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Related Experiment Videos

A selective protein sensor for heparin detection.

Shenshen Cai1, Jodi L Dufner-Beattie, Glenn D Prestwich

  • 1Department of Medicinal Chemistry and Center for Cell Signaling, The University of Utah, 419 Wakara Way, Suite 205, Salt Lake City, UT 84108-1257, USA.

Analytical Biochemistry
|February 11, 2004
PubMed
Summary

A novel heparin sensor was engineered using a hyaluronan-binding domain. This sensor accurately detects unfractionated and low-molecular-weight heparin in human plasma, enabling precise therapeutic level monitoring.

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Area of Science:

  • Biochemistry
  • Biotechnology
  • Analytical Chemistry

Background:

  • Direct clinical assays for heparin detection in blood are currently unavailable.
  • Heparin monitoring is crucial for managing anticoagulant therapy and preventing complications.
  • Existing methods for heparin quantification can be complex and time-consuming.

Purpose of the Study:

  • To engineer a sensitive and selective sensor for the direct detection of heparin in biological samples.
  • To develop a novel assay for measuring unfractionated heparin (UFH) and low-molecular-weight heparin (LMWH) in human plasma.
  • To assess the sensor's performance characteristics, including sensitivity, selectivity, and precision.

Main Methods:

  • Engineered hyaluronan (HA)-binding domains (HABD) into GST fusion proteins with varying copy numbers.

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  • Purified GST-HABD constructs and determined their binding affinities for heparin and HA using competitive ELISA.
  • Assessed the sensor's ability to detect UFH and LMWH in human plasma through dose-response studies.

    • Main Results:

    • The three-copy HABD construct (GST-HB3) demonstrated high affinity and selectivity for heparin over other glycosaminoglycans.
    • GST-HB3 detected unfractionated heparin as low as 39 ng/ml (0.1 U/ml) with a signal-to-noise ratio of 5.6.
    • The sensor linearly detected UFH and LMWH in human plasma within clinically relevant ranges, showing high precision (CV < 12%).

    Conclusions:

    • The engineered GST-HB3 construct serves as a sensitive and selective heparin sensor.
    • This novel sensor has the potential for accurate measurement of therapeutic heparin levels in clinical settings.
    • The developed assay offers a promising tool for real-time heparin monitoring in patients.