1. The development of an NDV-Fusion protein ELISA and its place in a full NDV diagnostic strategy
Luuk Stooker, DVM
Sales Director EMA BioChek
Lunch Meeting IPPE, Wednesday January 31st

2. NDV diagnostic approach
Poultry
Antibody detection
NDV ELISA
Antigen detection
NDV PCR

3. OIE states the following:
Why monitoring for NDV
OIE states the following:
1. To demonstrate historical freedom from infection in a defined population (country/zone/compartment/flock)
2. To determine immune status in individual animals or populations (post-vaccination)
3. To monitor infection or disease in unvaccinated populations
4. To estimate prevalence of infection to facilitate risk analysis in non-vaccinated populations (surveys/flock health schemes/disease control)
Monitor for negatives
Monitoring for disease exposure
Vaccination monitoring

5. Why did we develop the NDV-F ELISA?
One of the primary reasons to monitor for NDV is confirmation of success of vaccination
The current BioChek NDV ELISA has a low temporal sensitivity for monitoring vector NDV responses
Hence, we developed a new ELISA kit suitable for monitoring NDV vector vaccine responses

6. Technical background
Vector vaccine technology is based upon inserting a part of the genome (=the insert) of a pathogen into the DNA of a carrier vaccine to induce protection for both pathogens using the replication route of the carrier
The result is a specific antibody response against the part of the pathogen which has been inserted
Whole virus coated ELISA’s contain a relative low amount of binding sites specifically targeted to the insert epitope  reduced sensitivity
By coating for the insert epitope, the sensitivity of the assay will be increased

7. HI vs NDV-F Temporal Sensitivity
SPF chickens vaccinated with r-HVT-ND at day-old s.c.
Sampling (n=18) at 14 days post-vaccination
SPF VACCINATED
14 Days p.v.
HI (LaSota, 4 HAU)
NDV-F ELISA
Mean Titer
2.7
2919
% Positive Titers
72%
74%
Data Ceva, in spf al na 14 dagen zie je de eerste respons (no MAB’s)
In SPF birds positive vaccination response detected as early as 14 days post vaccination with both HI and recombinant NDV-F ELISA

8. HI vs NDV-F Temporal Sensitivity
Commercial broilers vaccinated with r-HVT ND at day-old s.c. versus non-vaccinated birds
Sampling (n=20) at 14 and 21 days post-vaccination
NON VACINATED
14 days
21 days HI
(LaSota 4 HAU)
% POS 5% 0%
NDV-F ELISA
25%
VACCINATED
14 days
21 days 5%
25%
30%
50%
Chickens with MAB’s have response delayed with one week. Data Ceva phylaxia, HI less sensitive
In commercial broilers (MDA +) positive vaccination response detected as early as 21 days post vaccination

9. NDV vs NDV-F: ELISA Temporal Response
R&D data, samples supplied by Ceva. Mooi verschil te zien in titer en positivity in huidige en nieuwe kit

10. NDV vs NDV-F ELISA Temporal Response
Own data. See slide 11 and 10

11. NDV-F : live vaccines enhance r-HVT ND vaccination reponse
Combination live plus vector is boosting response

12. Classic NDV vs NDV-F: ELISA Field Results
Field data Turkey, correlation with protection needs to be checked

13. Intended use Reason for testing Assay Vaccination check vector vaccine
NDV-F
Vaccination check classical NDV vaccine
NDV
Vaccination check classical NDV vaccine plus vector vaccine
NDV or NDV-F
Differentiation vector vaccine response vs field infection

14. NDV-F ELISA summary
The new NDV-F ELISA is suitable to detect specific vector vaccine induced antibodies
Combining the two NDV ELISA kits together gives you all tools in hand to:
Confirm success of NDV vaccination
Monitor for possible field challenge

15. NDV Diagnostic approach
Poultry
Antibody detection
Routine Monitoring
NDV ELISA
Monitoring vector vaccine response
NDV-F ELISA
Antigen detection
NDV PCR

16. BioChek PCR NDV with NDV marker and virulence marker

19. NDV diagnostic approach
Poultry
Antibody detection
Routine Monitoring
NDV ELISA
Monitoring vector vaccine response
NDV-F ELISA
Antigen detection
Vaccine strain
Field Strain
NDV PCR

20. Acknowledgements Dr. Bart van Leerdam, Holland CEVA Phylaxia, Hungary
MSD Boxmeer, Holland
Polimed, Turkey
VIC, Russia
Cabinet veterinaire, Maroc
Immunovet, South Africa