Spot-Trap Affinity Reagents
Capture Spot-tagged Proteins
Simple purification with ultra-low background
The Spot-Trap is a high quality Spot-Tag binding protein coupled to a resin matrix designed for use with immunoprecipitation or affinity purification protocols. It will only interact with the novel 12 amino acid sequence found on Spot-Tag fusion proteins. Spot-Tag has many advantages as compared with other short peptide “tag” systems. Like all Nanobody-Traps, the Spot-Trap binding protein is a small (14.7 kDa), monovalent, high-affinity monoclonal alpaca antibody (“nanobody”), and perfectly recognizes the Spot-Tag sequence motif (PDRVRAVSHWSS) at either the N-terminus or C-terminus of the expressed fusion protein. The Spot-Trap is covalently coupled to the surface of agarose or magnetic-agarose beads for fast and effective immunoprecipitation and affinity purification. It is optimized for biochemical analysis of Spot-Tagged proteins and their interacting partners. It has been tested for multiple applications and works efficiently in various systems, such as bacteria, yeast, mammalian cell lines, and insect cells. For biochemical analysis including mass spectrometry and enzyme activity measurements, Spot-Tag-fusion proteins and their interacting factors can be isolated quickly and efficiently by immunoprecipitation using the Spot-Trap.
When the results matter use Spot-Trap
To the left, immunoprecipitation was performed on a Spot-Tagged GFP. It is apparent that Spot-Trap not only lacks heavy and light chain antibody contaminations, but also lacks non-specific binding of contaminating proteins. This makes Spot-Trap a superior affinity capture tool for single band purification of Spot-tagged proteins. To the right, the same Spot-Tagged GFP was bound to matrix and then competitively eluted with 100 µM Spot-Peptide. This gentle affinity purification technique is a perfect way to preserve protein structure and function, and to look at larger protein complexes.
Spot-Trap is specially designed for spotless background
To the right, immunoprecipitations were conducted with HEK293T cell lysates without Spot-Tag or any other expressed tagged proteins present (each lane was normalized for equal binding capacity). The IPs were performed according to respective manufacturers’ protocols. Both versions of Spot-Trap affinity reagent show almost no non-specific binding of endogenous mammalian cell proteins because of a superior, re-engineered wash buffer. This promises clean results every time!
Camelidae single-domain antibodies are like IgGs on steroids
The family of animals known as Camelidae (camels, llamas, and alpacas) produce functional antibodies devoid of light chains, so-called "heavy chain" antibodies. These heavy chain antibodies recognize and bind their antigens via a single variable domain. When cleaved from their carboxy tail, these barrel-shaped structures (2x3 nm) are extraordinarily small, naturally-occurring, and intact antigen binding fragments (MW of 13 kDa). These fragments, called Nanobodies, are characterized by high specificity, affinities in the low nanomolar range, and dissociation constants in the sub-nanomolar range (typically 10- to 100-fold better than mouse IgGs). The compact size of Nanobodies makes them extremely stable at temperatures up to 70°C, and functional even in 2M NaCl or 0.5% SDS. These small and powerful antibody fragments can be used in a variety of unique applications. They will open up your research possibilities.
Spot-Traps Coupled to Agarose
Spot-Traps Coupled to Magnetic Agarose Beads
Spot-Trap Non-Coupled
Spot peptide
Binding Controls
Spin Columns for agarose beads
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SPECIFICATIONS
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| Configuration: | Specific Camelidae antibody linked to agarose bead |
| Particle Size: | ~90 µm when cross-linked to 4% agarose beads
~40 µm when coupled to magnetic agarose |
| Binding Capacity: | 10µl Spot-Trap Agarose slurry binds more than 7µg of a 30 kDa recombinant Spot-tagged protein |
| Storage Buffer: | 20% EtOH |
| Washing Conditions: | Stable up to: 65°C 2M NaCl 2M Urea |
| Part Numbers | ETA010, ETA020, ETA100, ETA200,ETA400, ETAK020, ETMA010, ETMA020, ETMA100, ETMA200, ETA400, ETMAK020, EP1, etb250 |
| Storage and Stability: | store at 4°C; stable for one year. Do not freeze. |