% was achieved for LPEI-siRNA nanoparticles at N/P ratios greater than 30.

AB - Gene delivery from a substrate depends, in part, on the vector-nucleic acid complex that is bound to the surface and the cell adhesive properties of the surface. Here, we present a method to deliver patterns of small interfering RNA (siRNA) that capitalize on a forward transfection method (transfection by introducing siRNA transfection reagent complexes onto plated cells); herein denoted as multilayer mediated forward transfection (MFT). This method separates the substrate-mediated delivery from the cell adhesive properties of the surface. pH responsive layer-by-layer (LbL) assembled multilayers were used as the delivery platform and microcontact printing technique (μCP) was used to pattern nanoparticles of transfection reagent-siRNA complexes onto degradable multilayers. Efficient MFT depend on optimal formulation of the nanoparticles. 25 kDa linear polyethylenimine (LPEI) was optimized as the siRNA transfection reagent for normal forward transfection (NFT) of the nanoparticles. A broad range of LPEI-siRNA nitrogen/phosphate (N/P) ratios (ranging from 5 to 90) was evaluated for the relative amounts of siRNA incorporated into the nanoparticles, nanoparticle size and NFT efficiencies. All the siRNA was incorporated into the nanoparticles at N/P ratio near 90. Increasing the amount of siRNA incorporated into the nanoparticles, with increasing N/P ratio correlated with a linear blue shift in the ultraviolet/visible (UV/vis) absorbance spectrum of the LPEI-siRNA nanoparticles. NFT efficiency greater than 80% was achieved with minimal cytotoxicity at N/P ratio of 30 and siRNA concentration of 200 nM. Similarly, MFT efficiency ≥60% was achieved for LPEI-siRNA nanoparticles at N/P ratios greater than 30.

KW - Layer-by-layer (LbL) assembly

KW - LPEI-siRNA nanoparticles

KW - N/P ratio

KW - Patterned transfection

KW - siRNA transfection

UR - http://www.scopus.com/inward/record.url?scp=67349175996&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=67349175996&partnerID=8YFLogxK

U2 - 10.1016/j.actbio.2009.01.004

DO - 10.1016/j.actbio.2009.01.004

M3 - Article

VL - 5

SP - 1474

EP - 1488

JO - Acta Biomaterialia

T2 - Acta Biomaterialia

JF - Acta Biomaterialia

SN - 1742-7061

IS - 5

ER -