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Fab Preparation Kits


Make antibody Fab fragments from whole IgG.

Pierce Fab Preparation Kits

Thermo Scientific Pierce Fab Preparation Kits use immobilized papain protease to digest human or mouse IgG antibodies to make separate Fab and Fc fragments and subsequently to purify the Fab using Protein A agarose.

These Fab Preparation Kits are suitable for human, rabbit, mouse and other species and subclasses of IgG. The papain antibody digestion reaction 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 Papain (beaded agarose resin) provides for control of the digestion reaction and complete removal of resulting antibody fragments from the proteolytic enzyme
  • Suitable for human and other species of IgG – the kit procedure is optimized for human, mouse and rabbit IgG. Papain-based digestion is effective for certain other species and subclasses of IgG (e.g., goat and pig), although the purification step requires that the antibody effectively binds to Protein A. (Note: for best results with mouse IgG1, use Part No. 44980.)
  • High capacity – kit provides 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.
  • Provides ready-to-use Fab – digestion and final recovery of purified Fab fragments occurs in neutral pH sodium phosphate buffer, suitable for storage or immediate use in typical applications
  • Complete – kits include all reagents needed to prepare and purify antibody fragments
  • Fast – spin format greatly reduces sample processing time
  • Flexible – Protocols are included for multiple species and IgG subclasses, as well as sample size and concentration.
  • Efficient – enhanced yield and sample purity

Product Details:

The kits use papain, a nonspecific thiol-endopeptidase, to enzymatically cleave whole IgG just above the hinge region to create two separate Fab fragments and one Fc fragment per antibody molecule. Because the papain protease is supplied in immobilized form as beaded agarose resin, the digestion reaction is easily stopped by removing the resin from the IgG solution; the result is digest products that are enzyme-free.

Fab antibody fragment preparation with immobilized papain
Digestion and purification scheme for preparing Fab fragments from IgG antibodies.

 

Efficient IgG fragmentation and production of Fab fragments with Thermo Scientific Pierce Fab Preparation Kit
Typical SDS-PAGE (10% Bis-Tris) result of reduced and non-reduced Fab and Fc fragments from rabbit IgG.

 

Recommended digestion times for various species and concentrations of IgG.
  Digestion Time (hours)
[IgG] (mg/mL) Rabbit IgG Human IgG Mouse IgG
8 8 to 9 5 to 6 4 to 5
4 6 to 7 5 to 6 3 to 4
1.5 4 to 5 3 to 4 2 to 3
0.5 3 to 4 2 to 3 2 to 3

References:

  1. Lindner I. et al. (2010) {alpha}2-Macroglobulin inhibits the malignant properties of astrocytoma cells by impeding {beta}-catenin signaling. Cancer Res. 70, 277-87.
  2. Kaufmann B. et al. (2010) Neutralization of West Nile virus by cross-linking of its surface proteins with Fab fragments of the human monoclonal antibody CR4354. PNAS. 107, 18950-5.
  3. Chen X. et al. (2010) Requirement of open headpiece conformation for activation of leukocyte integrin {alpha}x{beta}2. PNAS. 107, 14727-32.
  4. Uysal H. et al. (2009) Structure and pathogenicity of antibodies specific for citrullinated collagen type II in experimental arthitis. J. Exp. Med. 206, 449-62.
  5. Thomas G. M. et al. (2009) Cancer cell-derived microparticles bearing P-selectin glycoprotein ligand 1 accelerate thrombus formation in vivo. J. Exp. Med. 206, 1913-27.
  6. Kong F. et al. (2009) Demonstration of catch bonds between an integrin and its ligand. J. Cell Biol. 185, 1275-84.

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:

F(ab')2 Preparation Kit
Mouse IgG1 Fab and F(ab')2 Preparation Kit


Ordering Information
 
Product # Description Pkg. Size Instructions MSDS CofA Price
44685 Pierce Fab Micro Preparation Kit
Formulation: Multi-component kit
Sufficient For: 10 antibody samples, each containing 25 to 250µg IgG

Kit Contents:
Pierce Papain Agarose, 0.5mL
Cysteine-HCl, 1g
Fab Digestion Buffer, 55mL
NAb Protein A Spin Columns, 0.2mL, 2 columns
PBS Packs (each makes 500mL), 2 packs
IgG Elution Buffer, 50mL
Zeba Desalt Spin Columns, 7K MWCO, 0.5mL, 10 columns
Spin Columns, 0.8mL, 10 columns
Microcentrifuge Tubes, 2mL, 30 tubes
0.5mL-resin kit Product Instructions for product #44685 Pierce Fab Micro Preparation Kit MSDS for product #44685 Pierce Fab Micro Preparation Kit Certificate of Analysis for product #44685 Pierce Fab Micro Preparation Kit $372.00

Add to cart
44985 Pierce Fab Preparation Kit
Formulation: Multi-component kit
Sufficient For: 10 antibody samples, each containing 0.25 to 4mg IgG

Kit Contents:
Pierce Papain Agarose, 1.25mL
Cysteine-HCl, 1g
Fab Digestion Buffer, 120mL
NAb Protein A Spin Column, 1mL, 1 column
PBS Packs (each makes 500mL), 2 packs
IgG Elution Buffer, 120mL
Zeba Desalt Spin Columns, 7K MWCO, 2mL, 10 columns
Spin Columns, 0.8mL, 10 columns
Microcentrifuge Tubes, 2mL, 30 tubes
1.25mL-resin kit Product Instructions for product #44985 Pierce Fab Preparation Kit MSDS for product #44985 Pierce Fab Preparation Kit Certificate of Analysis for product #44985 Pierce Fab Preparation Kit $505.00

Add to cart

Instructions | MSDS | CofA
Product Instructions | MSDS | CofA  

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