Post on 28-Jun-2020
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Trial report of Anigen
Bovine Brucella Diagnostics
ANIGEN B. Brucella Antibody
Rapid Test Kit & ELISA Kit
Introduction
Principle : Indirect Sandwich Method – (LPS capture)-(blood,milk)-(Protein G detector) – LPS extracted from cell
Purpose : Brucella abortus antibody titration in bovine whole blood, plasma, serum and milk
[Rapid Test Kit] [ELISA Kit]
Anigen B. brucella Ab Diagnostics
Principle of B. Brucella Rapid Test Kit
Rose Bengal & Milk Ring Test Anigen Rapid Test Anigen’s
Advantage
Principle -Serum : Rose Bengal
-Milk : Milk Ring Test
Chromatographic Immunoassay
Specimen -Rose Bengal : Serum
-Milk Ring Test: Milk
Whole blood, serum, plasma, Milk
available
All specimen
are available
Method
- Rose Bengal
1) Preparation of serum
2)1 drop of Rose bengal reagent +
1 drop of serum : Mixing
3) Interpret the result within 4 min.
- Milk Ring Test(MRT)
1) Adding of 1ml of milk into a tube
2)Adding of 30ul of MRT reagent
3) Waiting for 1 hour at 37℃
4) Reading the result
1) Adding 1 drop of specimen
2) Adding one drop of assay
diluent
3) Reading result at 20 min.
Easy Test
procedure
Reading
Method
- Rose Bengal : Agglutination?
- MRT : Changing of blue color of Milk
Positive : 2 line
Negative : 1 line
Simple
Required
testing time
Min.one day 20 Min. Rapid Testing
1st Screening Method [Rose Bengal Test, Milk Ring Test vs. Rapid Kit]
SVANOVIR : Sweden Anigen : Korea
Principle ELISA ELISA
Specimen Serum & Milk Serum & Milk
Method Similar Similar
Reading Method Complicated result calculation
OD of neg. standard PP= ------------------------------------- X 100 OD of pos. standard
Serum cut off : PP>25
Milk cut off : PP>10
Simple result calculation
Required testing time 2 hours 10 min. 2 hours
2nd Confirmatory Method [Reference ELISA kit vs Anigen ELISA Kit]
Clinical Field Trial - Anigen B. Brucella Ab Rapid Test -
Korea Kyonggi-Do Province Livestock and Veterinary Service
KLVS
Performed by
Sensitivity and Specificity - of serum specimen -
Anigen
Rapid
+ 145
- 182
SVANOVIR
ELISA
+ -
139 6
3 179
142 185
Anigen
Rapid
+ 118
- 35
Rose
Bengal
+ -
118 0
35 0
153 0
Sensitivity : 97.9%
Specificity : 96.8%
- Rose Bengal Test
: Current 1st screening assay
- Specimens : serums from KLVS
- SVANOVIR ELISA Kit
: Current confirmatory assay
-Specimens : serums from KLVS
Sensitivity : 77.1%
Specificity : 100.0%
Sensitivity : 87.1%
Specificity : 98.6%
Anigen
Rapid
+ 30
- 36
SVANOVIR ELISA
+ -
30 0
1 35
31 35
Anigen
Rapid
+ 30
- 36
Milk Ring Test
+ -
26 4
1 35
27 39
Sensitivity : 96.7%
Specificity : 97.2%
- Milk Ring Test : Current 1st screening assay
- Milk specimens : from random 66 cows KLVS
- SVANOVIR ELISA : Current confirmatory assay
- Milk specimens : from random 66 cows KLVS
Sensitivity : 96.2%
Specificity : 89.7%
Sensitivity : 96.5%
Specificity : 93.4%
Sensitivity and Specificity - of Milk specimen -
A. Specimens: Serum and Milk specimens from same cow of randomly selected 66 cows of KLVS
B. Assay Method :Anigen B. Brucella Ab Rapid Kit
Serum Total
+ -
Milk
+ 30 0 30
- 1* 35 36
Total 31 35 66
* Serum test showed weak positive
The serum and milk shows similar result between serum and milk:
Agreement rate: 98.5%
Agreement rate - Serum and Milk specimen -
Detection Limit Study - serum and milk specimens -
No. of serum specimens No. of milk specimens
Titer(2n) Titer
Result of Detection Limit Study
A. Serum : Anigen rapid test is more 256 ~ 1,024 times sensitive than Rose Bengal
B. Milk: Anigen rapid test is more 2 ~ 4 times sensitive than Milk Ring Test
Study principle : Comparison of detection limit of diluted serum and milk specimens
12
4
13
5
13
5
17
7
15
7
0
2
4
6
8
10
12
14
16
18
1 2 3 4 5
래피드 진단시약 Rose BengalAnigen Rapid
5120
1280
160 80 16040
5120
1280
0
1000
2000
3000
4000
5000
6000
1 2 3 4
래피드 진단시약 MRTAnigen Rapid
OIE Evaluation
Result
Clinical Field Trial - Anigen B. Brucella Ab ELISA Kit -
Korea Kyonggi-Do Province Livestock and Veterinary Service
KLVS
Performed by
Anigen
ELISA
+ 130
- 141
SVANOVIR
ELISA
+ -
127 3
0 141
127 144
Sensitivity : 100%
Specificity : 97.9%
( SVANOVIR ELISA : current confirmatory assay)
Sensitivity and Specificity - serum specimens -
A. Specimens: Total 271 of positive and negative cow serums from KNVRQS
B. Assay Method : Comparison of Anigen ELISA and SVANOVIR ELISA Kit(Sweden)
( KNVRQS: Korea National Veterinary Research and Quarantine Service)
Anigen
ELISA
+ 69
- 116
SVANOVIR
ELISA
+ -
68 1
0 116
68 117 185
Sensitivity and Specificity - of Milk specimen -
A. Specimens: Total 185 of positive and negative cow milk from KLVS
B. Assay Method : Comparison of Anigen ELISA and SVANOVIR ELISA Kit(Sweden)
Sensitivity : 100%
Specificity : 99.1%
0.000
0.200
0.400
0.600
0.800
1.000
1.200
1 2 3 4 5 6 7 8 9
Anigen SVANOVA
0.000
0.500
1.000
1.500
2.000
2.500
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16
Anigen SVANOVA
Dilution (2n) Dilution (2n)
Comparison of detection limit in serum Comparison of detection limit in milk
OD OD
Detection Limit Study - of serum and milk specimens -
Study principle :
Comparison of detection limit of diluted positive serum confirmed by Rose Bengal
and milk confirmed by Milk Ring Test
Anigen B. Bruella Ab ELISA kit showed the detection limit:
- Anigen ELISA Kit is 2,048 times sensitive than Rose Bengal Test in serum
- Anigen ELISA Kit is 128 times sensitive than Milk Ring Test in milk
It showed that Anigen ELISA showed similar detection limit with SVANOVIR ELISA Kit in serum and milk
Anigen
Comparison
Anigen Rapid Kit Anigen ELISA Kit
Serum Milk Serum Milk
Sensitivity Specificity Sensitivity Specificity Sensitivity Specificity Sensitivity Specificity
Rose Bengal
/Serum 77.1% - - - - - - -
MRT/Milk - - 96.2% 89.7% - - - -
SVANOVA
B.Brucella ELISA 97.9% 96.8% 96.7% 97.2% 100% 97.9% 100% 99.1%
- Anigen B.Brucella Ab Rapid test kit has better sensitivity and specificity than Rose
Bengal test kit and Milk Ring test.
- Anigen B.Brucella Ab ELISA test kit has better sensitivity and specificity than a
commercially available SVANOVIR ELISA
- Using Anigen B.Brucella Ab Rapid test kit is very simple, quick and it does not
need any special equipments. Therefore, Anigen B.Brucella Ab Rapid test kit will be
suitable to be adopted in clinical laboratory of basic hospital for screening of
Brucella abortous.
Summary of Accuracy
A. Specimen : 65 serums of Rose Bengal test positive
B. Comparative assay: Tube agglutination reaction
C. Positive Result : Anigen Rapid Test-91%, Tube Agglu.-88%
Assay Tube agglutination
Total Positive Negative
Anigen B. brucella
Ab Rapid Test
Positive 55 4 59
Negative 2* 4 6
Total 57 8 65
Anigen B.brucella Ab Rapid Kit (91%) is more sensitive than the Tube
agglutination reaction method (88%)
Additional Validation 1
Bovine
Brucellosis
Conventional Test Method
Rose Bengal Test
Milk Ring Test
PPD Skin Test
(purified protein derivative)
Result in within 10 Min.
Weak points :
- Laboratory Testing
- 1 day required
- RB : Serum only
- MRT : Milk only
Bovine TB
Weak points :
- 3 days required
- Low specificity
Immediate Testing at Farm
Testing in whole blood, serum, Milk.
Anigen Rapid Test Kit
(Bovine Brucella Ab, Bovine TB Ab)
Immediate Preventive Action
Isolation, Disinfections,
Protective slaughter
Reducing the damage
Clean
Application of Rapid Kit at Cattle Farm
Thank you for your attention