SPADA

Detection techniques for botulinum neurotoxins

Method

Limit of Detection Multiplex Format Sample‐Matrix Automation

(ml

‐1

) Interference

1. Mouse Bioassay * 20 pg

No Limited No

ELISA*

Li it d M bl Li it d

5 pg ‐ 2 ng

m e

anagea e

m e

3. ECL* 5 pg ‐50 ng Limited Manageable Limited

Lateral flow assay* 5‐50 ng

No High N/A

5. Column flow assay* 1‐50 ng

No High N/A

6. Flow cytometry assay 50 pg‐20 ng Yes Manageable Yes

7. Immuno‐PCR 1 pg‐ 5 pg Limited Manageable No

8. L‐PCR 0.02 fg Limited N/A No

9. BDG assay 100 ng Yes Low Yes

10. Array biosensor assay 40‐200 ng Yes Low Yes

11. Aptamer electrochemical assay 40 pg Yes Low Yes

12. Peptide array assay 3 pg Yes High Yes

13. ALISSA

0.5 fg No Low No

14. Endopep‐MS assay* 0.4 ‐6 pg Yes High Yes

15.

Cell based assays 1‐10 ng No High No

(* assay used for food analysis); Enzyme-linked immunosorbent assay (ELISA);

Electro-chemiluminiscent (ECL) assay; Liposomal-PCR (L-PCR): Bidiffractive grating (BDG) assay;

Assay with a large immunosorbent surface area (ALISSA)

Fitness for Purpose

To validate a rapid multi plex

Clostridium botulinum

neurotoxin detection assay that has the ability to:

• Detect and differentiate serotypes A through G

• Detect bivalent toxins effectively and also

determine the dominant type

• Differentiate complex toxin with associated

proteins from purified toxin (di-chain) and toxoid