reporter enzymes
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Author(s):  
Caitlin E. Sharpes ◽  
John B. McManus ◽  
Steven M. Blum ◽  
Glory E. Mgboji ◽  
Matthew W. Lux
Keyword(s):  


2017 ◽  
Vol 217 (3) ◽  
pp. 1143-1154 ◽  
Author(s):  
Tino Pleiner ◽  
Mark Bates ◽  
Dirk Görlich

Polyclonal anti–immunoglobulin G (anti-IgG) secondary antibodies are essential tools for many molecular biology techniques and diagnostic tests. Their animal-based production is, however, a major ethical problem. Here, we introduce a sustainable alternative, namely nanobodies against all mouse IgG subclasses and rabbit IgG. They can be produced at large scale in Escherichia coli and could thus make secondary antibody production in animals obsolete. Their recombinant nature allows fusion with affinity tags or reporter enzymes as well as efficient maleimide chemistry for fluorophore coupling. We demonstrate their superior performance in Western blotting, in both peroxidase- and fluorophore-linked form. Their site-specific labeling with multiple fluorophores creates bright imaging reagents for confocal and superresolution microscopy with much smaller label displacement than traditional secondary antibodies. They also enable simpler and faster immunostaining protocols, and allow multitarget localization with primary IgGs from the same species and of the same class.



2013 ◽  
Vol 8 (10) ◽  
pp. 2127-2132 ◽  
Author(s):  
Sambashiva Banala ◽  
Stijn J.A. Aper ◽  
Werner Schalk ◽  
Maarten Merkx


2011 ◽  
Vol 93 (1) ◽  
pp. 381-392 ◽  
Author(s):  
Norma Welsch ◽  
Georg Homuth ◽  
Thomas Schweder




1997 ◽  
Vol 204 (1) ◽  
pp. 89-98 ◽  
Author(s):  
H Schäfer ◽  
A Schäfer ◽  
A.F Kiderlen ◽  
K.N Masihi ◽  
R Burger


1996 ◽  
Vol 42 (9) ◽  
pp. 1542-1546 ◽  
Author(s):  
I Bronstein ◽  
C S Martin ◽  
J J Fortin ◽  
C E Olesen ◽  
J C Voyta

Abstract A series of enzyme-activated chemiluminescence-based assays of reporter gene expression, useful in many biomedical applications, has been developed. The chemiluminescence detection systems for beta-galactosidase, beta-glucuronidase (GUS), and secreted placental alkaline phosphatase (SEAP) reporter enzymes are all based on use of 1,2-dioxetane substrates. This detection technology also permits the combined luminescence detection of two different reporter enzymes in the same tube, e.g., a dual assay for beta-galactosidase and luciferase. The sensitivity of these chemiluminescence assays is several orders of magnitude greater than that of conventional colorimetric or fluorometric detection methods; e.g., the detection limit for beta-galactosidase by the chemiluminescence assay is 8 fg and by a fluorometric assay is 2 pg. Furthermore, chemiluminescence enables detection of beta-galactosidase, GUS, and SEAP enzyme concentrations over a dynamic range of more than five to six orders in magnitude. These assays offer highly sensitive, quantitative, rapid, nonisotopic detection of reporter enzymes that are widely used in both mammalian cells and plant cells.



Author(s):  
Hyone-Myong Eun
Keyword(s):  


1994 ◽  
Vol 2 (1) ◽  
pp. 23-27 ◽  
Author(s):  
Karen Pardy




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