GlyCLICK Biotin

925.001,495.00

GlyCLICK Biotin enables site-specific affinity-tagging of IgG with Biotin

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Description

GlyCLICK Biotin is a kit that contains all reagents needed for site-specific conjugation of IgG using Biotin as an affinity tag. Spin columns with GlycINATOR (EndoS2) enzyme immobilized on agarose beads deglycosylates IgG from several species and subclasses by hydrolyzing any Fc glycoform after the inner most GlcNAc. The GalT enzyme supplied in solution enables incorporation of the UDP-GalNAz regent to the exposed GlcNAc sugar for subsequent click chemistry incorporating Biotin. GlyCLICK Biotin is available in kit formats to facilitate affinity-tagging of 250 ug or 2 mg of IgG.

Detailed Information

Application

Antibody Conjugation, Bioassays, ELISA, Western Blot

Product Group

GlyCLICK

Source/Origin

Cloned from S. pyogenes, Produced in E. coli (His-tagged), Engineered galactosyltransferase (Y289L)

Product Composition

Enzymatic components: Immobilized GlycINATOR (EndoS2) on agarose beads (20% EtOH), beta-1,4-galactosyltransferase (GalT)

Digestion Site

IgG Fc-glycans

Specificity

GlycINATOR hydrolyzes Beta-1,4-linked GlcNAc residues on all IgG Fc-glycoforms for site-specific azide activation using GalT and complete labeling using SPAAC

Substrates

Human IgG1, Human IgG2, Human IgG3, Human IgG4, Cow IgG, Fc-fusion proteins, Goat IgG, Horse IgG, Monkey IgG, Mouse IgG, Rabbit IgG, Rat IgG, Sheep IgG

Reaction Conditions

Deglycosylation (GlycINATOR): 30 min incubation at 37°C, Azide Activation (GalT): O/N incubation at 30°C, Labeling (SPAAC): O/N incubation at 25°C

Digestion Buffers

TBS (pH 7.4)

Buffer Compatibility

GlyCLICK steps are performed in TBS buffer, provided in the kit., For purification of azide activated Ab, TBS or PBS can be used., Sodium azide interferes with the click reaction and must be avoided., Avoid phophate buffers, not compatible with buffer additive (precipitation).

Additives/Co-factors

GalT requires buffer additive for activity., Buffer additive is included in the GlyCLICK kit.

Inhibitors

Denaturated IgG

Unit Definition

Components for antibody conjugation of up to 0.25-2 mg IgG

Label

Biotin

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