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Quantification of oligonucleotide modifications of small unilamellar lipid vesicles

Pfeiffer, Indriati and Höök, Fredrik LU (2006) In Analytical Chemistry 78(21). p.7493-7498
Abstract
We present a new method for quantification of the coupling efficiency between amphiphilic oligonucleotides and suspended small unilamellar lipid vesicles (SUVs). The method employs a supported (phospho) lipid bilayer (SLB)-modified sensor template, which upon exposure to a mixture of SUVs and amphiphilic DNA reacts neither with free SUVs nor with DNA-modified SUVs, but with free DNA only. Using calibration curves obtained by recording the concentration dependence of the initial binding rate of free amphiphilic DNA (in the absence of SUVs), it is demonstrated how concentration determinations of both free and bound DNA in the two-component mixture (amphiphilic DNA and lipid vesicles) can be obtained. The calibration curves and the binding... (More)
We present a new method for quantification of the coupling efficiency between amphiphilic oligonucleotides and suspended small unilamellar lipid vesicles (SUVs). The method employs a supported (phospho) lipid bilayer (SLB)-modified sensor template, which upon exposure to a mixture of SUVs and amphiphilic DNA reacts neither with free SUVs nor with DNA-modified SUVs, but with free DNA only. Using calibration curves obtained by recording the concentration dependence of the initial binding rate of free amphiphilic DNA (in the absence of SUVs), it is demonstrated how concentration determinations of both free and bound DNA in the two-component mixture (amphiphilic DNA and lipid vesicles) can be obtained. The calibration curves and the binding analysis were obtained using a quartz crystal microbalance with dissipation (QCM-D) monitoring. The binding efficiency of DNA coupled to SUVs (theta similar to 50 nm) with two cholesterol moieties revealed that the bivalent coupling is essentially 100% in the range of similar to 1 to similar to 35 oligonucleotides per vesicle, whereas reversible coupling was confirmed in the case of monovalent coupling. Coupling of DNA via two cholesterol moieties was obtained by prehybridization of two single-stranded DNA strands modified with single cholesterol moieties in their 3' and 5' ends, respectively, and the monovalent coupling was obtained using single-stranded DNA. In the latter case, the analysis of the amount of free DNA at different DNA-SUV ratios also allowed for a determination of the maximum number of available binding sites on the SUVs, shown to be in good agreement with data obtained for DNA coupling on planar surfaces. With the only requirement that the SLB-modified sensor template react with one of the components in the two-component mixture only, as verified through fingerprint analysis of frequency, f, and energy dissipation, D, QCM-D measurements, it is emphasized that the method is generic and offers a fast and reliable method for evaluations of biomolecular modifications of any type of colloidal nanoparticles. (Less)
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author
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type
Contribution to journal
publication status
published
subject
in
Analytical Chemistry
volume
78
issue
21
pages
7493 - 7498
publisher
The American Chemical Society (ACS)
external identifiers
  • wos:000241670000032
  • scopus:33750725109
  • pmid:17073417
ISSN
1520-6882
DOI
10.1021/ac061280p
language
English
LU publication?
yes
id
2b5e37de-b2dc-4974-ba84-5442071bc1d8 (old id 384244)
date added to LUP
2016-04-01 11:41:11
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2021-09-22 04:51:44
@article{2b5e37de-b2dc-4974-ba84-5442071bc1d8,
  abstract     = {We present a new method for quantification of the coupling efficiency between amphiphilic oligonucleotides and suspended small unilamellar lipid vesicles (SUVs). The method employs a supported (phospho) lipid bilayer (SLB)-modified sensor template, which upon exposure to a mixture of SUVs and amphiphilic DNA reacts neither with free SUVs nor with DNA-modified SUVs, but with free DNA only. Using calibration curves obtained by recording the concentration dependence of the initial binding rate of free amphiphilic DNA (in the absence of SUVs), it is demonstrated how concentration determinations of both free and bound DNA in the two-component mixture (amphiphilic DNA and lipid vesicles) can be obtained. The calibration curves and the binding analysis were obtained using a quartz crystal microbalance with dissipation (QCM-D) monitoring. The binding efficiency of DNA coupled to SUVs (theta similar to 50 nm) with two cholesterol moieties revealed that the bivalent coupling is essentially 100% in the range of similar to 1 to similar to 35 oligonucleotides per vesicle, whereas reversible coupling was confirmed in the case of monovalent coupling. Coupling of DNA via two cholesterol moieties was obtained by prehybridization of two single-stranded DNA strands modified with single cholesterol moieties in their 3' and 5' ends, respectively, and the monovalent coupling was obtained using single-stranded DNA. In the latter case, the analysis of the amount of free DNA at different DNA-SUV ratios also allowed for a determination of the maximum number of available binding sites on the SUVs, shown to be in good agreement with data obtained for DNA coupling on planar surfaces. With the only requirement that the SLB-modified sensor template react with one of the components in the two-component mixture only, as verified through fingerprint analysis of frequency, f, and energy dissipation, D, QCM-D measurements, it is emphasized that the method is generic and offers a fast and reliable method for evaluations of biomolecular modifications of any type of colloidal nanoparticles.},
  author       = {Pfeiffer, Indriati and Höök, Fredrik},
  issn         = {1520-6882},
  language     = {eng},
  number       = {21},
  pages        = {7493--7498},
  publisher    = {The American Chemical Society (ACS)},
  series       = {Analytical Chemistry},
  title        = {Quantification of oligonucleotide modifications of small unilamellar lipid vesicles},
  url          = {http://dx.doi.org/10.1021/ac061280p},
  doi          = {10.1021/ac061280p},
  volume       = {78},
  year         = {2006},
}