Question 1

Can I re-use FabRICATOR Immobilized columns?

kk · Accepted Answer

We can only guarantee optimal digestion for one-time use. We do not have a cleaning or regeneration protocol to provide. However, depending on the antibody and the following application the column can be reused. We do not recommend using with different antibodies due to the risk for contamination of carry-over from previous sample. If column reuse is desired store the column in 10-20% ethanol at +4-8°C.

Question 2

Can I release Fc fragments from the CaptureSelect column after isolating the F(ab')2 fragments?

kk · Accepted Answer

It is possible to elute the Fc fragments from the CaptureSelect column using a low pH buffer, for example a 0.1M Glycine buffer pH=3.0. The pH must immediately after elution be adjusted to neutral pH by adding 0.1x elution volume of 1M Tris pH=8.0.

Question 3

Can I use FabRICATOR Fab2 Kit for generation and purification of fragments from Rabbit IgG?

Felicia Mejare · Accepted Answer

FabRICATOR Fab2 Kit digests rabbit IgG but to capture the Fc fragments in the affinity step you need another resin than the CaptureSelect&trade; 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.

Question 4

Can I use protein A or MabSelectSure (GE) for removal of Fc-fragments?

kk · Accepted Answer

Genovis has chosen CaptureSelect in the FabRICATOR Fab2 Kit for removal of Fc fragments. CaptureSelect has a very high specific affinity for the Fc. Both protein A and MabSelect Sure (GE) may have affinity for the Fab which may result in a lower recovery of the generated F(ab’)2 fragments and the degree of affinity is antibody dependent. We have many customer reports on this subject, and they usually come back to us for the CaptureSelect columns for the above reason.

Question 5

Does FabRICATOR introduce modifications?

kk · Accepted Answer

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.

Question 6

Where does FabRICATOR cleave the IgG molecule?

kk · Accepted Answer

FabRICATOR® cleave the IgG molecule in one specific site in the lower hinge region of the anitbody leaving a F(ab')2 fragment and a Fc-fragment.
Human IgG1:  ..CPAPELLG / GPSVF..
Human IgG2:  ..CPAPPVA / GPSVF..
Human IgG3:  ..CPAPELLG / GPSVF..
Human IgG4:  ..CPAPEFLG / GPSVF..

FabRICATOR Fab2 Kit Microspin 0.5 mg quantity	FabRICATOR Fab2 Kit Microspin 0.5 mg (A2-FR2-005)	€514.00
FabRICATOR Fab2 Kit Microspin 5 x 0.5 mg quantity	FabRICATOR Fab2 Kit Microspin 5 x 0.5 mg (A2-FR2-025)	€1,367.00
FabRICATOR Fab2 Kit Microspin 10 x 0.5 mg quantity	FabRICATOR Fab2 Kit Microspin 10 x 0.5 mg (A2-FR2-050)	€2,605.00
FabRICATOR Fab2 Kit Midispin 10 mg quantity	FabRICATOR Fab2 Kit Midispin 10 mg (A2-FR2-100)	€1,717.00
FabRICATOR Fab2 Kit Maxispin 100 mg quantity	FabRICATOR Fab2 Kit Maxispin 100 mg (A2-FR2-1000)	€5,118.00

Application	Antibody Characterization, Antibody Fragmentation, Purify F(ab')2 and Fc Fragments
Product Group	FabRICATOR
Source/Origin	Cloned from S. pyogenes, Produced in E. coli (His-tagged), 37.7 kDa
Product Composition	Immobilized on agarose beads (20% EtOH), CaptureSelect Fc columns included
Digestion Site	Below Hinge
Specificity	CPAPELLG / GPSVF (below the hinge)
Substrates	Human IgG1, Human IgG2, Human IgG4, Monkey IgG, Rat IgG, Sheep IgG
Reaction Conditions	30 minute reaction at 37°C according to protocol specifications
Digestion Buffers	PBS (pH 7.4)
Buffer Compatibility	Other commonly used buffers at physiological pH (6.0 – 8.0) and ionic strength can also be used. Optimization may then be required.
Inhibitors	Iodoacetate 20 mM, Iodoacetamide 1 mM, pH < 5.1
Unit Definition	0.1 ml settled resin digests ≥ 95% of 0.5 mg Human IgG1

FabRICATOR Fab2 Kit

Description

Detailed Information

FAQs

Can I re-use FabRICATOR Immobilized columns?

Can I release Fc fragments from the CaptureSelect column after isolating the F(ab’)2 fragments?

Can I use FabRICATOR Fab2 Kit for generation and purification of fragments from Rabbit IgG?

Can I use protein A or MabSelectSure (GE) for removal of Fc-fragments?

Does FabRICATOR introduce modifications?

Where does FabRICATOR cleave the IgG molecule?

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