Reaktionsgleichung
N[C@H](C(O)=O)CS.C1(=O)NC(=O)C=C1.[OH:15][C:16]([CH2:18][CH2:19][CH2:20][CH2:21][C@H:22]1[C@@H:30]2[C@@H:25]([NH:26][C:27]([NH:29]2)=[O:28])[CH2:24][S:23]1)=[O:17]>>[OH:17][C:16]([CH2:18][CH2:19][CH2:20][CH2:21][C@H:22]1[C@@H:30]2[C@@H:25]([NH:26][C:27]([NH:29]2)=[O:28])[CH2:24][S:23]1)=[O:15]Komponenten
Versuchsvorschrift
The term “thiol reactivity value” is a quantitative characterization of the reactivity of free cysteine amino acids. The thiol reactivity value is the percentage of a free cysteine amino acid in a cysteine engineered antibody which reacts with a thiol-reactive reagent, and converted to a maximum value of 1. For example, a free cysteine amino acid on a cysteine engineered antibody which reacts in 100% yield with a thiol-reactive reagent, such as a biotin-maleimide reagent, to form a biotin-labelled antibody has a thiol reactivity value of 1.0. Another cysteine amino acid engineered into the same or different parent antibody which reacts in 80% yield with a thiol-reactive reagent has a thiol reactivity value of 0.8. Another cysteine amino acid engineered into the same or different parent antibody which fails totally to react with a thiol-reactive reagent has a thiol reactivity value of 0. Determination of the thiol reactivity value of a particular cysteine may be conducted by ELISA assay, mass spectroscopy, liquid chromatography, autoradiography, or other quantitative analytical tests. Thiol-reactive reagents which allow capture of the cysteine engineered antibody and comparison and quantitation of the cysteine reactivity include biotin-PEO-maleimide ((+)-biotinyl-3-maleimidopropionamidyl-3,6-dioxaoctainediamine, Oda et al (2001) Nature Biotechnology 19:379-382, Pierce Biotechnology, Inc.) Biotin-BMCC, PEO-Iodoacetyl Biotin, Iodoacetyl-LC-Biotin, and Biotin-HPDP (Pierce Biotechnology, Inc.), and Nα-(3-maleimidylpropionyl)biocytin (MPB, Molecular Probes, Eugene, Oreg.). Other commercial sources for biotinylation, bifunctional and multifunctional linker reagents include Molecular Probes, Eugene, Oreg., and Sigma, St. Louis, Mo.
Quellen und Nachweise
- ORD-Eintrag
- ord-6cd038edc8b74fefb06a74826f89bb35
- Quelldatensatz
- uspto-grants-2013_09
- Patentnummer
- US08524865B2
Verknüpfte Notizen
Wird geladen…