€1,876.00
FabRICATOR Validation kit enables validation of FabRICATOR-based analytical methods
FabRICATOR Validation kit contains three batches of lyophilized FabRICATOR (IdeS) enzyme. FabRICATOR is an IgG-specific cysteine protease that digests antibodies or Fc-fusions at one single amino acid position below the hinge generating Fc and F(ab’)2 fragments. FabRICATOR Validation kit contains 3 x 2000 U of lyophilized enzyme for digestion of 3 x 2 mg of antibodies or Fc-fusion proteins.
| Application | |
|---|---|
| Product Group | |
| Source/Origin | Cloned from S. pyogenes, Produced in E. coli (His-tagged), 37.7 kDa |
| Product Composition | 3 x 2000 U Lyophilized FabRICATOR in PBS Purified to ≥ 95% homogeneity |
| Digestion Site | |
| Specificity | |
| Substrates | Human IgG1, Human IgG2, Human IgG4, Monkey IgG, Rabbit IgG, Rat IgG, Sheep IgG |
| Reaction Conditions | 30 minute reaction at 37°C according to protocol specifications |
| Digestion Buffers | PBS (pH 6.0 – 8.0), TBS (pH 7.0 – 8.0), MES (pH 5.5 – 6.5), HEPES (pH 7.0 – 8.0), Ammonuim Bicarbonate (pH 6.0 – 7.0), Sodium Acetate (pH 6.0) |
| Buffer Compatibility | |
| Inhibitors | |
| Unit Definition |
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.
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.
FabRICATOR is a very specific enzyme and small changes in the sequence in or around the digestion site (…CPAPELLG / GPSVF…) may reduce or completely impair the ability to digest the IgG. It is very difficult to predict the outcome for each specific modification. It might work, but we cannot guarantee it unfortunately, the only way of knowing is to perform a digestion test. Some suggestions that are worth testing if you experience incomplete digestion: Increase the incubation time, increase the amount of added enzyme, use a PBS with greatly reduced salt concentration (5-10 mM instead of 140 mM) and pH 6.5 instead of 7.4.
