HCA002 binds to both free FITC and FITC conjugated to protein (tested for FITC-BSA).
Fluorescein isothiocyanate (FITC) is widely used to attach a fluorescent label to proteins via the amine group. The isothiocyanate group reacts with amino terminal and primary amines in proteins.
Product Form
A lyophilised monovalent human recombinant Fab selected from the HuCAL®GOLD phage display library. Expressed inE. coliand purified using Streptactin affinity chromatography. The antibody is tagged with a DYKDDDDK-tag and a strepII-tag (NWSHPQFEK) at the C-terminus of the antibody heavy chain.
Activity
Activity was tested by indirect ELISA: FITC-BSA (5 µg/ml) and unrelated control proteins were immobilized on a microtiter plate. Specific binding was monitored by first adding HCA002 (2 µg/ml), then adding a secondary antibody (goat anti-human F(ab’)2 fragment specific, AP conjugate. 1:5000 diluted). A fluorescent signal was created by adding the AP substrate Attophos. The signal on the antigen is at least 5-fold above background, whereas the signal on the control antigens is less than 1.5-fold above background.
Affinity
1. Affinity to FITC conjugated to BSA (FITC-BSA): kD = 7 nM. The monovalent intrinsic affinity of HCA002 was measured by BIAcore on immobilized FITC-BSA.
2. Affinity to soluble FITC: kD = 47nM. The monovalent intrinsic affinity to soluble FITC was determined by solution inhibition essentially as described by Friguet, B.et al.1using immobilised FITC-BSA competing with soluble FITC for binding to HCA002.
Application
This product has been reported to work in the following applications. This information is derived from testing within our laboratories, peer-reviewed publications or personal communications from the originators. Please refer to references indicated for further information. For general protocol recommendations, please visitwww.abdserotec.com/protocols.
Application Name
Yes
No
Not Determined
Suggested Dilution
ELISA
2ug/ml
Where this product has not been tested for use in a particular technique this does not necessarily exclude its use in such procedures. Suggested working dilutions are given as a guide only. It is recommended that the user titrates the product for use in their own system using appropriate negative/positive controls.
Prior to reconstitution store at +4oC.
After reconstitution store at -20oC.
Storage in frost-free freezers is not recommended. Avoid repeated freezing and thawing as this may denature the antibody. Should this product contain a precipitate we recommend microcentrifugation before use.
Shelf Life
6 months from date of reconstitution.
Licensed Use
For in vitro research purposes only, unless otherwise specified in writing by AbD Serotec.
Sold under license of U.S. Patents 6,300,064, 6,696,248, 6,708,484, 6,753,136, European Patent 0,859,841 and corresponding patents.
This antibody was developed byAbD Serotec, a wholly owned subsidiary of Bio-Rad Laboratories Inc., Zeppelinstr. 4, 82178 Puchheim, Germany.
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