Technological advances and new tools for food safety control
Katrina Campbell Lecturer in Bioanalytical Systems
2nd International Symposium on Innovation in marine products and food industry Vigo, 15th September 2014
Global Food Safety System Food Safety has always been essential for existence
Supply versus
demand
Impact: Biological
Economical Social
Political
Farm to Fork Under the current EU Food Hygiene legislation Producing safe food is
the responsibility of Food Business Operators (FBOs)
Hazard Analysis and Critical Control Points (HACCPs) checks
• Source of raw materials • Production site • Processing sites and distribution • End product testing
FOOD CONTAMINANT TESTING IS MAINLY ONLY PERFORMED IF
LEGISLATIVELY REQUIRED AND IF SUITABLE METHODS ARE AVAILABLE
What is there in food worth measuring? Basic components (protein, carbohydrate, fibre fats etc)
Additives (vitamins, colourants, stabilizers, nanoparticles)
Contaminants
Chemical Microbiological Process Natural Man made Bacterial / Viral
Heat Contact
RASFF Contaminants
Why food safety analysis?
Measuring Food Contaminants Verify the presence or absence of contaminant in a sample:
Qualitative Test
Quantify the amount of contaminant in a sample: Quantitative Test
An analytical method which will give a strong indication that there is some form of chemical contaminant present
Screenings tests
Confirmatory tests An analytical method which gives unequivocal proof that there is some form of chemical contaminant present
1. Functional Assays a. Animal assays b. Cell based c. Receptor based d. Enzyme based e. Fluorescence based
2. Biochemical Assays a. ELISA b. Lateral flow devices c. Biosensor
Level of contaminant measured is relative to the biological effect of the sample
May detect new toxic analogues Contaminant identification is not unequivocal Technology transfer of methods is difficult
Binder assays Toxicity does not always correlate with binder
cross-reactivity Sample preparation and data analysis is fast Screening tools for HACCP management and
rapid response
Methods applied to food analysis
4. Analytical methods a. HPLC b. LC-MS c. GC-MS d. ICP-MS
Contaminants can only be identified and quantified for available analytical standards
Toxicity equivalent factors must be applied Sample clean-up is extensive with oxidation
steps being required in cases Data analysis is laborious LC-MS is unequivocal for identification
3. Spectroscopic methods a. Near IR b. Mid IR c. RAMAN d. SERS
Fingerprinting techniques Non-destructive methods little to no sample prep Require chemometric models of known samples Sensitivity is questionable
Methods applied to food analysis
ANALYTICAL METHODS TRADITIONAL CONFIRMATORY METHODS
Screening analysis by animal assays, functional methods,
immunochemical
Selected samples taken from farms (blood) and slaughterhouses (meat
and urine) and feedmills (feed), production sites (various foods)
Monitoring plan for food analysis
Compliant result No further action -VE
Confirmatory analysis and quantification by
physiochemical methods usually LC-MS
+VE
BELOW
Non compliant result Action required
ABOVE
Screening Tests for Food Analysis Animal Based Cell Based Receptor Based Antibody Based
Antibiotics residues in milk
Dioxins in feed & food
Chemical contaminants in foods
Toxins in food Botulism
Marine toxins
Screening tests that require special facilities for use
Low molecular
weight Contaminant
Antibody Production
Protein Conjugation
Protein carrier
Immunisation Programme Polyclonal
antibody production
Titre Specificity
Major Problems:
Toxicity
Source of contaminant
Specificity
Antibodies specific for a desired antigen can be conjugated with a radiolabel, fluorescent label, or colour-forming enzyme & are used as a "probe" for detection.
Immunological methods for food analysis
1960: Radioimmunoassay was first described in a scientific paper by Rosalyn Sussman Yalow and Solomon Berson published in 1960 1971: Peter Perlmann and Eva Engvall at Stockholm University invented ELISA 1975: Generation of the 1st monoclonal antibodies (George Kohler & Cesar Milstein)
Lateral Flow Tests Rapid & simple
Antibody Coated Competitive ELISA
Antibody coated
No toxin in sample Antibody captures labelled toxin
High response
Toxin in sample Antibody captures toxin
Wash step removes labelled toxin Low response
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Antigen Coated Competitive ELISA
Toxin protein conjugate (TPC)
No toxin in sample Antibody binds to TPC
Labelled antibody binds to antibody High response
Toxin in sample Antibody binds to toxin
Wash step removes antibody Low response
Screening Tests for Food Analysis Rapid Reliable Low cost Low false positives No false negatives Safe
Speed – higher throughput Cost Portability Multiplexing Remote or in situ sensing Requirements of regulators or industry End product testing for release systems
Food is produced on an ever-increasing scale
To enhance and ensure GLOBAL FOOD SECURITY
new biosensor technologies and applications are needed
Antibody Based
Chemical contaminants in foods
Biosensors
http://packagingtech.net/34-biosensor-technology-for-food-packaging-applications.html
“A biosensor is an analytical device incorporating a
biological or biologically derived sensing element either intimately associated with or
integrated within a physicochemical transducer. The usual aim is to produce a digital
electronic signal which is proportional to the
concentration of a specific analyte or group of analytes”
Turner, A.P.F., Karube, I. and Wilson, G.S. (1987). Biosensors: Fundamentals and Applications. Oxford University Press, Oxford.
Biacore’s methods: Vitamin analysis for fortified foods Antibiotics in honey and screening for veterinary drugs Mycotoxin analysis in cereals and feeds Marine toxins: Domoic, okadiac and DTXs, PSPs, TTX, BTX Freshwater toxins: Microcystins and cylindrospermospin
SPR biosensor technology
Benefits: Simplicity of operation Minimal sample preparation Interchangeable chips for multiple targets Single channel operation
SPR Biosensor Invented by
Liedberg, Nylander,
Lunström (1983)
Drawbacks: Single target analysis, primarily laboratory based and relatively expensive
The use of multiplexing surface plasmon resonance
platforms is a major research focus at IGFS.
• 3 channel, fully automated SPR system • Control and data analysis software • Temperature controlled reagent racks
Nomadics SensiQ Platform
SPR biosensor technology
Individual SPR assays for DON & NIV and T2 &HT2 combined as a proof of concept
Meneely, J. P., Fodey, T., Armstrong, L., Sulyok, M., Krska, R., Elliott, C. T., 2010, Rapid surface plasmon resonance immunoassay for the determination of deoxynivalenol in wheat, wheat products, and maize-based baby food, Journal of Ag and Food Chem, 58, 8936-8941. Meneely, J. P., Sulyok, M., Baumgartner, S., Krska, R., Elliott, C. T., 2010, A rapid optical immunoassay for the screening of T-2 and HT-2 toxin in cereals and maize-based baby food, Talanta, 81, 630-636.
•Thermally insulated flow-cell and fluidics • Multiple sample injection options • Process over 300 samples in 24 hours
Multichannel Multiplexing
Sample compartment
Chip dockingcompartment
IFC
Running buffers
and waste
Peristaltic pumps
Injection needles
A 16 channel SPR biosensor
developed as part EU Biocop project
Campbell, K., McGrath, T., Sjölander, S., Hanson, T., Tidare, M., Jansson, O., Moberg, A., Mooney, M., Elliott, C., Buijs, J. (2011) Use of a novel micro-fluidic device to create arrays for multiplex analysis of large and small molecular weight compounds by surface plasmon resonance. Biosensors and Bioelectronics, 26: 3029-3036. McNamee, SE, Elliott, CT, Delahaut, P & Campbell, K 'Multiplex biotoxin surface plasmon resonance method for marine biotoxins in algal and seawater samples 2013 Environmental science and pollution research international, vol 20, no. 10, pp. 6794-807.
Toxin Antibody Titre IC50 (ng/ml) IC20 – IC80 (ng/ml)
Domoic Acid DA-Ab 1/200 2.6 1.0 – 6.4
Okadaic Acid OA-Ab 1/4000 4.9 1.7 – 14.4
Neosaxitoxin NEO-Ab 1/25 2.6 1.1 – 6.0
Saxitoxin STX-Ab 1/1000 1.9 1.0 – 3.7
Assay principle: indirect competitive assay with chemiluminescence detection
Munich MCR3 - Chemiluminescence
MCR3 Instrumentation
Toxin (ng/ml)
Domoic Acid Okadaic Acid Saxitoxin
Toxin IC50
(ng/ml) Saxitoxin 1.3
Domoic Acid 6.5 Okadaic Acid 2.9
Why nanosensors? Smaller and faster Require less power to run Greater sensitivity Better specificity Cost-effective Remote use Simple to use
SPR Biosensor
Bio to nanosensors
High Tech MS analysis
Multi mycotoxin methods Multi pesticide methods Untargeted analysis Fingerprint profiling
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On site or end product testing
Production of nanosensors
Localised SPR analysis - miniaturisation
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Multiplex approaches for emerging concerns
ergotamine
ergocristine
CTRL
Ergot Alkaloid
scopolamine
atropine
CTRL
Tropane Alkaloid
CTRL
Pyrrolizidine Alkaloid
Analysis time = 45mins Detection of 15 PA compounds
Semi-portable multiplexing technology Luminex
Technology Toximet
Technology
Mycotoxin analysis - Aflatoxins
Portable SPR
Sensing Unit: high resolution camera Operation via touch screen Graphics processing unit Analysis Tools: Smart applications Large memories Data transmission via mobile networks
Seattle Sensors Multiple target detection for
remote analysis 4 x 3 channel
0.00%
20.00%
40.00%
60.00%
80.00%
100.00%
120.00%
140.00%
0.01 0.1 1 10
[OA] (ng/ml)
Perc
ent R
ate
of B
indi
ng
Okadaic Acid Detection
Electrochemical Sensors
Environmental or chemical sensors for toxin analysis
Calibration curve for a biosensor prototype with 40 um gold microdisc array
Calibration curve for a biosensor prototype with 10 um gold microband array
Rapid Multiplex Portable Diagnostics
Aim to produce a lower cost platform offering • Low cost analysis • Simplicity in use • Highly specific single target analysis • Multiplexing – multiple target analysis • Bespoke sensitivity • Robust – high performance • Field deployable
Suitable for source to product testing
Up to 100 feature microarray
Planar waveguide with custom surface
chemistry
Multiplex Mbio Diagnostics
Scientific know how is based in the cartridge Different approaches in how this can be designed
• Molecular level – DNA / RNA for pathogen and speciation testing • Protein Level – Allergen testing eg milk, nuts, eggs, seafood • Residual level – Low molecular weight toxins / antibiotics / contaminants
Nanotechnology in Microarrays
Competitive inhibition assay
• Toxins are immobilised on the cartridge surface through nanospotting.
• Sample/antibody are mixed and added to the cartridge.
• Incubation with a secondary labelled antibody .
• Light directed to the bottom of the cartridge allows excitation of the fluorophores of the labelled antibody.
• The sensitive detection of binding events on the cartridge surface is observed.
• When toxin is present there is reduced antibody binding to surface.
Important to implement simple testing regimes to allow FBOs to perform testing
Offer a simple device requiring minimal sample preparation through either simple fluid application (blood, milk, juice) or dissolution of solid foods in buffering reagents
Simplicity in Use
Depending on user Qualitative – YES/NO Answer Quantitative – Concentration
Add Sample to Assay Buffer
Dye-labeled Ab
Transfer to Cartridge
Read at 5-10 min
Biotoxin Assay n IC50 (ng/ml)
Dynamic Range IC20 – IC80 (ng/ml)
Domoic acid Single 2 0.13 0.02 – 1.13
Multi 4 1.86 0.52 – 5.71
Okadaic acid Single 2 0.35 0.12 – 0.68
Multi 4 0.42 0.20 – 0.88
Saxitoxin Single 2 0.04 0.02 – 0.06
Multi 4 0.06 0.02 – 0.16
Microcystin Single 2 1.48 0.52 – 3.71
Multi 4 1.40 0.48 – 3.64
Cylindrospermopsin Single 2 0.23 0.07 – 0.55
Multi 4 0.19 0.08 – 0.52
Aquatic Toxins
Marine and fresh water toxin assay
Antibiotics in milk
Multiplex Test for detecting
antibiotics at MRL values
Synthetic Binders
Recombinant antibodies No use of animals
Phage display technology Synthetic binders from libraries Tolerant solvent and pH
Aptamers for food contaminants
single stranded DNA or RNA oligonucleotides that range in size
from 20-80 bases
Spectroscopic techniques “fingerprinting” technique giving unique spectra Little or no sample preparation Ideal technique for use with adulteration of food eg fats and oils Multivariate techniques can be used to extrapolate the desired chemical information
Image Detector Processor Data Answer
Animal Dog
Alsatian German Shepherd
Speed – higher throughput Simplicity in use Minimal sample preparation Relatively low cost Multiplexing Portability Remote sensing Requirements of regulators or industry End product testing for release systems
Summary for Rapid methods
EU Project Collab4Safety
http://web.spi.pt/collab4safety/collab4safety
Technological advances and new tools for food safety control
Katrina Campbell Lecturer in Bioanalytical Systems
2nd International Symposium on Innovation in marine products and food industry Vigo, 15th September 2014