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Mouse IgG1 Fab and F(ab’)2 Preparation Kits


Prepare and purify Fab or F(ab')2 fragments from mouse IgG1 antibodies.

Mouse IgG1 Fab and Fab2 Preparation Kits

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.

Highlights:

  • 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

Product Details:

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.

Fab or F(ab')2 antibody fragment preparation with immobilized ficin
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.

 

Efficient IgG fragmentation and production of Fab fragments with Thermo Scientific Pierce Fab Preparation Kit # 44985
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.

References:

  1. Beale, D. (1987). Molecular fragmentations. Some applications in immunology. Dev Comp Immunol 11:287-96.
  2. Boguslawski, S.J., et al. (1989). Improved procedure for preparation of F(ab')2 fragments of mouse IgGs by papain digestion. J. Immunol. Meth. 120:51-56.
  3. Kerbel, R.S. and Elliot, B.E. (1983). Detection of Fc receptors. Method. Enzymol. 93:113-147.
  4. Kulkami, P.N., et al. (1985). Conjugation of methotrexate to IgG antibodies and their F(ab')2 fragments and the effect of conjugated methotrexate on tumor growth in vivo. Cancer Immunol. Immunother. 19:211-14.
  5. Mariani, M. et al. (1991). A new enzymatic method to obtain high-yield F(ab')2 suitable for clinical use from mouse IgG1. Mol. Immunol. 28:69-77.
  6. Milenic, D.E., et al. (1989). Comparison of methods for the generation of immunoreactive fragments of a monoclonal antibody (B72.3) reactive with human carcinomas. J. Immunol. Meth. 120:71-83.
  7. Nisonoff, A., et al. (1960). Properties of the major component of a peptic digest of rabbit antibody. Science 132:1770.
  8. Parham, P.L., et al. (1982). Monoclonal antibodies: Purification, fragmentation and application to structural and functional studies of class I MHC antigens. J. Immunol. Meth. 53:133-73.
  9. Parham, P. L. (1983). On the fragmentation of monoclonal IgG1, IgG2a, and IgG2b from BALB/c mice. J. Immunol. 131:2895-2902.
  10. Raychaudhuri, G., et al. (1985). Human IgG1 and its Fc fragment bind with different affinities to the Fc receptors on the human U937, HL-60, and ML-1 cell lines. Mol. Immunol. 22(9):1009-19.
  11. Wilson, K.M., et al. (1991). A new enzymatic method to obtain high-yield F(ab')2 suitable for clinical use from mouse IgG1. Mol. Immunol. 28:69-77.

Related Resources:

Review of antibody fragmentation
Review of antibody structure and classes of immunoglobulins
Tech Tip #59: Choosing a secondary antibody: a guide to fragment specificity

Related Products:

Fab Preparation Kit – for other species and subclasses besides mouse IgG1
F(ab')2 Preparation Kit – for other species and subclasses besides mouse IgG1


Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
44680 Mouse IgG1 Fab and F(ab')2 Micro Preparation Kit
Formulation: Multi-component kit
Sufficient For: 10 antibody samples, each containing 25 to 250µg IgG

Kit Contents:
Pierce Ficin Agarose, 0.8mL
Cysteine-HCl, 1g
Mouse IgG1 Digestion Buffer, 120mL
NAb Protein A Spin Columns, 0.2mL, 2 columns
Protein A Binding Buffer, 50mL
IgG Elution Buffer, 50mL
Zeba Spin Desalting Columns, 7K MWCO, 0.5mL, 10 columns
Spin Columns, 0.8mL, 10 columns
Microcentrifuge Tubes, 2mL, 30 tubes
0.8mL-resin kit Product Instructions for product #44680 Mouse IgG1 Fab and F(ab')2 Micro Preparation Kit MSDS for product #44680 Mouse IgG1 Fab and F(ab')2 Micro Preparation Kit Certificate of Analysis for product #44680 Mouse IgG1 Fab and F(ab')2 Micro Preparation Kit $431.00

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44980 Mouse IgG1 Fab and F(ab')2 Preparation Kit
Formulation: Multi-component kit
Sufficient For: 10 antibody samples, each containing 0.25 to 4mg IgG

Kit Contents:
Pierce Ficin Agarose, 2.5mL
Cysteine-HCl, 1g
Mouse IgG1 Digestion Buffer, 120mL
NAb Protein A Spin Column, 1mL, 1 column
Protein A Binding Buffer, 120mL
IgG Elution Buffer, 120mL
Zeba Desalt Spin Columns, 7K MWCO, 2mL, 10 columns
Spin Columns, 0.8mL, 10 columns
Microcentrifuge Tubes, 2mL, 30 tubes
2.5mL-resin kit Product Instructions for product #44980 Mouse IgG1 Fab and F(ab')2 Preparation Kit MSDS for product #44980 Mouse IgG1 Fab and F(ab')2 Preparation Kit Certificate of Analysis for product #44980 Mouse IgG1 Fab and F(ab')2 Preparation Kit $558.00

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Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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