Prepare and purify Fab or F(ab')2 fragments from mouse IgG1 antibodies.
Thermo Scientific Pierce Mouse IgG1 Fab and F(ab')2 Preparation Kits use immobilized ficin protease to efficiently digest mouse IgG1 into Fab or F(ab')2 fragments, depending on the chosen reaction conditions.
These regular and micro-scale kits provide include all the reagents and accessories necessary to produce either Fab or F(ab')2 fragments. The antibody digestion reaction with ficin agarose is performed in convenient disposable spin columns that allow efficient removal of the immobilized protease and maximum recovery of the IgG fragments. Also included in the kits are Thermo Scientific NAb Protein A Spin Columns and buffers to efficiently purify the resulting fragments. Protein A binds the Fc fragments and undigested IgG, allowing the pure Fab fragments to be recovered in the flow-through fraction. The kits also include Thermo Scientific Zeba Desalting Spin Columns for preparing the IgG sample quickly without dilution instead of utilizing time-consuming dialysis steps.
Enzyme-free digestion products – Immobilized Ficin (beaded agarose resin) provides for control of the digestion reaction and complete removal of resulting antibody fragments from the proteolytic enzyme
Optimized for mouse IgG1 – the kit procedure is optimized for mouse IgG1; the only effective method for this IgG species and subclass (see Related Products for Fab and F(ab')2 preparation kits that are optimized for other species and subclasses of IgG)
Provides ready-to-use antibody fragments – digestion and final recovery of purified Fab or F(ab')2 fragments occurs in neutral pH sodium phosphate buffer, suitable for storage or immediate use in typical applications
High capacity – kits provide for digestion and purification from as much as 4mg of IgG at a time. Use the standard size kit for fragmenting 0.25 to 4mg IgG, and the micro kit for 25 to 250µg IgG.
Complete – kits include all reagents needed to prepare and purify antibody fragments
Fast – spin format greatly reduces sample processing time
Both regular and micro-scale kits provide include all the reagents and accessories necessary to produce either Fab or F(ab')2 fragments depending on the concentration of cysteine that is used in the digestion reaction. When incubated with whole IgG1 and 1 to 4mM cysteine, ficin produces F(ab')2 fragments. When 25mM cysteine is used, ficin digestion produces Fab fragments.
Digestion scheme for preparing F(ab')2 fragments from mouse IgG1 antibodies. Depending on the concentration of cysteine used in the digestion buffer, primarily F(ab')2 or Fab can be made.
Fab and F(ab')2 fragments generated with the Thermo Scientific Pierce Kits. Analysis of various experimental fractions by non-reducing SDS-PAGE (8-16%). Each well was loaded with 2µL of protein.
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