FMDv NSP Ab Rapid test kit

Passion, Integrity, Professional, and Ethics

Excelsior Bio-System

P r o fe s s i o n a l a n d Re l i a b l e

Profe s s i ona l and Re l i ab l e

Costs of eradicating 1997 outbreak estimated to be US$ 378.9 million,

Potential export losses of approximately US$ 1.2 billion (Data analysis Yang et al. (1999))

Foot & Mouth Disease (FMD) – A big economic Threat


Sentinel® FMDV Non Structural Protein Antibody Rapid Test

Sentinel® FMDV Non Structural Protein Antibody ELISA kit

What is NSP?

4

Foot and mouth disease virus

Viral structural proteins, SP

Viral non structural proteins (NSPs)

Pro fe s s i ona l and Re l i ab l e

http://upload.wikimedia.org/wikipedia/commons/c/c9/Foot-and-Mouth-Disease-Virus.png

Vaccine and viral protein

Viral structural proteins

Viral non structural proteins (NSPs)

Early vaccine

New vaccine

Not present inFMDV structure,Not in New vaccine!

Could be present

Only appeared in viral replication process



Removing NSP in vaccines

Distinguish between animals that have been vaccinated and animals that are infected!

FMD Vaccine (NSP removed)

inoculation

blood

Antibody : OnlyAnti-structural protein

infection

FMDV

Antibody : Anti-structural protein & Anti-nonstructural protein

blood











Differentiation Infected and Vaccinated Animal

Results serology

FMDV NSP test

Vaccinated Infected +

Vaccinated Not infected -Not

vaccinatedInfected +

Not vaccinated

Not infected -


To prove absence of infection or virus circulation

To certify individual animal free of FMD prior to move or trade

To confirm suspected case

To distinguish from infected and vaccinated animal (DIVA)

NSP Ab testing for FMD are performed in support for 4 main purposes


Vaccinated or infected animals

Extremely Easy

High specificity and sensitivity

Monitor FMD Virus activity

Time saving


Sentinel® FMDV NSP Ab Raipid test

ELISA

Sentinal® FMDV NSP Ab Rapid Test

16 hours 60 mins 20 mins

Read by ELISA ReaderStop reactionWash

Add Secondary Antibody Develop with Substrate

Wash

Add Sample and Control

30 mins

Add Running buffer

Interpret by eyesAdd Sample

Over 17 hours!!


Protocol


Validation test and Field test


• Swine sera provided by Animal Health Research Institute (AHRI).

• Total 62 positive swine samples from pigs experimentally infected

with FMDV O/Taiwan/1997 strain at 14 days post-infection(dpi)

were tested, and results are as follows:

Sensitivity :98.39 % (61/62)

Lab tests-Sensitivity


Lab tests-Specificity

• Animal Health Research Institute (AHRI) provided swine sera and analyzed.

• Total 412 FMD negative swine sera from vaccinated pigs, pre-outbreak pigs and SPF pigs were tested.

Specificity :92.96 % (383/412)


•A serum panel collected sequentially from 32 secondary SPF pigs

experimentally challenged by the FMDV O/Taiwan/1997 strain was

tested. The sera in the panel were sampled at different days post

infection(dpi).

•Results were compared with PrioCHECK, UBI and Chekit ELISA kit.testing limited

Tes

t p

osit

ive

%Lab tests-early testing

The validation test was executed by the Animal Technology Institute Taiwan (ATIT).

169 swine sera were blind tested.Then compared results with PrioCHECK® FMDV NS (Ceditest)

and UBI® FMDV NS ELISA(swine), respectively.The data had showed that the Kappa value of Sentinel were

more than 0.8, it indicate the data of Sentinel® were almost the same result with PrioCHECK® and UBI®.


Observed agreement

Kappa value Sensitivity Specificity

PrioCHECK 0.95 0.87 90.2 % 96.9 %

UBI 0.95 0.86 86.4 % 97.6 %

Field tests-comparison with other kit

Kappa（ K coefficient of agreement； K）

P = (observed agreement)：

P (chance agreement)：=

─────P

-Po Pc

c-1K=

o

c



Kappa（ K coefficient of agreement； K）

0.81~1 (almost perfect)

0.61~0.80 (substantial)

0.41~0.60 (moderate)

0.21~0.40 (fair)

0.0~0.20 (slight)

K value is between -1 and 1. If the raters are in complete agreement then κ = 1. If there is no agreement among the raters other than what would be expected by chance


Sentinel ® FMDV NSP Ab ELISA kit


Sentinel ® FMDV NSP Ab ELISA kit

Reliability


Two internal controls (IC-SPC , IC-PC )* were run in 2 replicates on 5 different plates in the same lot.(The data presented here are CV)

within run precision

PI Value Plate-A Plate-B Plate-C Plate-D Plate-E

SPC 0.27% 0.24% 0.00% 0.07% 0.62%

PC 0.31% 0.69% 1.46% 1.31% 2.32%


between run precision Two internal controls (IC-SPC , IC-PC )* were run 10 replicates in the same lot.

*Strong positive control-SPC*positive control-PC

PI Value AVE. STD CV

SPC 96.5 0.31 0.33%

PC 89.0 1.26 1.42%

different lot precision

Two internal controls (IC-SPC , IC-PC ) were run in PD601631 and PD601648, 2 different lots.

PI Value Ave. STD. CV

SPC 96.4 0.14 0.15%

PC 86.8 3.11 3.58%

P r o f e s s i o n a l a n d R e l i a b l e

When tested the same sample in different lot, the result were high consistency


Lab tests- Sensitivity and Specificity Animal Health Research Institute (AHRI) provided swine and goat sera than

analyzed.

1. Swine sera30 swine sera infected with O type FMD virus50 FMDV-free swine seraSensitivity： 96.7% (29/30)Specificity： 100% (50/50)

2. Goat sera30 goat sera infected with FMD virus50 FMDV-free goat seraSensitivity： 100% (30/30)Specificity： 100% (50/50)

Field data

P r o f e s s i o n a l a n d R e l i a b l e

　　

　

PrioCHECK®

　　 Total+ -

Sentinel®+ 47 5 52- 4 57 61

Total 51 62 113

113 pigs sera were collected from field farm,Sentinel ® NSP Ab ELISA test kit and PrioCHECK ® NS ELISA were compared

• Observed agreement = 0.92

• Kappa value= 0.83

• Sensitivity※ = 92.20%

• Specificity※ = 91.90%

• ( ※ PrioCHECK ® as standard)

Sentinel ® FMDV NSP Ab ELISA test kit compared with PrioCHECK ® FMDV NS ELISA kit


PrioCHECK®

Total+ -

Sentinel®+ 17 2 19- 3 159 162

Total 20 161 181

181 cattle sera were collected from field farm,Sentinel ® NSP Ab ELISA test kit and PrioCHECK ® NS ELISA were compared

• Observed agreement = 0.97• Kappa value= 0.86• Sensitivity※ = 85%• Specificity※ = 98.1%• ( ※PrioCHECK ® as standard)




PrioCHECK®

Total+ -

Sentinel®+ 3 1 4- 1 127 128

Total 4 128 132

132 goats sera were collected from field farm,Sentinel ® NSP Ab ELISA test kit and PrioCHECK ® NS ELISA were compared

• Observed agreement = 0.98

• Kappa value= 0.74

• Sensitivity※ = 75%

• Specificity※ = 99.2%

• ( ※ PrioCHECK ® as standard)



Field tests-validation： The agreement of analytical performance compare to commercial ELISA kit (PrioCHECK)

Species Sample No. Observed agreement Kappa value

Swine 113 0.92 0.83

Cow 181 0.97 0.86

Goat 132 0.98 0.74


The performance of our FMDV NSP Ab Testing kits

• Sensitivity• Specificity• Reliability


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FMDv NSP Ab Rapid test kit