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Immobilized molecular beacons: a new strategy using UV-activated poly(methyl methacrylate) surfaces to provide large fluorescence sensitivities for reporting on molecular association events.

Publication ,  Journal Article
Situma, C; Moehring, AJ; Noor, MAF; Soper, SA
Published in: Analytical biochemistry
April 2007

We have designed appropriately prepared solid supports consisting of poly(methyl methacrylate) (PMMA) that provide enhanced performance levels for molecular beacons (MBs) that are used for recognizing and reporting on signature DNA sequences in solution. The attachment of primary amine-containing MBs to the PMMA surface was carried out by UV activating the PMMA to produce surface-confined carboxylate groups, which could then be readily coupled to the MBs using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) chemistry. The fluorescence properties of the MBs covalently attached onto this UV-activated PMMA surface were evaluated and compared with the same MBs immobilized onto glass supports. We observed improved limits of detection for the solution complement to the MBs when immobilized onto PMMA, and this was attributed to both the lower autofluorescence levels exhibited by PMMA at the detection wavelengths used and the improved quenching efficiency of the MBs when in their closed hairpin configuration when strapped to a PMMA surface as opposed to glass. As an example of the utility of the PMMA-based immobilization strategies developed for MBs, we report on the analysis of complementary DNAs specific for fruitless (fru) and Ods-site homeobox (OdsH) genes extracted from Drosophila melanogaster fruit flies. The fru gene functions in the central nervous system, where it is necessary for sex determination and male courtship behavior, whereas the OdsH gene is involved in the regulation of transcription.

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Published In

Analytical biochemistry

DOI

EISSN

1096-0309

ISSN

0003-2697

Publication Date

April 2007

Volume

363

Issue

1

Start / End Page

35 / 45

Related Subject Headings

  • Ultraviolet Rays
  • Transcription Factors
  • Polymethyl Methacrylate
  • Nerve Tissue Proteins
  • Molecular Sequence Data
  • Microfluidics
  • Homeodomain Proteins
  • Fluorescence
  • Drosophila melanogaster
  • Drosophila Proteins
 

Citation

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Situma, C., Moehring, A. J., Noor, M. A. F., & Soper, S. A. (2007). Immobilized molecular beacons: a new strategy using UV-activated poly(methyl methacrylate) surfaces to provide large fluorescence sensitivities for reporting on molecular association events. Analytical Biochemistry, 363(1), 35–45. https://doi.org/10.1016/j.ab.2006.12.029
Situma, Catherine, Amanda J. Moehring, Mohamed A. F. Noor, and Steven A. Soper. “Immobilized molecular beacons: a new strategy using UV-activated poly(methyl methacrylate) surfaces to provide large fluorescence sensitivities for reporting on molecular association events.Analytical Biochemistry 363, no. 1 (April 2007): 35–45. https://doi.org/10.1016/j.ab.2006.12.029.
Situma, Catherine, et al. “Immobilized molecular beacons: a new strategy using UV-activated poly(methyl methacrylate) surfaces to provide large fluorescence sensitivities for reporting on molecular association events.Analytical Biochemistry, vol. 363, no. 1, Apr. 2007, pp. 35–45. Epmc, doi:10.1016/j.ab.2006.12.029.
Journal cover image

Published In

Analytical biochemistry

DOI

EISSN

1096-0309

ISSN

0003-2697

Publication Date

April 2007

Volume

363

Issue

1

Start / End Page

35 / 45

Related Subject Headings

  • Ultraviolet Rays
  • Transcription Factors
  • Polymethyl Methacrylate
  • Nerve Tissue Proteins
  • Molecular Sequence Data
  • Microfluidics
  • Homeodomain Proteins
  • Fluorescence
  • Drosophila melanogaster
  • Drosophila Proteins