Process multiple serum samples for 2D or 2D-LC in less than 40 minutes.

The Thermo Scientific Pierce Albumin/IgG Removal Kits are optimized to decrease the abundant albumin and antibody components of human serum sample in preparation for 2D electrophoresis and other protein profiling methods. By eliminating most of these two proteins, which account for nearly 70% of total serum proteins, other less abundant proteins of interest can be more easily detected and studied. Both the Antibody-based and Dye/Protein A-based Kits are scalable and sufficient to process 25 samples, each containing 600 µg of total serum protein (about 10 µl of serum).

The Antibody-Based Kit (Product # 89876) uses specific antibodies against human serum albumin (HSA) and human gamma immunoglobulin (IgG) immobilized to beaded agarose resin. When samples are incubated with the affinity resin, both HSA and IgG bind. Centrifugation to separate the supernatant from the resin results in sample in which HSA and IgG concentrations have been decreased to negligible levels. The Antibody-Based Kit provides the most specific method for removing HSA and IgG, and it is suitable only for human samples.

The Dye/Protein A-Based Kit (Product # 89875) uses a mixture of Cibacron Blue Dye and Protein A agarose affinity resin. The dye binds to albumin with relatively good specificity, and Protein A binds to many species and subclasses of IgG. The kit is more economical than the Antibody-Based Kit and can be used to treat samples from other species besides human (e.g., monkey, swine, rabbit).

Highlights:

• Optimized for 2D sample prep – microcentrifuge spin cup method is scaled and optimized for treating 10 µl human serum samples for 2D electrophoresis
• Efficient – kits remove nearly all HSA and IgG, enabling lowabundance proteins to be detected in 2D gels
• Fast – kits process samples in less than 40 minutes
• Scalable – use different amounts of affinity resin to match sample volume and concentration
• Two kit options – choose the system appropriate for your application: highest specificity (Antibody-Based Kit) or greater ecomony (Dye/Protein A-Based Kit)

Low-abundant proteins in human serum and plasma can provide information about human diseases. However, human fluid analysis is often complicated by the presence of high concentrations of albumin and IgG, which can make up more than 70% of total serum protein (Figure, Panel A). The Antibody-based Kit processes 12 samples of approximately 600 µg of human serum. It is suitable for detailed proteomic analysis because it uses highly specific immobilized antibodies for the removal of albumin and IgG, producing minimal nonspecific interactions (Figure, Panel B). The Dye/Protein A-based Kit uses a classical mixture of Cibacron Blue Dye and Protein A resin for economical removal of albumin and IgG. The kit can process up to 25 samples of approximately 600 µg of serum using an optimized protocol to minimize nonspecific protein interactions and sample dilution (Figure, Panel C).

2D gels of unprocessed vs. processed human serum. Approximately 30 µg of unprocessed human serum was isoelectric-focused on a pH 5-8 strip and then separated in the second dimension on an 8-16% SDS polyacrylamide gel. The Pierce Silver Stain Kit (Product # 24612) was used to visualize bands. Panel A. Unprocessed serum. Large poorly resolved bands corresponding to human serum and IgG obscure much of the gel.
Panel B. Processed with the Pierce Antibody-Based Albumin/IgG Removal Kit. With albumin and IgG removed, low-abundant proteins are readily identified.
Panel C. Processed with the Pierce Albumin/IgG Removal Kit (dye- and Protein A-based). With albumin and IgG removed, low-abundant proteins are readily identified.