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.

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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

  • Target

    Human IgG1-4, IgG of some classes from monkey, rat, rabbit and sheep

  • Time

    30 min reaction

  • Box

    No need for reducing agents or co-factors

  • Digestion Site

    CPAPELLG / GPSVF (below the hinge)

Product Formats

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.

FabRICATOR (IdeS) is a cysteine protease that digests antibodies at a specific site below the hinge, generating a homogenous pool of F(ab')2 and Fc/2 fragments.

Applications

Performance Activity

Citations

Resources

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FAQ and Support

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.

FabRICATOR is a cysteine protease. It should not result in any other modifications than the digestion of the peptide bond. Digestion will be between the two Glycines (…CPAPELLG / GPSVF…) and the C-terminus will regain the carboxyl group – that is addition of one OH-group and the N-terminal amino group will be regained (addition of one H).

In summary, R1-CO-NH-R2 + H2O –> R1-COOH + NH2-R2.

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.

Yes, for human IgG1 molecules both enzymes can be used in combination to generate a specific hinge peptide. The incubation time is significantly longer for FabALACTICA and it is recommended to first run the FabALACTICA digest and then add FabRICATOR for the last 30 min.

We have tested PBS and 150 mM ammonium acetate, pH7 with good results. More buffers were found to be compatible with FabRICATOR digestion in solution (more info) and should be suitable for FabRICATOR HPLC as well. However, the ionic strength of the buffer influences retention of the antibody fragments on the FabRICATOR HPLC column and a minimum of 100-150 mM salt is necessary for IgG1 subunits to elute as a single, narrow peak. Other IgG subclasses or fusion proteins might need some optimization of buffer composition.

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Legal & Disclaimers

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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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