Nanomolar Level Specific Detection and Quantification of Phosphatidic Acid with a Protein-based Probe

 

Executive Summary

 

Phosphatidic acid is an important lipid for metabolism, membrane curvature, and signal transduction within cells. Although essential for all life, this molecule exists in relatively low concentrations and presents a challenge for both researchers and industry to monitor.  Current methods of detection include two-dimensional thin lay chromatography (2D TLC) coupled with gas-liquid chromatography (GLC) and tandem mass spectrometry (MS/MS), both of which are time consuming and require expensive instrumentation.  MSU researchers have developed a biologically based assay to distinguish phosphatidic acid from all other abundant forms of lipids in both plants and animals. This technology represents a fast, accurate, low-cost, and highly effective method in which to measure nanomolar amounts of phosphatidic acid.

 

Description of Technology

 

This MSU technology utilizes a lipid-protein overlay assay to detect phosphatidic acid specifically and unambiguously on a nanomolar scale. No special equipment is needed beyond that available in routine clinical lab facilities and can be used for high throughput samples. This technology uses a recombinant protein probe that includes a binding motif of the Arabidopsis TGD4 protein fused to a fluorophore such as DsRED. The TDG4 motif binds solely to phosphatidic acid, and works on material sourced from either animal or plant material. Purified probes can be obtained through routine use of a plasmid-based bacterial expression system and Ni-NTA columns.  A lipid-protein incubation on nitrocellulose membranes allows quantification through the use of anti-His antibodies and standard imaging software.

 

Key Benefits

  • Nanomolar level of detection
  • Specific binding to phosphatidic acid over other lipids
  • No specialized clinical lab materials/equipment required
  • Faster than conventional methods (2D TLC w/ GLC; MS/MS)

 

Applications

  • Quantitative assay for phosphatidic acid

 

Patent Status: 

 

U.S. patent 8,629,251 issued January 14, 2014.

 

Licensing Rights Available:

 

Full licensing rights available

 

Inventors: Christoph Benning, Zhen Wang

 

Tech ID: TEC2012-0055

 

 

Patent Information:

For Information, Contact:

Thomas Herlache
Assistant Director
Michigan State University
herlache@msu.edu
Keywords: