TY - JOUR
T1 - Fluorescent biosensor for the detection of hyaluronidase
T2 - intensity-based ratiometric sensing and fluorescence lifetime-based sensing using a long lifetime azadioxatriangulenium (ADOTA) fluorophore
AU - Chib, Rahul
AU - Mummert, Mark
AU - Bora, Ilkay
AU - Laursen, Bo W.
AU - Shah, Sunil
AU - Pendry, Robert
AU - Gryczynski, Ignacy
AU - Borejdo, Julian
AU - Gryczynski, Zygmunt
AU - Fudala, Rafal
N1 - Publisher Copyright:
© 2016, Springer-Verlag Berlin Heidelberg.
PY - 2016/5/1
Y1 - 2016/5/1
N2 - In this report, we have designed a rapid and sensitive, intensity-based ratiometric sensing as well as lifetime-based sensing probe for the detection of hyaluronidase activity. Hyaluronidase expression is known to be upregulated in various pathological conditions. We have developed a fluorescent probe by heavy labeling of hyaluronic acid with a new orange/red-emitting organic azadioxatriangulenium (ADOTA) fluorophore, which exhibits a long fluorescence lifetime (∼20 ns). The ADOTA fluorophore in water has a peak fluorescence lifetime of ∼20 ns and emission spectra centered at 560 nm. The heavily ADOTA-labeled hyaluronic acid (HA-ADOTA) shows a red shift in the peak emission wavelength (605 nm), a weak fluorescence signal, and a shorter fluorescence lifetime (∼4 ns) due to efficient self-quenching and formation of aggregates. In the presence of hyaluronidase, the brightness and fluorescence lifetime of the sample increase with a blue shift in the peak emission to its original wavelength at 560 nm. The ratio of the fluorescence intensity of the HA-ADOTA probe at 560 and 605 nm can be used as the sensing method for the detection of hyaluronidase. The cleavage of the hyaluronic acid macromolecule reduces the energy migration between ADOTA molecules, as well as the degree of self-quenching and aggregation. This probe can be efficiently used for both intensity-based ratiometric sensing as well as fluorescence lifetime-based sensing of hyaluronidase. The proposed method makes it a rapid and sensitive assay, useful for analyzing levels of hyaluronidase in relevant clinical samples like urine or plasma. [Figure not available: see fulltext.]
AB - In this report, we have designed a rapid and sensitive, intensity-based ratiometric sensing as well as lifetime-based sensing probe for the detection of hyaluronidase activity. Hyaluronidase expression is known to be upregulated in various pathological conditions. We have developed a fluorescent probe by heavy labeling of hyaluronic acid with a new orange/red-emitting organic azadioxatriangulenium (ADOTA) fluorophore, which exhibits a long fluorescence lifetime (∼20 ns). The ADOTA fluorophore in water has a peak fluorescence lifetime of ∼20 ns and emission spectra centered at 560 nm. The heavily ADOTA-labeled hyaluronic acid (HA-ADOTA) shows a red shift in the peak emission wavelength (605 nm), a weak fluorescence signal, and a shorter fluorescence lifetime (∼4 ns) due to efficient self-quenching and formation of aggregates. In the presence of hyaluronidase, the brightness and fluorescence lifetime of the sample increase with a blue shift in the peak emission to its original wavelength at 560 nm. The ratio of the fluorescence intensity of the HA-ADOTA probe at 560 and 605 nm can be used as the sensing method for the detection of hyaluronidase. The cleavage of the hyaluronic acid macromolecule reduces the energy migration between ADOTA molecules, as well as the degree of self-quenching and aggregation. This probe can be efficiently used for both intensity-based ratiometric sensing as well as fluorescence lifetime-based sensing of hyaluronidase. The proposed method makes it a rapid and sensitive assay, useful for analyzing levels of hyaluronidase in relevant clinical samples like urine or plasma. [Figure not available: see fulltext.]
KW - Azadioxatriangulenium fluorophore
KW - Fluorescence lifetime-based sensing
KW - Fluorescence-based assay
KW - Hyaluronidase sensing
KW - Ratiometric sensing
UR - http://www.scopus.com/inward/record.url?scp=84961211458&partnerID=8YFLogxK
U2 - 10.1007/s00216-016-9472-5
DO - 10.1007/s00216-016-9472-5
M3 - Article
C2 - 26993308
AN - SCOPUS:84961211458
SN - 1618-2642
VL - 408
SP - 3811
EP - 3821
JO - Analytical and Bioanalytical Chemistry
JF - Analytical and Bioanalytical Chemistry
IS - 14
ER -