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Multiplexed GPCR Characterization Using SPR

Award Information

Agency:
Department of Health and Human Services
Branch:
N/A
Award ID:
Program Year/Program:
2011 / STTR
Agency Tracking Number:
R42MH084372
Solicitation Year:
2011
Solicitation Topic Code:
NIMH
Solicitation Number:
PA11-097
Small Business Information
Wasatch Microfluidics, LLC
4909 BROWN VILLA COVE SALT LAKE CITY, UT 84123-
View profile »
Woman-Owned: No
Minority-Owned: No
HUBZone-Owned: No
 
Phase 2
Fiscal Year: 2011
Title: Multiplexed GPCR Characterization Using SPR
Agency: HHS
Contract: 2R42MH084372-02A1
Award Amount: $811,093.00
 

Abstract:

DESCRIPTION (provided by applicant): The goal of this Phase II STTR project is to develop a real-time label-free biosensor that can analyze 96 samples at a time, compared to the 6 samples possible with current technologies. This platform will initially bedemonstrated with G-protein-coupled receptors (GPCRs) and antibodies. What both applications have in common is the need for higher- throughput sensing, and demonstrating the integrated system for these will illustrate its versatility and potential contributions across the wide spectrum of biosensor applications. For the GPCR demonstration, our work with standard SPR instruments has shown that the choice of detergent(s) is critical for obtaining active solubilized receptor. However, standard low throughput SPR biosensors have two overwhelming drawbacks: (1) The analysis of 96 solubilization conditions requires more than two days and the receptor loses significant activity during this time, which makes it difficult to compare the results obtained at the beginning and end of the analysis and (2) the SPR instrument is limited to testing only one analysis buffer at a time, which means that the success of the entire assay depends on the initial choice of analysis buffer. In Phase I, we developed a 96-channel Continuous Flow Microspotter (CFM) printhead and demonstrated the ability to print GPCRs onto a sensor surface directly from crude media using up to 96 different analysis buffers. The GPCRs were also kept wetted and active throughout the printing process by theCFM's enclosed microchannel printing network. In the final experiment, we solubilized the GPCR CCR5 from whole cells using 192 different detergent conditions and spotted them onto an SPR sensor surface using our CFM printhead. We then tested the activity of the receptor and used the ligand binding results to determine that a certain combination of detergents best enhanced receptor activity. To run this analysis with a standard Biacore technology (e.g. T100) would have required four days of instrument time.In comparison, we were able to perform the analysis in less than 2 hours. In Phase II, we propose to integrate the 96-channel CFM from Phase I with a commercial SPR imager to enable automated interaction analysis in a highly parallel format. The followingspecific aims detail the combination of Wasatch's microfluidic technologies with the commercial IBIS SPR imager to produce a high-throughput label-free biosensor. 1. Refine the 96-channel CFM printhead design from Phase I to enable optimal performance whenmounted on the IBIS SPR imager. 2. Mount the 96-channel CFM onto the IBIS SPR imager and optimize the fluidic parameters that affect platform sensitivity and uniformity. 3. Automate the CFM and SPR components within one seamless instrument: Automation ofthe flow cell positioning, sealing, fluid handling, valving, in-line degassing and temperature control. Integration of the CFM and SPR imager control software and data collection/analysis software. 4. Demonstrate use of the automated system with GPCRs andantibodies. PUBLIC HEALTH RELEVANCE: Antibody analyses are one of the most common applications of biosensor technology and are typically straightforward. GPCRs are the hottest and most challenging system that biosensor users are tackling. Up to halfof the drugs on the market today modulate some form of GPCR activity, and it is estimated that 25-50% of the total drug targets are in the GPCR families. GPCRs are the most studied of the major drug target classes, yet they are challenging to work with because they are normally membrane associated, present in low abundance, and unstable. By enabling the high throughput study of GPCRs, there is enormous potential for speeding drug development, treatments, and the associated health of patients with hundredsof different diseases.

Principal Investigator:

James S. Smith
801-532-4486
jsmith@microfl.com

Business Contact:

Josh Eckman
801-532-4486
josh@microfl.com
Small Business Information at Submission:

WASATCH MICROFLUIDICS
4909 BROWN VILLA COVE SALT LAKE CITY, UT -

EIN/Tax ID: 183041304
DUNS: N/A
Number of Employees: N/A
Woman-Owned: No
Minority-Owned: No
HUBZone-Owned: No
Research Institution Information:
UNIVERSITY OF UTAH
UNIVERSITY OF UTAH
75 South 2000 East Room 111
SALT LAKE CITY, UT 84112-
Contact: