1. Cell by Date Bio-Indicator for Meat Spoilage

2. Introduction

3. Introduction

4. Introduction

5. Problem with Food Industry PROBLEM: Printed sell-by date is based on predictive challenge testing PROBLEM: Printed sell-by date is based on predictive challenge testing –Not reflective of product exposure to environment –Printed Sell By Date is unreliable SOLUTION: Monitor level of exposure and report it SOLUTION: Monitor level of exposure and report it – Tell Consumer when meat is not fit for consumption SpecificationDesignModellingImplementationTesting/Validation

6. Motivations Fresh meat is a highly perishable food Fresh meat is a highly perishable food Survey studies show temperature conditions >10°C are usual during transportation, retail storage, and consumer handling. Survey studies show temperature conditions >10°C are usual during transportation, retail storage, and consumer handling. 1/3 of Adults use spoilt food 1/3 of Adults use spoilt food Colouring and packaging methods are deceptive Colouring and packaging methods are deceptive Win-win situation for producers-consumers Win-win situation for producers-consumers Recycling of old projects – pH, temperature, BFP Recycling of old projects – pH, temperature, BFP SpecificationDesignModellingImplementationTesting/Validation

7. Specification 1. Report When food is ‘off’ –Report cumulative Temperature exposure of food Temp is largest factor in spoiling Temp is largest factor in spoiling Easiest way to report if food is bad Easiest way to report if food is bad 2. Easy To Use – Visual Reporter – Wow ! factor 3. Improve Accuracy of other attempts: – SSM, Intelligent Cooling, Antimicrobial Only Threshold devices Only Threshold devices 4. System must be stable for length of shelf-life 5. No contact with food – Need to obey HSA standards SpecificationDesignModellingImplementationTesting/Validation

8. 1. Report Cumulative Temperature exposure of food  Need memory to do this / Integrator fct 2. Visual Reporter  Needs to have long degradation time 3. Improve Accuracy  Limited Crosstalk with chassis  Easily tuneable threshold 4. Stable for length of Shelf Life 5. HSA standards  Project must not contaminate food Considerations/Problems SpecificationDesignModellingImplementationTesting/Validation

9. Solutions 1. Integrator 3. Improved Accuracy 4. Stability 5. HSA standards 2. Long Degradation Time –Firefly FP SpecificationDesignModellingImplementationTesting/Validation Cell Free System

10. Cell-free system Cell-free system –Widely-used in vitro tool used to study biological reactions in vitroin vitro –Less of the complex interactions cf. whole cell. –Provides molecular machinery that can be used in reactions with the absence of many cellular components. Solutions SpecificationDesignModellingImplementationTesting/Validation

11. Solutions 1. Integrator 3. Improved Accuracy 4. Stability 5. HSA standards CFS increases stability due to lack of degrading enzymes CFS increases stability due to lack of degrading enzymes CFS stable for up to 4 days CFS stable for up to 4 days CFS agrees with HSA standards CFS agrees with HSA standards SpecificationDesignModellingImplementationTesting/Validation Cell Free System

12. Design SpecificationDesignModellingImplementationTesting/Validation Photinus luciferin + O 2 + ATP oxidized Photinus luciferin + CO 2 + AMP + diphosphate + light ATPAMPATPAMP PcPc RBS Te Luciferase Temp. Increase Luciferin O2O2 Cell-free extract medium LIGHT!!!

13. Modelling Concept of predictive microbiology Concept of predictive microbiology –Involves knowledge of microbial growth responses to environmental factors –Expressed in quantitative terms using mathematical equations –Data used to interpret the effect of varying conditions on microbial growth. Integrative Temp function of product Integrative Temp function of product –Leads to “intelligent” product management systems –Optimization of food quality and safety at the time of consumption SpecificationDesignModellingImplementationTesting/Validation

14. Implementation SpecificationDesignModellingImplementationTesting/Validation Bio indicator Colour scheme Simplest System : Phase 1 GoodOK BAD

15. Implementation SpecificationDesignModellingImplementationTesting/Validation Simplest System : Phase 2 Discrete Change In addition Phase 1 Good Bad Eg. Discrete Change At 25oC for more than 2 hours

16. Testing Focus on proof of concept Focus on proof of concept –Period of 24 hours as opposed to 4 days –Regulated luciferase synthesis that corresponds to predicted outcome –Light intensity as a measurement (compare with colour scheme strip) SpecificationDesignModellingImplementationTesting/Validation

17. Time line Week 3-4 – Modelling + GeneART Week 3-4 – Modelling + GeneART Week 4-6 – Experiment implementation Week 4-6 – Experiment implementation Week 7-8 – Testing / Validation Week 7-8 – Testing / Validation Week 9-10 – Polish the project + documentation Week 9-10 – Polish the project + documentation

18. Conclusion Cell by date: Cell by date: –Mimics printed ‘sell by date’ with additional temperature exposure tracking –More accurate than conventional method –Will improve consumer health –Focuses on use of experimentation & proof of concept

19. End Questions? Questions?