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FabRICATOR® – Below Hinge Digestion of IgG (IdeS)
FabRICATOR (IdeS) is an IgG-specific cysteine protease that digests antibodies at a single amino acid site below the hinge, generating homogenous F(ab’)2 and Fc fragments.
SmartEnzymes™
FabRICATOR is widely used in characterization, quality control, stability testing, production monitoring and clone selection of antibody-based therapeutics such as mAbs, ADCs, biosimilars and Fc-fusion proteins.
Why FabRICATOR®?
- Highly specific – one digestion site below the hinge
- Fast – generates a homogenous pool of fragments within 30 minutes
- Enables characterization and quality control of antibody-based biotherapeutics
- Available immobilized in ready-to-use spin columns
Human IgG1-4, IgG of some classes from monkey, rat, rabbit and sheep
30 min reaction
No need for reducing agents or co-factors
CPAPELLG / GPSVF (below the hinge)
Product Formats
FabRICATOR® Immobilized
Immobilized enzyme for below hinge digestion of IgG in spin columns
Buy productFabRICATOR® Fab2 Kit
Immobilized enzyme and affinity resin for below hinge digestion of IgG and purification of fragments
Buy productFabRICATOR® MagIC
Immobilized enzyme on magnetic beads for automated digestion of IgG below the hinge
Buy productFabRICATOR® Low Endotoxin
Lyophilized enzyme with low levels of endotoxin for below hinge digestion of IgG
Buy productFabRICATOR® Validation Kit
Three different lyophilized batches for validation of FabRICATOR-based analytical methods
Buy product
About FabRICATOR®
FabRICATOR (IdeS) is a unique enzyme that digests IgG at a specific site below the hinge region, generating a homogenous pool of F(ab’)2 and Fc/2 fragments.
No other substrate besides IgG is known. The IgG is digested within 30 minutes, and there is no risk of overdigestion if the incubation time is prolonged. Since FabRICATOR digests IgG under physiological reaction conditions, the immunoreactivity is preserved. The best activity is obtained at 37°C, but digestion can also be performed at room temperature using a prolonged reaction time.
FabRICATOR digests all subclasses of human and some classes of monkey, rabbit and sheep IgG. It has limited activity on mouse IgG2a and IgG3 – for digestion of these antibody species, we recommend using FabRICATOR Z.
FabRICATOR is cloned from Streptococcus pyogenes and recombinantly expressed in E. coli. It is produced by Genovis AB for analytical and biotech applications requiring the highest standards and consistency. The enzyme contains a His-tag and has a molecular weight of 38 kDa.
A method to detect the FabRICATOR enzyme is available. Learn more about Anti-FabRICATOR, a goat polyclonal antibody.
Applications
Fast and reliable middle-level analysis of antibody oxidation using FabRICATOR and GlycINATOR, supporting robust QC of critical quality attributes.
Performance Activity
Generation and purification of F(ab’)2 and scFc fragments without residual enzyme, enabling rapid preparation of antibody fragments for downstream applications.
Generation of defined IgG subunits via single-site, below hinge cdigestion, enabling high-resolution middle-level LC-MS and improved PTM characterization.
Citations
Resources
How FabRICATOR® works
Watch video
SmartEnzymes™ for Automated and High-throughput Characterization of Antibodies
Download Scientific Poster
AppNote FabRICATOR® MagIC for Automated Analysis in Complex Samples
Download Application Notes
AppNote SmartEnzymes™ for mAb Oxidation
Download Application Notes
AppNote FabRICATOR® for Glycan Analysis
Download Application Notes
SmartEnzymes™ for Generation of Homogeneous Antibody Fragments
Download Scientific Poster
SmartStories
FabRICATOR® MagIC produce F(ab)2 subunits in less than 30 minutes
Dan Bach Kristensen
Symphogen A/S
FabRICATOR® Supports Analysis of a Complex Antibody-Oligonucleotide Conjugate
Yury Tsybin
Spectroswiss
FabRICATOR® for Intact Analysis of Biologics
Min Kyung So
KBIO Osong Medical Innovation Foundation
FAQ and Support
Can I use FabRICATOR (IdeS) with very low amounts of IgG?
It is possible to use FabRICATOR with low concentrations of IgG. Low concentrations of IgG will lead to decreased reaction rate, so for concentrations below 0.5mg/mL it is therefore recommended to increase the amount of added enzyme and/or increase the incubation time. There is no risk of over-digestion and the incubation time can be prolonged to O/N if necessary.
Does FabRICATOR (IdeS) have protease activity on IgM?
No. According to the following reference, FabRICATOR does not digest IgM. Pavel-Rammingen et al., 2002. IdeS, a novel streptococcal cysteine proteinase with unique specificity for immunoglobulin G.
Which digestion buffers are compatible with FabRICATOR MagIC?
The protocol for FabRICATOR MagIC is based on using standard PBS pH 7.4 as digestion buffer. FabRICATOR in solution is compatible with most commonly used buffers at pH=6-8 and the same buffers should work for FabRICATOR MagIC, but the incubation time might need to be optimized.
Is FabRICATOR stable in the presence of denaturing agents such as guanidine hydrochloride or SDS?
We have tested 4-6 M GuHCl which the enzyme does not withstand without significant decrease of activity. SDS have not been evaluated but 0.1% RapiGest from waters have negative influence on activity. Taken together the enzyme does not well tolerate denaturing agents although we have not tested this in more detailed studies.
Can I use FabRICATOR Fab2 Kit for generation and purification of fragments from Rabbit IgG?
FabRICATOR Fab2 Kit digests rabbit IgG but to capture the Fc fragments in the affinity step you need another resin than the CaptureSelect™ Fc normally included in the kit. Please contact the Genovis team if you are interested in processing rabbit IgG and we will provide columns with resin compatible with rabbit Fc in the FabRICATOR Fab2 Kit.
Legal & Disclaimers
All rights reserved. Genovis products may be covered by one or more patents, trademarks and copyrights owned by Genovis AB or licensed from third parties. For more information about commercial rights, please contact the Genovis team at licensing@genovis.com.
Genovis products are intended for research use only. They are not intended to be used for therapeutic or diagnostic purposes in humans or animals. All goods and services are sold subject to Genovis’ General Terms and Conditions of Sale.
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