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HaloTag Technology: a multifunctional technology enabling protein purification, analysis of protein interactions, cellular imaging, and surface display for overall study of protein function

In the post-genomic era a major objective is to fully characterize the human proteome involving identification of proteins, determination of their localization, modifications, interactions, activities and ultimately their function. Essential to this ambition is development of new affinity and imaging tools allowing comprehensive analysis of protein function. Here we present a new technology based upon the use of a protein fusion tag, termed HaloTag, which allows for highly specific, oriented, and covalent immobilization on surfaces, and also protein labeling based on covalent binding with a variety of fluorescent ligands. Because of the multifunctional nature of HaloTag technology, analysis of many different aspects of protein function can be successfully achieved using a single construct.


Abstract: "HaloTag� Protein Arrays"

Protein microarray technologies are increasingly being employed to study protein-protein interactions, enzyme activities, protein profiling, and antibody screening. Despite great promises, applications of protein arrays in functional proteomics have lagged because of time consuming steps required for generating purified protein content, difficulties in capturing proteins at high density in stable conformation and need for complex robotics for arraying proteins. To address the needs of scientists interested in medium throughput custom protein arrays we used HaloTag multifunctional technology to provide a HaloLink� Array platform for covalent and oriented capture of proteins on solid substrates directly from the protein expression systems without any prior purification step. HaloLink� Arrays use glass slides coated with polyethylene glycol polymer known to resist non-specific adsorption of unwanted protein and prevent surface-induced denaturation of specific proteins. An easily removable silicone gasket is used to create 50 wells on the glass slides, allowing multiple assays to be performed manually on same slide without any specialized equipment.

To create a custom protein array, proteins of interest are expressed as HaloTag� fusion (in cell-free or cell based protein expression systems) and then covalently captured on a hydrogel coated HaloLink� glass slide by simple 1hr incubation. Silicone gasket allows upto 50 different proteins to be captured in parallel for multiplexed studies. HaloLink� arrays have been used for protein:protein and protein:DNA interaction studies as well as capture of protein biomarkers related to autoimmune diseases. The ability to easily create a protein array will allow proteomics researchers to shift their efforts from creating arrays to studying interesting biological questions using arrays.



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