Chris Elliott (+ many many others…),

Novel approaches to screening for food contaminants

Broad Spectrum Testing for Compounds

The requirement to test for more chemical contaminants in food in a shorter period of time has become a necessity within routine laboratories.

How can we achieve this?

Broad Spectrum Contaminant Analysis

In their wise & far sighted approach to the subject the European Commission published in the FP6 Food Quality Work Programme:

“New methods to prevent & monitor occurrence of multiple contaminants such as pesticides, toxins, drugs, & EDs in foods.

This should be achieved through use of advanced sample preparation techniques & emerging biotechnological screening approaches based on development of novel biomarkers”

Question…….

How can such a radical set of requirements be met?

The birth of the Biocop consortium……….

The Biocop principles:

The fundamental scientific objective is to utilise emerging life science technologies to create a powerful new approach to detect & control chemical contaminants in foods.

Biocop: New Technologies to Screen Multiple Chemical Contaminants in Foods

Objectives:

Development of novel screening methods to detect multiple chemical contaminants in foods

Training of scientists in developed technologies

Widespread dissemination of project information

Project Architects

Hanspeter Naegli: Switzerland

Aldert Bergwerff: The Netherlands

Timo Lovgren: Finland

Esa Stenberg: Sweden

Han van Egmond: The Netherlands

Luis Botana: Spain

Matthew Sharman: United Kingdom

Michel Nielen: The Netherlands

Jana HajŠlová: Czech Republic

Edwin de Pauw: Belgium

Guiseppe Palleschi: Italy

Project Timelines

Written & submitted : May 2002 (1156 in food)

Shortlisted by EC : October 2002 (53 in food)

Appeared in FP6: Food Quality & Safety Work programme: December 2002

Specific call appeared in November 2003

The Current Status of the The Current Status of the BioCop BioCop ProposalProposal

Submitted February 2004

First phase of evaluation: Passed as top proposal

Second Phase evaluation: Passed as top proposal

Contract negotiations commenced with the Commission June 2004

The PlanThe Plan……....

Completion of contracts October 2004

Initiation of five year project from January 2005

33 partners from 16 countries

Budget 15M EURO (9.6M for European Commission)

BiocopBiocop: The Science: The Science

Veterinary drugs, heavy

metals, mycotoxins, EDs

shellfish toxins, pesticides,

Transcriptomics,

Proteomics, Molecular

Engineering, Biosensors

WP5 Shellfish Toxins

WP 10 Mycotoxins

WP6 Pesticides

WP8 Therapetics

WP7 Endocrine Disrupters

WP9 Heavy metals

WP1 WP2 WP3 WP4 Binding

Molecules

T’omics Proteomics Biosensors

WP11 Consumer Sci &

Dissemination

The The Biocop Biocop MatrixMatrix

Transcriptomics Transcriptomics (1)(1)Identification of chemical residues through genomic fingerprints

Exposure of Target Cells

cDNA

Total RNA isolation

In vitro

transcription

*biotin-labeled cRNA

Hybridization to the

Affymetrix®U133 GeneChip

(sequences of 33‘000 human genes)

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SAPE Staining

Scanning &

Data Analysis

Reverse transcription

TranscriptomicsTranscriptomics (2)(2)

The selected genes will then be employed to print tailored DNA microchips for the development of a low cost high-throughputmicroarray platform.

0 5

10

15

20

Induction

SFRS2

FLJ11838

JAK1

HNRPH3

MCM2

MELK

ANLN

HELLS

FLJ10055

SDF1

PIGA

DUT

CXCL12

IER3

HCAP-G

H11

SFXN2

GJA1

CTSD

FKBP4

OLFM1

SERPINA6

IGFBP4

MYB

HNOEL-iso

PTPLA

HSPC154

IGSF4

CDC6

NASP

RRM2

UHRF1

KIAA0101

ACAT2

MYBL

TYMS

TSSC3

AMD1

ARL3

FLJ10540

RAMP

genis

tein

bis

phenol A

Comparison betw

een bisphenol A, genistein and estradiol

Transcriptom

icsTranscriptom

ics(3)(3)

Proteomics Proteomics (1)(1)

Identification of candidate biomarkers

Development of methodology to screen animals for the presence of identified biomarkers

Selection of most appropriate biomarkers

Fingerprints reporting contamination of food-producing animals

catabolism ⇔ anabolismcatabolism ⇔ anabolismcatabolism ⇔ anabolismcatabolism ⇔ anabolism

Identification of alteration of protein expression using a wide range of proteomic tecnhiques

ProteomicsProteomics (2)(2)

•Species / race

•Zootechnical aspects

•Health

•Age

•Gender

•Feed regime

•Oestrous cycle

•Gestation

•Stress

•Concentration of the target substance

•…….

ProteomicsProteomics (3)(3)

Confusing signals…..

ProteomicsProteomics (4)(4)

Antibody production to selected biomarkers

Source existing mabs Produce new panels of mabs

Gene ManipulationSource existing enzymes

DNA databases Receptor Expression

Assay Developers

Production of Unique Binding ProteinsProduction of Unique Binding Proteins

From a very compound specific antibody to a broad spectrum binding protein.

R&D work based at University of Turku

Antibody Engineering

A wide range of existing mabs available to the consortium. These will undergo engineering

The Fusion Technology PlatformThe Fusion Technology Platform

High-throughput

Platform

Receptor

Modelling

Fragments

cloned with

fusion tags

Over-express in

E. coli

solubility

Scale-up

low

pressure

purification

Incorporation

into assay

www.fusionantibodies.co.uk

BiosensorsBiosensors

Optical Optical ElectrochemicalElectrochemical

⇒⇒⇒⇒⇒⇒⇒⇒ High throughputHigh throughput

⇒⇒⇒⇒⇒⇒⇒⇒ Technology couplingTechnology coupling⇒⇒⇒⇒⇒⇒⇒⇒ Low cost Portable devicesLow cost Portable devices

Biosensors(2)Biosensors(2)

Biosensors(3)Biosensors(3)

Identification & Quantification

Sample injection

Measurable molecular interactions

Immobilised receptor on chip surface

No response, no MS analysis

Biosensor Coupled to Mass Spectrometry

Phase 1: Research, e.g.

Development of binders, surface chemistries, chip production,

equipment design

Phase 2: Research, e.g.

Development of assays, technology coupling, sample preparation

Phase 3: Comparison and validation

Compare new assays with state of the art & fully validate

Phase 4: Demonstration

Technology transfer events, e.g. Workshops

YEAR

1

2

3

4

5

Decision Points Decision Points

Decision Points Decision Points

Decision Points Decision Points

BioCop BioCop PhasesPhases

BiocopBiocop……....

We will try to keep you informed…..

Keep looking out for information

The skies the limit…..