Amanda Bukhtia and Kaitlyn Tingley

Lawrence Technological University

Advisors: Dr. Auner, Dr. Bou-Akl, Dr. Li and Dr. Nasir

Need

“Develop a quick and cost effective method to detect if a

patient has (MRSA)”

Solution

• A novel impedance biosensor

Project Summary

.

1

What is MRSA?

An antibiotic resistant

staph infection

Can enter the bloodstream

through open wounds

which can lead to death

75,309 people are

estimated to have MRSA

every year

http://www.cdc.gov/mrsa/tracking/index.html

2

Current Diagnostic Methods3

http://laboratory-manager.advanceweb.com/Archives/Article-Archives/MRSA-Screening-Culture-or-Molecular.aspx

http://ac.els-cdn.com/S0956566311005197/1-s2.0-S0956566311005197-main.pdf?_tid=55edafa6-5648-11e4-beaa-

00000aacb35f&acdnat=1413583120_a5737397e845f21a215313800649bf76http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1287783/ http://www.rapidmicrobiology.com/test-method/methicillin-resistant-

staphylococcus-aureus-mrsa-detection-and-identification-methods/

http://www.biomedcentral.com/1471-2180/6/83

https://workspace.imperial.ac.uk/opticalandsemidev/public/Publications/LAWES.pdf

http://www.aist.go.jp/aist_e/latest_research/2012/20121121/20121121.html

• Penicillin binding protein

2A (PBP2a)

• Responsible to antibiotic

resistance in MRSA

• Regulates the cell wall of

MRSA bacteria

PBP-2a Antibody4

Device Design5

Device Design

.

6

Device A Device B

Device Fabrication

Glass Wafer Fabrication1. Electrodes (Au)

2. Passivation (SiO2)

PDMS Fabrication

Device Bonding

7

Device A Device B

Electrode Fabrication8

Passivation Layer Fabrication9

Channel Fabrication10

Device Bonding

• Oxygen plasma

treated PDMS and

glass

• Bonded through

minimal compression

and heat

11

Impedance Testing12

Impedance

Analyzer

Syringe

Pump

Device

Probe

Impedance Testing (Part 1)

• Four different concentrations of PBS

13

Impedance Testing (Part 2)

• Impedance (Ω) is inversely hyperbolic to ion concentration

14

Impedance Testing (Part 3)

• Device was placed on

magnetic platform

• Steel particles were

mixed into a 1x PBS

solution

• Impedance decreases

as steel particles are

captivated by the magnet

onto the electrode

15

Antibody Binding Testing

Testing Conditions

• Three concentrations of anti-PBP-2a

• Whole MRSA

- Experimental

• Whole methicillin-sensitive staphylococcus aureus

MSSA

-Control

• Fluorescent staining of antibody/bacteria complex

-Using Alexa Fluor

16

Antibody Binding Testing17

Results

• Positive detection of anti-PBP-2a/<RSA complex

-Green fluorescence

• Failure to detect anti-PBP-2a/MSSA

-No fluorescence

Antibody Immobilization 18

Results19

• PBP-2a antibody will bind to MRSA bacteria but not to

MSSA

- This shows the specificity of our biological reagent

• Fluid dynamics and impedance mechanisms of device

have been validated

- Device A provides superior results due to enhanced

PDMS adhesion

Timeline20

Team Structure

Amanda Kaitlyn

OSHA Training

Fabrication

Impedance

Testing

Antibody

Immobilization

Antibody

Binding Testing

Device Design

21

Future Outlook22

http://www.nature.com/srep/2013/130521/srep01863/full/srep01863.html#suppl

ementary-information

http://www.sciencedirect.com/science/article/pii/S1549963413005893

Establish flow through testing conditions using

COMSOL modeling

Validate anti-PBP-2a immobilization

Conduct device impedance testing with MRSA

Integrate impedance analyzer

Create packaging for device

Acknowledgements

Wayne State University’s Smart Sensors and Integrated

Microsystems (SSIM)

Dr. Auner

Lawrence Technological University

Dr. Li

Dr. Nasir

Dr. Bou-akl

The LEGENDS Entrepreneurial Student Awards

23

Questions?

MRSA Diagnostics Amanda Bukhtia and Kaitlyn TingleyDr. Auner, Dr. Bou-Akl, Dr. Li and Dr. Nasir

Objective: To create a novel device for the rapid detection of MRSA.

Approach: Create a microfluidic device to detect MRSA due to these devices quick turn around time and their cheap manufacturing costs.

Impact: Began proof of concept for a novel solution to detect MRSA quickly while maintaining high specificity

Progress and Results:

1. We have validated the impedance mechanism of our device

2. We have verified the specificity mechanism of our device

3. We have shown device A has better binding properties

Antibody Testing

Impedance Testing