Annual Report - Center for Food Safety Engineering - Purdue ...

The Center for Food Safety Engineeringg 

2006 - 2007 Research Report 

“Collaborating to make our food safer”

The mission of the Center for Food Safety 

Engineering is to develop new knowledge, 

technologies and systems for detection 

and prevention of chemical and microbial 

contamination of foods. 

Through CFSE, Purdue University positions 

itself as a national leader in multidisciplinary 

food safety research. Our multidisciplinary 

approach, including a strong 

engineering component, makes Purdue 

University truly unique.

2006-2007 Research Report 

2 Welcome from the Director 

2 Message from USDA 

3 A discussion with Dan Fung: 

The past, present, and future of microbial detection 

4 Engineering of biosystems for the detection of Listeria monocytogenes in foods 

• Michael Ladisch, Rashid Bashir, Arun Bhunia, J. Paul Robinson 

6 Multipathogen screening using immunomicroarry 

• Arun Bhunia, Mark Morgan, B.K. Hahm, Andrew Gehring 

7 Optical biosensors for food pathogen detection 

• Arun Bhunia, Mark Morgan, Viswaprakash Nanduri, Andrew Gehring, Shu-I Tu 

8 Optical forward scattering for bacterial colony differentiation and identifi cation 

• Arun Bhunia, E. Daniel Hirleman, J. Paul Robinson 

9 Immunocapture real-time PCR to detect mycotoxigenic mold spores in grains 

• Maribeth Cousin, Charles Woloshuk 

10 Nanoparticle based DNA multiplexed probes for pathogen detection using confocal 

raman microscopy 

• Joseph Irudayaraj 

11 Rapid, quantitative, and reusable immunosensors for bacteria detection on a 

microfl uidic platform 

• Chang Lu, Arun Bhunia, Zhongyang Cheng 

12 Continuous monitoring of chemical agents in aqueous media using bioreporterbased 

sensors 

• David Nivens, Michael Franklin, Carlos Corvalan 

13 Scientifi c Publications and Presentations 

16 Center Staff 

Visit us @ www.cfse.purdue.edu

Welcome from the Director 

The Center for Food Safety Engineering (CFSE) celebrates its seventh anniversary. Our 

collaboration with USDA-ARS Eastern Regional Research Laboratory continues to breed success 

and has led to 21 peer-reviewed research publications and 22 presentations and proceedings 

this year. Purdue and USDA-ARS scientists teamed up and presented a series of lectures for 

“Molecular Day” at Kansas State University’s 27 th annual Rapid Methods and Automation in 

Microbiology Workshop. This effort alone led to a series of 11 peer-reviewed manuscripts published 

in the March and June issues of the Journal of Rapid Methods and Automation in Microbiology. 

Dr. Richard H. Linton 

Director of the Center for 

Food Safety Engineering 

Our scientists continue to work on technology platforms to improve microbial and chemical 

detection. Research is being done to develop detection systems for many bacterial pathogens, 

including Listeria monocytogenes, Escherichia coli O157:H7, Campylobacter spp., Salmonella spp., 

and for chemical toxins like arsenite. We are utilizing different technologies to improve detection 

capabilities, including enzyme-linked immunosorbant assays, polymerase chain reactions, 

impedance-based microbiology, infrared spectroscopy, scanning microscopy, confocal Raman 

microscopy, bioluminescense, DNA/RNA probes, and bioreporter-based chemical sensors. 

One of our most signifi cant breakthroughs this year was the further development of the BActeria 

Rapid Detection using Optical scattering Technology, or BARDOT system. This technology uses 

optical forward scattering techniques to identify different microorganisms after scanning colonies 

that grow on an agar surface. The resulting image produces a “fi ngerprint” that can be used to 

specifi cally identify organisms of interest. 

I continue to be impressed with the collaborative research effort of Purdue University and USDA- 

ARS scientists and feel privileged to serve as director of the center. If you are interested in learning 

more about CFSE, please visit our Web site at www.cfse.purdue.edu, or feel free to contact me 

directly. 

Message from USDA 

Dr. Shu-I Tu 

Supervisory Research 

Chemist USDA-ARS, 

Eastern Regional 

Research Center 

As the collaboration between the Center for Food Safety Engineering (CFSE) at Purdue University 

and the Eastern Regional Research Center (ERRC) of the Agricultural Research Service (ARS) 

enters its eighth year, I am grateful to witness the growth and maturation of this collaborative 

arrangement. In 2007, we were pleased to learn that the research of the Purdue team has been 

selected for the ARS OSQR (Offi ce of Scientifi c Quality Review) process. This process is normally 

conducted for ARS in-house research projects. Thus, this selection signals that the Purdue project 

is now considered an integral part of ARS research efforts. I am proud to witness the continual 

growth of our Purdue colleagues’ research impact. Together, our Purdue-ERRC team has received 

increased recognition as an important contributor to the technology advancement of pathogen 

detection in food as evident by the invitation from the International Workshop on Rapid Methods and 

Automation in Microbiology to conduct a half-day symposium on molecular methodologies in August 

2006 and again in June 2007. We were pleased that a dedicated issue of the Journal of Rapid 

Methods and Automation in Microbiology to report the progress of our collaborative team has been 

published. More recently, our collaboration has attracted interest from the National Food Safety 

Center of China. They have expressed a desire to form an international collaboration arrangement 

under the general agreement between USDA and the Ministry of Science and Technology, China 

(MOST). Dr. Xianming Shi, the deputy director of the Chinese Food Safety Center, plans to attend 

our joint annual meeting this year at Purdue in October. I am looking forward to this opportunity to 

expand the ARS-Purdue collaboration into an international enterprise. 

2 

Center for Food Safety Engineering

A discussion with Dan Fung: 

The past, present, and future of microbial detection 

Dr. Fung started his career in microbial detection as a graduate student at the University of North 

Carolina-Chapel Hill when his professor asked him to identify several hundred cultures of bacteria 

from sewage. The work was so tedious that Dan thought “there must be a better way to do 

microbiology!” He then traveled to Iowa State University and pursued his Ph.D. working under Dr. 

Paul Hartman, whom Dan considers his mentor. Here, Dan began research on miniaturization of 

microbiology tests and developed the Miniaturized Microbiological Cultural and Viable Cell Count 

systems. Forty years later, Dr. Fung is recognized as a worldwide leader in microbial detection. He 

just completed advising his 100th graduate student; has instructed over 18,000 people; has lectured 

throughout the world; and has authored over 800 publications. 

This summer, Dan completed his 27th annual workshop on rapid methods at Kansas State University, 

which is a program that has attracted over 4,000 participants from more than 60 countries. In 2005, 

Dan fi rst invited a team from Purdue University and USDA-ARS to participate in his workshop. CFSE 

has now participated in this program for the past three years. We are glad for the opportunity and 

embrace our collaboration with Dr. Fung. 

We had a chance to catch up with Dan and ask him the following questions: 

1. What do you consider to be the most significant breakthroughs related to microbial detection in the 

past 40 years? 

Dr. Dan Fung 

Kansas State 

University 

• “Miniaturization from test tubes, miniaturized chambers, microwells, microarrays, 

biosensors, and nanotechnologies has greatly improved the capacity to 

study thousands of samples in a much more effi cient manner. 

• The effective use of ELISA tests, instrumentation, sensors, PCR, microarrays, microchips, 

biochips, nanotechnology, and a variety of physical, chemical, biochemical methods. 

• Ingenious and effective sampling technologies for food, water, air, medical, environmental, and 

industrial samples have greatly helped to make analyses more accurate and effi cient.” 

2. What was your most significant research finding or contribution? 

• “Miniaturization of microbial techniques (cultural methods and viable cell counts 

methods) directly and indirectly infl uenced the revolution of diagnostic kits 

and procedures in medical, food and environmental microbiology. 

• Development of detection methods for staphylococcal enterotoxins, Listeria 

monocytogenes, Escherichia coli O157:H7, and Clostridium perfringens. 

• Creation of dye-containing media for the rapid detection and differentiation of bacteria, yeast, and molds. 

• Advancement and promotion of enzymes systems, such as Oxyrase, 

for stimulating the rapid growth of foodborne pathogens.” 

3. Where have we made progress, and where have we failed to make progress? 

“We have made great progress in sample preparation and rapid detection of target molecules and cells by a 

variety of sophisticated technologies and approaches. We now can identify microorganisms and classify them 

to the species, subspecies, and molecular levels very effi ciently. We have not been able to effectively separate 

target organisms from the background food systems yet. And, there is no instant, on-line test for pathogens 

and may never be since microorganisms are so small compared with the food matrix. We still have much to 

do.” 

4. What technologies or approaches do you feel have the most promise for the future? 

“It depends on what you want to fi nd. I do believe nanotechnologies developed and used properly can greatly 

assist in the development of rapid detection and enumeration of pathogenic and non-pathogenic organisms. I 

think this is our future.” 

5. What do you think our greatest challenges will be in the future? 

“I think we need to be able to: 

• Predict and detect the arrival of new and exotic microorganisms. 

• Control or eliminate pathogenic organisms better in food systems. 

• Educate and stimulate another generation of bright people to put their minds 

together to solve the problems at hand and the problems unseen.” 

3 


La 

di 

sc 

h, 

Bas 

hi 

r, 

Bhu 

huni 

a and Ro 

bi 

nson 

Engineering of biosystems for the detection 

of Listeria monocytogenes in foods 

Investigators: Michael Ladisch (Department of Agricultural and Biological Engineering), Rashid Bashir (School of Electrical and Computer 

Engineering), Arun Bhunia (Department of Food Science), J. Paul Robinson (Weldon School of Biomedical Engineering) 

Project Rationale 

Current methods for detecting L. monocytogenes rely upon 

enrichment procedures to increase bacterial numbers for 

detection. The food or food extract is incubated in a special 

growth medium for 12 to 24 hours, and the resulting culture is 

tested for L. monocytogenes using procedures that require an 

additional 3 to 24 hours. An overall time of 2 to 3 days is typical 

of the time that elapses between when the food is sampled 

and when the test results are available. The elapsed time, 

referred to as time to result or TTR, is problematic since some 

perishable, ready-to-eat foods are consumed before test results 

would be available. Rapid and affordable technologies to detect 

low numbers of L. monocytogenes cells directly from food and to 

distinguish living from dead cells are needed. 

This multi-disciplinary, multi-departmental research project is 

addressing the fundamental engineering and science required 

for developing a microchip that is capable of rapidly detecting L. 

monocytogenes at the point of use. Our primary goals are to 

establish microscale detection of L. monocytogenes on a realtime 

basis with a time to result of 4 hours and to reduce the 

time of bacterial culture steps with rapid cell concentration and 

recovery based on membrane technology. 

Our multidisciplinary research team is addressing the 

development, engineering and validation of the microchip 

system that combines bioseparation and bionanotechnology. 

Bioseparations technology will allow rapid concentration and 

recovery of microbial cells. Bionanotechnology enables the 

construction of systems capable of interrogating fluids for 

pathogens. The combination of the two technologies facilitates 

devices for rapid processing and detection of food pathogens. 

Project Objectives 

• Develop a system for rapid cell concentration and recovery. 

Improve membrane chemistry and methodology for 

handling complex food samples presented by blended 

hotdog, hamburger, vegetables, milk, and meat and 

decrease the volume in which the cells are captured by 

selecting or constructing the appropriate membrane design 

and combining with other bioseparation techniques. 

• Correlate media composition to changes in growth 

characteristics and metabolism of L. monocytogenes cells 

(recovery of stressed cells, capture from mixed cultures, 

increase sensitivity through decreasing media conductance) 

and develop media that enhance pathogenic cell response 

to detection methods. We also are interested in developing 

low conductivity media for enhanced capture and detection 

of stressed cells by antibodies and other bioreceptors. 

• Combine antibody-based capture and growth detection 

for the biochip. Integrated devices have been designed 

and microfabricated to combine ATP, pH, and/or direct 

nucleotide and antibody-based detection on-chip using 

multi-channel, multi-functional designs. The intent 

is to obtain biochips that sense multiple parameters 

simultaneously and improve DEP (di-electrophoresis)- 

based selective capture of L. monocytogenes and other 

pathogens in mixtures of cells. Then we will test for 

sensitivity of detection and selectivity of capture. 

4 

Center for Food Safety Engineering 

“Our multidisciplinary research team is addressing the development, engineering and 

validation of the microchip system that combines bioseparation and bionanotechnology.”

Ladi 

disc 

sch, 

Bas 

ashi 

r, Bhu 

huni 

a an 

d Ro 

bi 

ns 

on 

Project Highlights 

The concentration and recovery of microbial cells for analysis 

requires rapid recovery of viable cells while minimizing retention 

or loss of cells during processing. This work further extended 

the methods for achieving rapid concentration of homogenized 

or stomached food samples. We showed the utility of using 

a small hollow fi ber module with an internal volume of 23 or 

101 μL. The module is operated in a dead-end mode for the 

rapid concentration and recovery of various types of microbial 

cells in a small sample volume that is compatible with microscale 

detection systems. Tests were carried out with Bacillus 

thuringiensis DUP-6040, Escherichia coli K12, Listeria 

innocua FY248, Pseudomonas fl uorescens ATCC13525, and 

Streptococcus faecalis CG110 in PBS at initial concentrations of 

100 cfu/mL as well as E. coli or L. innocua in washing solution 

from hotdogs (i.e., massaged hotdog solution). After treating 

massaged hotdogs with protease and lipase, up to 53% recovery 

of viable cells at a concentration of 10 5 to 10 6 cfu/mL in a 100 

μL sample was achieved when E. coli and L. innocua cells 

were concentrated from a 250 mL massaged hotdog solution 

containing an initial microbe concentration of 600 to 900 cfu/ 

mL. The design and fabrication of hollow fi ber modules and 

membrane/microbe interactions during cell concentration and 

recovery was developed to achieve reproducible devices and 

rapid concentration of the cells. 

“The design and fabrication of hollow fiber modules and membrane/ 

microbe interactions during cell concentration and recovery was developed to 

achieve reproducible devices and rapid concentration of the cells.” 

5 


Bh 

unia 

, Mo 

rgan 

an, Ha 

hm and 

Geh 

ring 

Multipathogen screening using immunomicroarry 

Investigators: Arun Bhunia (Department of Food Science), Mark Morgan (Department of Food Science), 

B.K. Hahm (Department of Food Science), Andrew Gehring (USDA) 


Antibody-based methods are widely regarded as rapid and 

effi cient for detecting pathogenic foodborne bacteria. Application 

of conventional ELISA-based assays and further adaptation in 

modern biosensor tools show promise for rapid detection. Most 

assays are developed for detection of a single target pathogen 

or toxin. As a result, these methods can be very expensive 

when testing for multiple pathogens because separate assay 

methods are required. In addition, large laboratory space is 

required to perform separate tests for each target pathogen, 

and separate enrichment reagents and procedures are needed 

for each pathogen detection method. The development of a 

single test capable of detecting multiple pathogens, enriched in 

a single enrichment broth, will reduce costs and provide needed 

results in a short period of time. This technology would benefi t 

regulatory agencies and the food industry when evaluating food 

products for key food pathogens. 


For simultaneous growth and detection of Salmonella spp., E. 

coli O157:H7 and L. monocytogenes, our laboratory formulated 

a selective enrichment broth SEL (Salmonella, E. coli O157:H7, 

and Listeria). This year we evaluated SEL for its suitability to 

recover pathogens from inoculated ready-to-eat meat samples 

(deli turkey and salami). We were impressed with the overall 

performance of SEL. When SEL performance was compared 

with the Universal Pre-enrichment Broth (UPB, a commercial 

multiplex medium), SEL performance was comparable. However, 

SEL was also able to inhibit the growth of food-associated 

spoilage or natural contaminants while UPB failed. These 

results indicate that SEL has the potential to be used as a single 

selective enrichment medium for multiple target pathogens. 

In the last decade, several rapid detection methods—such as 

antibody-based, nucleic acid-based, and biochemical-based— 

have been developed. Even though these methods have 

shortened analysis times, there is still a time requirement to use 

selective enrichment of samples prior to using conventional rapid 

detection methods. Antibody-based methods, such as ELISA, 

require a minimum of 10 6 CFU/ml for cell detection. To achieve 

that cell level, it is important to use proper enrichment media. 


• Develop a microarray assay in 96-well plate and glass 

slide using a sandwich immunoassay for Salmonella 

spp., E. coli O157:H7 and L. monocytogenes. 

• Optimize growth and enrichment of these pathogens 

(healthy or stressed) spiked into model food samples. 

• Evaluate performance of the Pathogen 

Enrichment Device (PED). 

6 


“This year we evaluated selective enrichment broth for its suitability to 

recover pathogens from inoculated ready-to-eat meat samples.”

Bhun 

unia 

, Mo 

rgan 

, Nand 

ndur 

i, Geh 

ehri 

ng and 

Tu 

Optical biosensors for food pathogen detection 

Investigators: Arun Bhunia (Department of Food Science), Mark Morgan (Department of Food Science), Viswaprakash Nanduri (Department of Food Science), 

Andrew Gehring (USDA), Shu-I Tu (USDA) 


Our goal was to develop a fi ber optic sensor for detecting 

foodborne pathogens, including Listeria monocytogenes, 

Escherichia coli O157:H7 and Salmonella Enteritidis. We have 

been able to develop a fi ber optic sensor for L. monocytogenes 

and E. coli O157:H7. We have also developed a sensitive 

and specifi c fi ber optic detection assay for S. Enteritidis 

in poultry. The assay was compared with time-resolved 

immunofl uorescence (TRF) for confi rmation. An effi cient multipathogen 

array, using a fl ow through immobilization protocol, 

has also been developed for detection of L. monocytogenes, 

E. coli and S. Enteritidis. Pilot studies are currently underway 

to study the binding effi ciencies of an antibody-pathogen 

complex using different surface chemistries in order to have a 

better understanding of the molecular nature of interactions. 

This approach will help us increase sensitivity and specifi city of 

binding on the sensor. 


The development of a fi ber optic biosensor for detection of S. 

Enteritidis was the most important accomplishment this year. 

The success of the project provides proof of the principle of 

detection of S. Enteritidis using an effi cient fl ow through antibody 

immobilization using a fi ber optic biosensor. The low detection 

level achieved also emphasizes that target specifi c antibodies 

developed in our laboratory can be used as bio-probes on fi ber 

optic sensor platforms. 


• Develop and evaluate an antibody-coupled fi ber optic 

biosensor [ANALYTE 2000] for detection of S. Enteritidis. 

• Develop an effi cient, multi-pathogen array 

using the fi ber optic biosensor. 

• Screen and identify monoclonal and polyclonal 

antibodies, developed in our laboratory, for L. 

monocytogenes, E. coli O157:H7 and S. Enteritidis. 

• Develop effi cient surface chemistry protocols for evaluation 

and quantifi cation of binding interactions of the antibodypathogen 

complex on the surface of the fi ber optic sensor. 

• Deploy the common selective (SEL) media developed in 

our laboratory for the enrichment of targeted pathogens. 

• Explore the viability of other biosensor platforms for the 

development of a multi-pathogen array biosensor. 

“The development of a fi ber optic biosensor for detection of S. 

Enteritidis was the most important accomplishment this year.” 

7 


Bhun 

unia 

ia, Hi 

rl 

eman 

and 

Rob 

obin 

inso 

n 

Optical forward scattering for bacterial colony 

differentiation and identification 

Investigators: Arun Bhunia (Department of Food Science), E. Daniel Hirleman (School of Mechanical 

Engineering), J. Paul Robinson (Weldon School of Biomedical Engineering) 


The CDC estimates that 76 million people get sick, more than 

300,000 are hospitalized, and 5,000 Americans die each year 

from foodborne pathogen infections. Preventing foodborne 

illnesses remains a major public health challenge. Listeria 

monocytogenes, Escherichia coli, and Salmonella are three 

major foodborne pathogens of concern in the U.S. There has 

been an increase in foodborne illnesses, multiple outbreaks, 

product recalls, and loss of lives resulting from the association 

of pathogens in raw and processed, ready-to-eat food products. 

Bacterial contamination in products not only places the public 

at risk, but it is also costly to companies due to the loss of 

production time, product recalls and liability. 

For detecting and evaluating foods contaminated with L. 

monocytogenes or E. coli, USDA/FSIS recommends initial 

enrichment and subsequent plating on a selective agar media, 

which is often followed by further identifi cation procedures. 

These procedures are often time consuming and lengthy, since 

they take as much as 5 to 7 days. The present industrial demand 

is to increase the speed of the detection and to decrease 

economical losses and the chance of public health concerns. 

Our main objective was to develop a simple light scattering 

sensory method called the BARDOT (BActeria Rapid Detection 

using Optical scattering Technology) system in order to reduce 

the time of identifi cation of these pathogens after plating. 


• Improve the BARDOT design, including supporting 

physics-based models, for more repeatability and maximum 

discrimination of forward scattering signatures of colonies. 

• Acquire scatter images of colonies of select 

foodborne bacterial colonies including pathogens. 

• Analyze bacterial colonies of different foodborne 

bacteria on non-selective and selective agar media. 

• Validate the technology by using inherently 

contaminated food samples and samples that have 

been inoculated with selected pathogens. 

• Analyze cellular composition, cell arrangement, 

refractive index and colony contents using 

electron microscopy, FT-IR or GC-MS. 

• Analyze the scatter signal images using “standard feature 

extraction” and “moments of shape analysis” methods. 


The most signifi cant accomplishment this year was the design 

of an automated BARDOT system and related algorithm. We 

introduced a model for optical forward scattering by bacterial 

colony based on scalar diffraction theory. The model treats 

the colony as an amplitude/phase modulator and suggests 

macroscopic factors that cause the distinctive features shown 

in forward scattering signatures of the three types of Listeria 

species. We used phase contrast and confocal microscopy 

to provide independent information on the structure and 

morphology of the colonies that are fi xed parameters on the 

scattering model. We validated the experimental system using a 

chrome mask reference sample with known diffraction properties. 

Distinctive scattering patterns measured for three important 

species of Listeria were found to show good agreement with the 

model predictions. The results provide a physical explanation 

for the unique and distinctive scattering signatures produced 

by colonies of closely related Listeria species and support 

the effi cacy of forward scattering for rapid detection and 

classifi cation of pathogens without the use of labeling molecules. 

This improvement has provided the groundwork for developing 

the portable, stand-alone type of bacterial characterization 

instrument now known as BARDOT. 

8 


“We introduced a model for optical forward scattering by 

bacterial colony based on scalar diffraction theory.”

Co 

usin 

and 

Wol 

olos 

oshu 

huk 

Immunocapture real-time PCR to detect 

mycotoxigenic mold spores in grains 

Investigators: Maribeth Cousin (Department of Food Science), Charles Woloshuk (Department of Botany and Plant Pathology) 


Currently, there are few commercial rapid methods to detect 

molds and their spores in agricultural commodities, grains, 

and foods. In previous research, a protocol was developed to 

identify Fusarium species that produce two major mycotoxins, 

including fumonisins and trichothecenes. The antibodybased 

method was developed for Fusarium species to 

capture antigens of these mycotoxin-producers, which was 

then combined with a real-time PCR assay that was based 

on species-specifi c and genus-specifi c primers to identify the 

Fusarium species. The previous research was limited in spore 

capture effi ciency because the Fusarium spores were diffi cult 

to lyse for DNA release. We have designed this new project 

to help resolve these limitations by: 1) studying physical, 

enzymatic, and mechanical methods to break mold spores to 

release DNA for use in real-time PCR; and 2) incorporating 

the method into a immunocapture-qPCR method that uses 

antibodies produced against Fusarium graminearum and F. 

verticillioides and primers that are specifi c for the Tri6 gene 

involved in trichothecene biosynthesis and for the Fum1 

gene involved in fumonisin biosynthesis. This project was 

also designed to develop a library of PCR primers to other 

mycotoxigenic genera (Aspergillus that produce afl atoxins 

and ochratoxin and Penicillium that produce ochratoxin and 

patulin) for real-time PCR and to use these primers multiplex 

PCR formats to detect all major mycotoxin producers in the 

same assay. Antibodies to afl atoxin-producing molds (Yong and 

Cousin, 2001) and Penicillium species (Tsai and Cousin, 1990) 

were produced in earlier CFSE-funded research. 


• Develop primer sets to detect Aspergillus 

and Penicillium species. 

• Determine specifi city and sensitivity of 

primer sets and multiplex format. 

• Experiment with different methods to break 

mold spores of Fusarium species. 

• Optimize the capture of mold spores and release of DNA 

and use to detect Fusarium species in foods and grains. 


We designed PCR probes specifi c to species of Aspergillus 

(three probes), Fusarium (one probe), and Penicillium (two 

probes). Multiple genus-probes are required to capture minor 

groups or individual species that have slight differences in 

their DNA sequence. We have determined the optimum PCR 

conditions to achieve specifi city under multiplex conditions and 

developed quantitative curves. The linear range of the probes 

was between 0.01 ng and 1000 ng of target DNA. Thus far, the 

major probes for each genus have been tested. 

“Currently, there are few commercial rapid methods to detect molds and 

their spores in agricultural commodities, grains, and foods.” 

9 


Irud 

uday 

ayar 

araj 

aj 

Nanoparticle based DNA multiplexed probes for pathogen 

detection using confocal raman microscopy 

Investigator: Joseph Irudayaraj (Department of Agricultural and Biological Engineering) 


The overall goal is to develop a framework with probe 

fabrication and assay synthesis protocols for multiplex DNA 

detection of food pathogens by surface-enhanced Raman 

scattering (SERS) utilizing non-fl uorescent, label-containing 

nanoparticles as DNA probes. Although research on SERSlabeled 

DNA examination is very active, it is still in its early 

stages with regard to multiplexing and detecting analytes at 

low levels. Our team is capitalizing on the unique spectroscopic 

signatures (down to ~1 nm resolution) of non-fl uorescing 

molecules as labels (Raman tags) to identify specific DNA 

sequences. Because of the distinct fi ngerprint of the labels due 

to SERS, simultaneous detection of multiple DNA hybridization 

without separation is feasible at sub femto molar (fM) 

sensitivity. 

There are a number of unique aspects to this project. We can 

use multiplex labeling in one system using a range of nonfl 

uorescing labels. A single platform for detection of food 

pathogens at sensitivities not afforded by fl uorescence methods 

is possible using our approach. Incorporation of a magnetic 

separation step will enable the separation of target sequences 

in complex media. Use of non-fl uorecent labels [~$10-20/ 

gm] for multiplexing is many orders cheaper than fl uorescent 

labels [~$10-20/mg]. Furthermore, the choice of SERS labels 

is enormous (~over 1000 labels) and extremely sensitive, and 

single-molecule identifi cation has been reported. This implies 

that eventually the detection can be accomplished without the 

amplifi cation step. 


• Optimize the SERS effect of dye/gold particle size 

and excitation wavelength and concentrations with 

multiplexing. We have demonstrated that up to 

eight non-fl uorescent Raman tags can be chosen 

with distinct signatures for visual multiplexing 

utilizing the SERS spectra. The fabrication step has 

also been optimized, and detection sensitivity of 

up to 1 fM is achievable for the chosen target. 

• Fabricate SERS-tagged DNA probes for each of the 

fi ve target pathogens and multiplex demonstrations. 

Multiplexing of up to eight probes has been demonstrated 

for a chosen DNA sequence. We have also demonstrated 

that hybridization of eight different DNA sequences 

(depicting eight probes) at one time can be detected. 

• Design and implement an analysis assay for 

pathogen detection using SERS DNA probes. 


Our key accomplishment this year is the demonstration of an 

eight-plex, non-fl uorescent DNA detection assay using Raman 

spectroscopy. We are now undertaking steps to standardize 

the assay for direct detection of target sequences without the 

PCR amplifi cation step. The developed technique could be 

utilized as a slide (lab-on-slide) or tube (lab-on-tube) format for 

pathogen and disease detection. 

10 


“Our key accomplishment this year is the demonstration of an eight-plex, 

non-fl uorescent DNA detection assay using Raman spectroscopy.”

Lu 

, Bh 

un 

ia and 

Che 

heng 

Rapid, quantitative, and reusable immunosensors for 

bacteria detection on a microfluidic platform 

Investigators: Chang Lu (Department of Agricultural and Biological Engineering), Arun Bhunia (Department of 

Food Science), Zhongyang Cheng (Materials Engineering, Auburn University) 


The development of portable, rapid and sensitive biosensors 

for food safety applications enables point-of-care contamination 

detection and immediate interpretation of results. In this 

research project, our multi-disciplinary team is developing an 

integrated biosensor system on a microfl uidic chip for detecting 

bacteria using immunoassay-based reactions. The device will 

offer a sensitivity to detect 10 2 to 10 3 bacterial cells and an 

assay time of less than 20 minutes per test. Our system will 

yield quantitative data for estimating the number of the target 

bacterium in a food sample. The microfl uidic system under 

development consists of individual devices for cell lysis, lysate 

purifi cation, and immunoassays. We will use an intracellular 

antigen, alcohol acetaldehyde dehydrogenase (Aad), and its 

antibody MAb-H7 to detect Listeria monocytogenes. In order 

to concentrate L. monocytogenes cells from food samples, 

we will fabricate magnetic nanobars with different sizes and 

geometries and develop protocols for immobilizing antibodies 

specifi c to L. monocytogenes on the surface. 

A portable, reusable, and low-cost device will be useful for 

bacterial testing in food manufacturing laboratories. Carrying 

out bacteria detection tests within the food manufacturing 

lab will decrease turnaround time for the results and avoid 

potential growth/decrease/contamination of bacteria during 

transit. Large, expensive equipment is often needed to perform 

conventional analytical methods. Lab-on-a-chip approaches, 

however, allow sophisticated functions of analytical techniques 

to be miniaturized on a microchip. With this technology, it 

will be possible to perform bacteria detection on a microchip 

using only basic laboratory equipment. This technology can 

signifi cantly benefi t the food industry by enhancing the food 

safety testing capability for food manufacturers, food testing 

laboratories, and public health and governmental agencies. 


• Fabricate magnetic nanobars with different sizes and 

geometries and develop protocols for immobilizing 

antibodies specifi c to L. monocytogenes on the surface. 

The amount of bacterial cells bound to the surface 

will be characterized under different conditions. 

• Develop electrophoresis-based immunoassay coupled 

with laser-induced fl uorescence on a microfl uidic 

chip. We will use this tool to quantitatively detect L. 

monocytogenes based on cell lysate, via the interaction 

between alcohol acetaldehyde dehydrogenase 

(Aad) and its monoclonal antibody (MAb-H7). 

• Demonstrate a prototype-integrated microfl uidic 

system that incorporates different steps such as 

manipulation of magnetic nanobars, cell lysis, 

lysate purifi cation, and immunoassay. 


We have created microfl uidic devices for concentration 

and lysis of bacterial cells in a continuous mode based on 

applying microspheres. Microspheres are accumulated in a 

microfl uidic channel, and the gaps between the microspheres 

retain bacterial cells. The electrical lysis can be carried out 

by applying an electrical pulse after cell concentration. This 

technology will allow incorporation of solid phase-based 

detection assays (e.g. ELISA) for bacterial identifi cation by 

functionalizing the microsphere surface. With concentration, 

lysis, and detection capabilities contained in one miniaturized 

device, we can produce a biosensor system that is both robust 

and low cost. 

“We have created microfl uidic devices for concentration and lysis of bacterial 

cells in a continuous mode based on applying microspheres.” 

11 


Ni 

vens 

ns, Fran 

klin 

and 

Corvala 

lan 

Continuous monitoring of chemical agents in aqueous 

media using bioreporter-based sensors 

Investigators: David Nivens (Department of Food Science), Michael Franklin (Center for Biofilm Engineering, 

Montana State University), Carlos Corvalan (Department of Food Science) 


The development of inexpensive sensors with analytical 

attributes to detect product tampering or adulterations from 

chemical contaminants would facilitate our nation’s ability 

to protect its food supply and minimize damage during a 

chemical contamination event (inherent or intentional). The 

bioreporter-based chemical sensors consist of genetically 

programmed cells (bioreporters), a porous microenvironment 

containing the bioreporters, an enclosure that houses the 

microenvironment, and a detection/communication system for 

on-line detection capabilities. With the successful development 

of this technology, it is anticipated that the bioreporter-based 

chemical sensors will have the analytical attributes required to 

fi ll a critical need in the food industry. Besides being potentially 

inexpensive, these biosensors are being developed to: 1) detect 

a hazardous agent(s) typically below immediately dangerous 

to health or life (IDHL) limits; 2) minimize false positives and 

negatives; 3) have a typical response time of less than one 

hour; and 4) be used in fi eld environments by personnel with 

minimal to no training. We envision that the sensors will be 

used with standard food defense practices to further facilitate a 

safe food supply. 

• Combine the most successful strategies of 

objectives 1 and 2 to enhance selectivity (reducing 

false positives) for constructing novel bioreporters 

with optimal analytical performance for point-ofuse 

and long-term monitoring experiments. 

• Model the systems to improve all aspects of analytical 

performance and develop application-specifi c 

biosensors for food and agriculture systems. 


Our bioreporter-based chemical sensor technology used to 

detect a sodium arsenite spiked in water, milk, and orange 

juice samples demonstrates that our sensor technology can 

be used to detect food contaminants in a complex food matrix 

with minimal or no sample preparation. Arsenite levels of ten 

parts per billion in undiluted milk matrices were detected in 

two hours (response time is inversely proportional to analyte 

concentration). This technology could potentially be used by 

minimally trained personnel in numerous food and agriculture 

environments at or below immediately dangerous to health or 

life (IDHL) limits. 


• Develop a dual-signaling bioreporter that minimizes false 

negatives by using bioluminescence as an indicator 

for viability and physiological status and a second 

fl uorescence signal for hazardous chemical detection. 

• Develop a microenvironment that contains 

a programmable biofi lm and slow-release 

nutrients to increase the stability and 

extend the lifetime of the biosensor. 

12 


“The development of inexpensive sensors with analytical attributes to detect product 

tampering or adulterations from chemical contaminants would facilitate our nation’s ability 

to protect its food supply and minimize damage during a chemical contamination event.”

Scientific Publications and Presentations 

Peer Reviewed Journal Publications (2006-2007) 

• Bae, E., Banada, P.P., Huff, K., Bhunia, A.K., Robinson, J.P., Hirleman, E.D. Bio-physical modeling of forward scattering from 

bacterial colonies using scalar diffraction theory. Applied Optics. v. 46 (17) p. 3639-3648 (2007). 

• Banada, P.P., Guo, S., Bayraktar, B., Bae, E., Rajwa, B., Robinson, J.P., Hirleman, E.D., Bhunia, A.K. Optical forward scattering for 

colony identifi cation and differentiation of Listeria species. Biosensors and Bioelectronics. v. 22 (8). p.1664–1671 (2007). 

• Banerjee, P., Morgan, M.T., Rickus, J.L., Ragheb, K., Corvalan, C., Robinson, J.P., Bhunia, A.K. Hybridoma Ped-2E9 cells cultured 

under modifi ed conditions can sensitively detect Listeria monocytogenes and Bacillus cereus. Applied Microbiology and 

Biotechnology. v. 73. p. 1423-1434 (2007). 

• Bayraktar, B., Banada, P.P., Hirleman, E.D., Bhunia, A.K., Robinson, J.P., Rajwa, B. Feature extraction from light-scatter patterns of 

Listeria colonies for identifi cation and classifi cation. Journal of Biomedical Optics. v. 11 (3) p. 034006 (2006). 

• Bhunia, A.K., Banada, P.P., Banerjee, P., Valadez, A., Hirleman, E.D. Light scattering, fi ber optic and cell-based sensors for 

sensitive detection of foodborne pathogens. Journal of Rapid Methods and Automation in Microbiology. v. 15 (2) p.121–145 

(2007). 

• Geng, T., Uknalis, J., Tu, S.-I., Bhunia, A.K. Fiber optic biosensor employing Alexa-Fluor conjugated antibody for detection of 

Escherichia coli O157:H7 from ground beef in four hours. Sensors. v. 6. p. 796-807 (2006) 

• Geng, T., Hahm, B.K., Bhunia, A.K. Selective enrichment media affect the antibody-based detection of stress-exposed Listeria 

monocytogenes due to differential expression of antibody-reactive antigens identifi ed by protein sequencing. Journal of Food 

Protection. v. 69. p. 1879-1886 (2006). 

• Kim, G., Morgan, M.T., Ess, D.R., Hahm, B.K., Kothapalli,A., Valadez, A., Bhunia, A.K. Detection of Listeria monocytogenes using an 

automated fi ber-optic biosensor: Raptor. Key Engineering Materials. v. 321-323. p. 1168-1171 (2006). 

• Kim, K.P., Hahm, B.K., Bhunia, A.K. The 2-cys peroxiredoxin defi cient Listeria monocytogenes displays impaired growth and 

survival in the presence of hydrogen peroxide in vitro but not in mouse organs. Current Microbiology. v. 54. p. 382-387 

(2007). 

• Leake, L.L. Advancing rapid microbial testing. Food Technology. v.60. p. 68-72 (2006) 

• Lathrop. A., Huff, K., Bhunia, A.K. Prevalence of antibodies reactive to pathogenic and nonpathogenic bacteria in preimmune 

serum of New Zealand white rabbits. Journal of Immunoassay and Immunochemistry. v. 27. p. 351-361 (2006). 

• Morgan, M.T., Kim, G., Ess, D.R., Kothapalli, A., Hahm, B.K., Bhunia, A.K. Binding inhibition assay using fi ber-optic based biosensor 

for the detection of foodborne pathogens. Key Engineering Materials. v. 321-323. p. 1145-1150 (2006). 

• Sage, L. Detecting pathogens on produce. Analytical Chemistry. pp 7-9. (2007) 

• Nanduri, V., Kim, G., Morgan, M.T., Ess, D. Hahm, B.-K., Kothapalli, A., Valadez, A.,Geng, T., Bhunia, A.K. Antibody immobilization 

on waveguides using a fl ow–through system show improved Listeria monocytogenes detection in an automated fi ber optic 

biosensor: RAPTOR. Sensors. v. 6. p. 808-822 (2006). 

• Nanduri, V., Bhunia, A.K., Tu, S.-I., Paoli, G.C., Brewster, J.D. SPR biosensor for the detection of L. monocytogenes using phage 

displayed antibody. Biosensors & Bioelectronics. 2007 (in press) 

• Sun, L., Yu, C., Irudayaraj, J. Surface enhanced raman scattering based nonfl uorescent probes for multiplex DNA detection. 

Analytical Chemistry, (accelerated publication, available online, Apr 27, 2007). 

• Wang, H.Y., Bhunia, A.K., Lu, C. A microfl uidic fl ow-through device for high throughput electrical lysis of bacterial cells based on 

continuous DC voltage. Biosensors & Bioelectronics. v. 22 p. 582-588 (2006). 

“Our collaboration with USDA-ARS Eastern Regional Research 

Laboratory continues to breed success and has led to 21 peer-reviewed 

research publications and 22 presentations and proceedings this year.” 

13 


Scientific Publications and Presentations 

Abstracts for major papers/posters presented (2006-2007) 

• Banada, P.P., Bernas, T., Robinson, J.P., Bhunia, A.K. Proteomic analysis and identifi cation of cytotoxic factors from Bacillus 

cereus. P-097, 107th American Society for Microbiology (ASM) Annual Meeting. May 21-25, Toronto, Canada pp 206., 2007. 

• Gross, B. D., Reed, T.S., Stoklosa, A.D., Schroeder, D.L., Nagel, A.C., Co, B., Nivens, D.E. Remote sensing of agricultural contaminants 

using biosensor networks over TCP/IP. 233rd Annual Meeting of the American Chemical Society. Presented April 23, 2007. 

• Hahm, B.K., Kim, H., Bhunia, A.K. Enrichment and detection of Escherichia coli O157:H7, Salmonella Typhimurium and Listeria 

monocytogenes using a sample preparation device, PEDD (Pathogen Enrichment and Detection Device). 107th American 

Society for Microbiology (ASM) Annual Meeting. May 21-25, Toronto, Canada. 2007. 

• Irudayaraj, J. No-glow tags for Raman Spectroscopy. Analytical Chemistry News. June 1, 2007 (page 3961). 

• Nagel, A.C., Mauer, L.J. , Stoklosa, A.D., Franklin, M.J., Nivens, D.E. Quantifi cation of cellular biomass and extracellular alginate 

from Pseudomonas aeruginosa biofi lms using fourier transform infrared (FT-IR) spectroscopy. American Society for 

Microbiology, 107th Annual General Meeting. 2007. 

• Schroeder, D.L. , Nagel, A.C., Gross, B.D., Reed, T.S., Nivens, D.E. Bioreporter-based chemical sensor of arsenic in agricultural 

samples. 233rd Annual Meeting of the American Chemical Society. Presented April 23, 2007. 

• Valadez, A.M., Tu, S.-I. Tu, Bhunia, A.K. Development of a fi ber-optic waveguide biosensor assay for the detection of Salmonella 

Enteritidis from poultry products. Institute of Food Technologist. Poster Abstract No. 098-31, 2007. 

Thesis/Dissertations (2006-2007) 

• Bae, E. Analysis and characterization of multi-scale scattering : Application to bacterial colonies. Ph.D. Dissertation. Dec 

2006. Purdue University. 148 p. 

• Kim, H.A. Selective enrichment medium for simultaneous growth and detection of Escherichia coli O157:H7, Listeria 

monocytogenes, and Salmonella Enteritidis from food. 2007. Purdue University. p. 126 

• Valadez, A.M. Development of a fi ber-optic waveguide biosensor assay for the detection of Salmonella Enteritidis in food. 

M.S. Thesis 2006. Purdue University. 105 p. 

Books and Book Chapters (2006-2007) 

• Geng, T., Bhunia, A.K. 2007. Optical biosensors in foodborne pathogen detection. Smart Biosensor Technology. CRC Press, 

(Editors: Knopf and Bassi), Taylor and Francis group, Boca Raton, FL p. 505-519. 

• Kizil, R., Irudayaraj, J. 2007. FT-Raman spectroscopy for food and biomaterial characterization. Nondestructive sensing for 

food quality. IFT Press, (Editors. Irudayaraj and Christoph), Blackwell Publishing Professional, Ames, IA 50014. 

14 


“At the Center for Food Safety Engineering, we direct our

Invited Lectures and Seminars (2006-2007) 

• Cheng, Z.-Y. Electroactive polymers for artifi cial muscles and biosensors. 3M Science and Engineering Faculty Day, St. Paul, 

Minnesota, USA, June 21, 2006. 

• Cheng, Z.-Y., Chin, B.A. A radically new approach to rapidly detect biological threat agent. SPIE Defense & Security, April, 

Orlando, Florida, USA, 2006. 

• Fu, L., Li, S., Zhang, K., Cheng, Z.-Y. A Novel Method for Monitoring Biological Quality of Surface Waters. 20th Annual Alabama 

Water Resource Conference, Orange Beach, Alabama USA, Sept. 7-8, 2006. — This poster was selected as the Best 

Student Poster (1st Place). 

• Li, S., Fu, L., Wang, C., Lea, S., Engelhard, M., Arey, B., Cheng, Z.-Y. Microstructure and composition of Fe-B Nanobars. Materials 

Research Society Fall Meeting, Boston, Massachusetts, USA, 2006. 

• Li, S.Q., Orona, L., Fu, L.L., Cheng, Z.-Y. Development of amorphous Fe-B nanorods and nanoarray as magnetostrictive acoustic 

resonator for sensor applications. US ONR Workshop on Transduction Materials and Devices, State College, Pennsylvania, 

May 10, 2006. 

Articles in Popular Press (2006-2007) 

• Advancing rapid microbial testing. Food Technology. 60 (9) p. 69-72. (2006) 

• Pinpointing bacteria. R&D Magazine. 48 (8) p. 68. (2006). 

• Detecting pathogens on produce. Analytical Chemistry. p. 7-9. (2007). 

Patents Granted (2006-2007) 

• 64803.P1.US - Multiplex Biosensor Using Gold Nanostructures. Provisional patent application fi led on February 23, 2007. 

efforts toward detecting problems and protecting consumers.” 

15 


Center Staff 

Dr. Richard H. Linton 

Director 

765.494.6481 

linton@purdue.edu 

Staff 

Kevin T. Hamstra, Multimedia Technical Specialist 

• 765.496.3833 • khamstra@purdue.edu 

Kiya A. Smith, Center Coordinator 

• 765.496.3827 • kiya@purdue.edu 

Dr. W. R. (Randy) Woodson, Dean of Agriculture 

• 765.494.8391 • woodson@purdue.edu 

Dr. Shu-I Tu 

Supervisory Research Chemist 

215.233.6466 

shui.tu@ars.usda.gov 

USDA-ARS 

Dr. Jeffrey Brewster • 215.233.6447 • jeffrey.brewster@ars.usda.gov 

Dr. Pina Fratamico • 215.233.6525 • pina.fratamico@ars.usda.gov 

Dr. Andrew Gehring • 215.233.6491 • andrew.gehring@ars.usda.gov 

Dr. Peter Irwin • 215.233.6420 • peter.irwin@ars.usda.gov 

Dr. James A. Lindsay • 301.504.4674 • james.lindsay@ars.usda.gov 

Dr. John B. Luchansky • 215.233.6620 • john.luchansky@ars.usda.gov 

Dr. George Paoli • 215.233.6671 • george.paoli@ars.usda.gov 

Dr. Gary Richards • 302.857.6419 • gary.richards@ars.usda.gov 

Dr. Christopher Sommers • 215.836.3754 • chris.sommers@ars.usda.gov 

Dr. Howard Zhang • 215.233.6583 • howard.zhang@ars.usda.gov 

Dr. Arun K. Bhunia 

PI 

765.494.5443 

bhunia@purdue.edu 

Co-PIs 

Andrew Gehring • agehring@errc.ars.usda.gov 

E. Daniel Hirleman • 765.494.5688 • hirleman@purdue.edu 

Mark Morgan • 765.494.1180 • mmorgan@purdue.edu 

J. Paul Robinson • 765.494.6449 • jpr@fl owcyt.cyto.purdue.edu 

Shu-I-Tu • 215.233.6466 • stu@errc.ars.usda.gov 

Staff / Graduate Students 

Euiwon Bae • 765.494.4762 • ebae@purdue.edu 

Padmapriya Banda • 765.496.3826 • pbanada@purdue.edu 

B.K. Hahm • 765.496.7356 • hahm@purdue.edu 

Karleigh Huff • khuff@purdue.edu 

Hyochin Kim • 765.496.7354 • kim399@purdue.edu 

Viswaprakash Nanduri • vnanduri@purdue.edu 

Bartek Rajwa • 765.494.0757 • rajwa@fl owcyt.cyto.purdue.edu 

Angela Valadez • 765.496.3824 

Dr. Maribeth A. Cousin 

PI 

765.494.8287 

cousin@purdue.edu 

Co-PI 

Charles P. Woloshuk • 765.494.3450 • woloshuk@purdue.edu 


Yenny Suanthie • ysuanthi@purdue.edu 

Janaka Morandage • jmoranpa@purdue.edu 

16 


Dr. Joseph Irudayaraj 

PI 

765.494.0388 

josephi@purdue.edu 

Staff / Graduate Student 

Lan Sun • sun22@purdue.edu 

Dr. Michael Ladisch 

PI 

765.494.7022 

ladisch@purdue.edu 

Co-PIs 

Padmapriya Banada • 765.496.3826 • pbanada@purdue.edu 

Rashid Bashir • basher@purdue.edu 

Arun Bhunia • 765.494.5443 • bhunia@purdue.edu 

Young-mi Kim • kim107@purdue.edu 

Xingya Liu • 765.494.7052 • xingya@purdue.edu 

Nathan Mosier • 765.494.7025 • mosiern@purdue.edu 

J. Paul Robinson • 765.494.6449 • jpr@fl owcyt.cyto.purdue.edu 

Andres Rodriguez • arodrigu@ecn.purdue.edu 

Miroslav Sedlak • 765.494.3699 • sedlak@purdue.edu 


Winnie Chen • chen52@purdue.edu 

Bala Jagadeesan • bjagadee@purdue.edu 

Yi-Shao Liu • liu63@purdue.edu 

Peter McKinnis • pmckinni@purdue.edu 

Heyjin Park • 765.434.7031 • heyjincpark@gmail.com 

Jaeho Shin • shin0@purdue.edu 

David Sung • ysung@purdu.edu 

Angela Valadez • 765.496.3824 • valadez@purdue.edu 

Shuaib Salamat • ssalmat@purdue.edu 

Dr. Chang Lu 

PI 

765.494.1188 

changlu@purdue.edu 

Co-PIs 

Ning Bao • nbao@purdue.edu 

Arun Bhunia • 765.494.5443 • bhunia@purdue.edu 

Zhongyang Cheng • chengzh@auburn.edu 


Hsiang-Yu Wang • hwang@purdue.edu 

Balamurugan Jagadessan • bala@purdue.edu 

Peixuan Wu • wupeixu@auburn.edu 

Suiqiong Li • lisuiqi@auburn.edu 

Dr. David E. Nivens 

PI 

765.494.0460 

dnivens@purdue.edu 

Co-PIs 

Carlos Corvalan • 765.494.8262 • corvalac@purdue.edu 

Michael Franklin • 406.994.5658 • umbfm@montana.edu 


Claudia Ionita • 765.496.7354 • cionita@purdue.edu 

Aaron Nagel • 765.496.3832 • acnagel@purdue.edu 

Xinyu Shen • 765.496.3825 • shenx@purdue.edu 

17 


It is the policy of the Purdue University Center for Food Safety Engineering, that all persons shall have equal 

opportunity and access to the programs and facilities without regard to race, color, sex, religion, national origin, 

age, marital status, parental status, sexual orientation or disability. 

Purdue University is an Affirmative Action employer. 

The Center for Food Safety Engineering 

Purdue University 

Food Science Building 

745 Agriculture Mall Drive 

West Lafayette, IN 47909 

Non-profit 

Organization 

U. Postage 

PAID 

Purdue 

University

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