1. Biosensor Microelectrodes Guido Drago Applied Enzyme Technology Ltd

2. Company History Applied Enzyme Technology Ltd formed in 1994 Smart Awards 1994 & 1995 protein stabilisation technology Patents for dry stabilisation fully granted in 1995 and 1996 Patent for solution stabilisation fully granted in 2000 AET acquired by LRH Ltd. and relocated June 2001 BIOWISE Demonstrator Award 2003 SIF Award 2009 Multiple EU, TSB, Home Office & DEFRA Grants

3. Contract research for the stabilisation of specific proteins Supply Protein Stabilisation Kits for stability troubleshooting Supply of bulk stabiliser formulations Supply pre-stabilised enzymes Design and develop biosensors Carry out prototype & contract manufacture of low volume sensors Production of our own environmental & agrifood sensors Products & Services

4. TSB SENSACELL-TOX PROJECT TASKS Stabilise GOX and LOX in a water based carbon ink Print microelectrode structures < 500 Micron Incorporate glucose and lactate biosensors on the final chip

5. 500 micron 7mm Microelectrode Design

6. Screen Printed Microelectrodes

7. Schematic of Oxidase Biosensor

8. Amperogram obtained using glucose biosensors in stirred solution. Arrows correspond to 70 µl additions of 50 mM glucose to 10 ml supporting electrolyte. Water Based GOX Ink

9. Glucose calibrations carried out on GOX/carbon ink stored for up to 215 days at room temperature GOX Ink Stability

10. Electrode Stability Water based GOX electrodes printed and stored at 4oC. Electrodes tested using 2mM glucose additions

11. Effect of sterilisation via gamma irradiation on GOX response Lactate Biosensor

12. Fig.5. (A) amperogram and (B) calibration plot for L-lactate in culture medium using a microband biosensor at 37 o C. (A) (B)

13. Lactate calibration carried out on sensors stored for 28 days at RT Lactate Biosensor

14. A + cells - cells Cell Measurements GlucoseLactate

15. Stencil Printing Electroformed apertures Positive advantage since no threads to reduce open area. Stencils provided by Tecan Ltd. Results from HIPER-ACT EU Project

16. Stencil Printing X10 MagX60 MagX 200 Mag Measured Line Widths (µm) 35 41 39 36 39 30 47 Ave39 Vertical print stroke 20 micron stencil

17. Ink Jet Printing

18. Stabilised GOX in the carbon ink for 215 days at room temperature Stabilised the LOX in carbon ink for 28 days at room temperature Successfully proved that the electrode performance was not effected by gamma irradiation sterilisation technique Successfully printed structures < 500micron square onto final device chips Final devices tested on cell media Conclusions

19. SENSACELL-TOX PARTNERS Funded by the Technology Strategy Board

20. Applied Enzyme Technology Ltd. Monmouth House Mamhilad Park Pontypool NP4 OHZ Tel: 01495 751 100 Fax: 0870 052 8250 Email: sales@gwent.org Website: www.gwent.orgwww.gwent.org