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Method of selective conjugation without unwanted cross-linking

Description:

Although there have been continuous efforts to improve various aspects of the assays that are commonly used in clinical medicine and research laboratories, there still exists a problem of preparing a large quantity of an analyte-enzyme conjugate of high purity by a simple and efficient process. This invention is an improved process of selectively preparing a conjugate of an analyte-enzyme without generating undesired multimers of the enzyme.

The invention involves blocking the carboxyl functional groups of an enzyme which would normally be responsible for enzyme-enzyme cross linking. This leaves mainly amines as unique reactive functional groups therefore having little to no effect on the kinetic properties of the enzymes.

This approach increases the yield of the reaction and reduces the need for (chromatographic) purification of analyte-enzyme conjugates due to lower proportion of cross-linked materials saving time and money. Analyte conjugates prepared according to this process can be employed in a variety of biochemical techniques and assays including, but not limited to,

  • enzyme-linked immunosorbent assays (ELISA),
  • competitive immunoassays,
  • in situ chemical staining, and
  • immunohistochemical assays

The technology is ideally applied in assays where the enzyme is a peroxidase, a phosphatase, an esterase or a galatosidase. An example application of the invention eliminated enzyme–enzyme cross-linking by blocking the solvent-accessible carboxyl groups of horseradish peroxidase and alkaline phosphatase, with dialysis being the only purification step necessary.

Potential Applications
For performance improvement in assays where ligand-enzyme conjugation is required.

Opportunity
Available for non-exclusive or exclusive licensing and UNR is seeking parties interested in collaborative research to further develop, evaluate, or commercialize this technology.

IP Status
UNR ID: UNR02-011
METHOD FOR SELECTIVE CONJUGATION OF ANALYTES TO ENZYMES WITHOUT UNWANTED ENZYME-ENZYME CROSS-LINKING
US Patent Appl. No.: 10/494,163

    Patent Information:
    For Information, Contact:
    Dan Langford
    Technology Commercialization, Manager
    University of Nevada, Reno and Desert Research Institute
    dlangford@unr.edu
    Inventors:
    Vincent Lombardi
    David Schooley
    Keywords: