Fluorescence detection of hyaluronidase

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

We labeled hyaluronan (HA) with two fluorophores, fluorescein amine and rhodamine B amine. These two fluorophores are suitable for a fluorescence (Foerster) resonance energy transfer (FRET) which results in a fluorescein quenching and an enhanced rhodamine emission. Such labeled HA (HA-FRET) is a potential sensor for HA degradation. We studied fluorescence properties of HA-FRET in the absence and presence of hyaluronidase enzyme (HA-ase). The time-resolved fluorescence measurements indicate more than 50% of FRET in the absence of HA-ase. In the presence of HA-ase FRET decreases with time, and relative fluorescence intensities of fluorescein and rhodamine shifts to fluorescein indicating a release of FRET. The kinetics of the digestion process of HA by HA-ase depends on the concentration of the enzyme. We demonstrate that simultaneous measurements of green and red emission of HA-FRET can be used in ratio metric detection of the HA-ase presence and activity. This in turn, can be utilized for the construction of a robust but reliable HA-ase sensing device.

Original languageEnglish
Pages (from-to)473-477
Number of pages5
JournalJournal of Photochemistry and Photobiology B: Biology
Volume104
Issue number3
DOIs
StatePublished - 2 Sep 2011

Fingerprint

Hyaluronoglucosaminidase
Hyaluronic Acid
Fluorescence
energy transfer
fluorescence
Energy Transfer
rhodamine
Fluorescein
enzymes
amines
rhodamine B
Rhodamines
resonance fluorescence
Amines
quenching
Fluorescence Resonance Energy Transfer
degradation
Enzymes
shift
sensors

Keywords

  • FRET
  • Fluorescence
  • Hyaluronan
  • Hyaluronidase
  • Ratio metric sensing

Cite this

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title = "Fluorescence detection of hyaluronidase",
abstract = "We labeled hyaluronan (HA) with two fluorophores, fluorescein amine and rhodamine B amine. These two fluorophores are suitable for a fluorescence (Foerster) resonance energy transfer (FRET) which results in a fluorescein quenching and an enhanced rhodamine emission. Such labeled HA (HA-FRET) is a potential sensor for HA degradation. We studied fluorescence properties of HA-FRET in the absence and presence of hyaluronidase enzyme (HA-ase). The time-resolved fluorescence measurements indicate more than 50{\%} of FRET in the absence of HA-ase. In the presence of HA-ase FRET decreases with time, and relative fluorescence intensities of fluorescein and rhodamine shifts to fluorescein indicating a release of FRET. The kinetics of the digestion process of HA by HA-ase depends on the concentration of the enzyme. We demonstrate that simultaneous measurements of green and red emission of HA-FRET can be used in ratio metric detection of the HA-ase presence and activity. This in turn, can be utilized for the construction of a robust but reliable HA-ase sensing device.",
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Fluorescence detection of hyaluronidase. / Fudala, Rafal; Mummert, Mark E.; Gryczynski, Zygmunt; Gryczynski, Ignacy.

In: Journal of Photochemistry and Photobiology B: Biology, Vol. 104, No. 3, 02.09.2011, p. 473-477.

Research output: Contribution to journalArticle

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