Quantitative comparison of long-wavelength Alexa Fluor dyes to Cy dyes: fluorescence of the dyes and their bioconjugates.
AuthorsBerlier JE, Rothe A, Buller G, Bradford J, Gray DR, Filanoski BJ, Telford WG, Yue S, Liu J, Cheung CY, Chang W, Hirsch JD, Beechem JM, Haugland RP, Haugland RP
JournalJ Histochem Cytochem
PubMed ID14623938
'Amine-reactive N-hydroxysuccinimidyl esters of Alexa Fluor fluorescent dyes with principal absorption maxima at about 555 nm, 633 nm, 647 nm, 660 nm, 680 nm, 700 nm, and 750 nm were conjugated to antibodies and other selected proteins. These conjugates were compared with spectrally similar protein conjugates of the Cy3, Cy5, ... More
Direct influence of S9 liver homogenate on fluorescence signals: impact on practical applications in a bacterial genotoxicity assay.
AuthorsDreier J, Breitmaier EB, Gocke E, Apfel CM, Page MG
JournalMutat Res
PubMed ID11719102
Assays based on the bacterial SOS-response offer the possibility of automatization of genotoxicity testing for screening of large compound libraries. While existing assays use colorimetric detection or luminescence read-out, we describe here the use of a fluorescence-based system to achieve high sensitivity of detection required for assay miniaturization. Three commonly ... More
beta-Galactosidase activity assay using far-red-shifted fluorescent substrate DDAOG.
AuthorsGong H, Zhang B, Little G, Kovar J, Chen H, Xie W, Schutz-Geschwender A, Olive DM,
JournalAnal Biochem
PubMed ID19103143
beta-Galactosidase (beta-gal) is commonly used as a reporter gene in biological research, and a wide variety of substrates have been developed to assay its activity. One substrate, 9H-(1,3-dichloro-9,9-dimethylacridin-2-one-7-yl) beta-d-galactopyranoside (DDAOG), can be cleaved by beta-gal to produce 7-hydroxy-9H(I,3-dichloro-9,9-dimethylacridin-2-one) (DDAO). On excitation, DDAO generates a far-red-shifted fluorescent signal. Using this substrate, ... More
Activation of membrane-associated procaspase-3 is regulated by Bcl-2.
AuthorsKrebs JF, Armstrong RC, Srinivasan A, Aja T, Wong AM, Aboy A, Sayers R, Pham B, Vu T, Hoang K, Karanewsky DS, Leist C, Schmitz A, Wu JC, Tomaselli KJ, Fritz LC
JournalJ Cell Biol
PubMed ID10085291
The mechanism by which membrane-bound Bcl-2 inhibits the activation of cytoplasmic procaspases is unknown. Here we characterize an intracellular, membrane-associated form of procaspase-3 whose activation is controlled by Bcl-2. Heavy membranes isolated from control cells contained a spontaneously activatable caspase-3 zymogen. In contrast, in Bcl-2 overexpressing cells, although the caspase-3 ... More
Quantification of beta-galactosidase activity after non-viral transfection in vivo.
AuthorsZinselmeyer BH, Beggbie N, Uchegbu IF, Schätzlein AG
JournalJ Control Release
PubMed ID12932652
The limited efficacy of non-viral gene delivery systems currently hampers their wider therapeutic use. In order to further develop novel gene delivery systems, it is important to quantify their efficacy. Many reporter gene assays have limitations when being used to quantify expression in vivo. We have developed a simple assay ... More