Download www .bio x.com Anti-VIRAL HAEMORRHAGIC SEPTICAEMIA

BIO 281
REAGENT FOR DETECTION OF VHSV ON TISSUE SECTION OR CELL CULTURE (IP5B11)
INTRODUCTION
Viral haemorrhagic septicaemia (VHS) is a rhabdovirus infection that is very common on the
European continent. It causes great losses on European trout farms - an estimated 20 to 30
thousand tons of trout are lost to this disease each year. Three distinct serotypes of this virus
have been identified. The rainbow trout (Oncorhynchus mykiss) appears to be very sensitive to
this virus. However, other Salmonidae, such as the brown trout (Salmo trutta fario L.), grayling
(Thymallus thymallus L.) and even white fish may develop the disease. Salmonidae hybrids
show variable degrees of sensitivity to the virus. The pike (Esox lucius L.) is also sensitive to
VHS, primarily as fry, but the adult fish is also vulnerable. The pathogenic power of this virus
is expressed only in water that is colder than 14° C. This explains why the disease strikes primarily in the winter.
The clinical signs of the disease are high mortality, especially during the young trout’s first winter. The subjects exhibit melanosis and exophthalmia. The paleness of their gills reflects their
anaemic condition. An autopsy will reveal the presence of numerous sites of haemorrhages
in the viscera and muscle mass. It is practically impossible to distinguish VHS from infectious
haematopoietic necrosis (IHN) - another Salmonidae viral infection likewise caused by a rhabdovirus - on the basis of clinical examination alone. A differential diagnosis must thus be performed in a laboratory. The clinical diagnosis is most often confirmed by isolating the virus in a
cell culture. BIO 281 is specific for VHSV N-protein.
EXAMPLE OF RESULTS
Bio-X Diagnostics - 38, Rue de la Calestienne (PAE) - 5580 Rochefort - Belgique
Tél : 0032(0)84.32.23.77 - Fax : 0032(0)84.31.52.63 - E-mail : [email protected]
www.biox.com
Anti-VIRAL HAEMORRHAGIC SEPTICAEMIA VIRUS
Monoclonal Antibody labelled with Fluorescein
Isothiocyanate
- Paraformaldehyde 2 % in PBS
- Acetone solution (9 volumes of acetone and 1 volume of water).
- Isopropanol
Rince with PBS.
Dilute the conjugate twentyfold with a PBS-Evans blue solution made up according to the following formula:
PBS - Blue Evans
________________
NaCl:8 gr
KH2PO4:0.2 gr
KCl:0.2 gr
Na2HPO4 . 2H2O:
1.15 gr
Blue Evans:
0.01 gr
NaN3:0.1 gr
H201 L
Incubate the sample with the fluorescein-labelled conjugate for 1 hour at room temperature. At
the end of this incubation period rinse the cell preparation with a PBS solution. Dry the cell preparation, then add the mounting medium prepared as follows:
Mounting medium
_________________
Glycerol
9 volumes
PBS1 volume
Examine the cell preparation under a microscope equipped for detecting fluorescence.
COMPOSITION: One vial of 500 µl
STORING THE CONJUGATE: The conjugate must be stored at 4°C. It must never be frozen.
STABILITY: One year at 4°C
Bio-X Diagnostics - 38, Rue de la Calestienne (PAE) - 5580 Rochefort - Belgique
Tél : 0032(0)84.32.23.77 - Fax : 0032(0)84.31.52.63 - E-mail : [email protected]
www.biox.com
Fix the cell preparation (cell cultures or tissue sections) for 15 minutes at room temperature with
one of the following fixators :
IO 281
B
BIO 281
BIO 281
1:20
1:40
1:80
None
None
None
RTG-2
RTG-2
RTG-2
0
0
0
IO 281
B
BIO 281
BIO 281
1:20
1:40
1:80
VHS903 (Fi13)
VHS903 (Fi13)
VHS903 (Fi13)
EPC
EPC
EPC
+
+
+
IO 281
B
BIO 281
BIO 281
1:20
1:40
1:80
None
None
None
EPC
EPC
EPC
0
0
0
IO
B
BIO
BIO
BIO
BIO
281
281
281
281
281
1:20
1:20
1:20
1:20
1:20
VHS 903 (Fi13)
VHS 922 DF72/94 (Isolate Wi)
VHS 917 DK-3592 «Voldbjerg»
VHS 958 07/71 (France)
VHS 919 1P8 (Isolate from herring)
RTG-2
RTG-2
RTG-2
RTG-2
RTG-2
+
+
+
+
+
IO
B
BIO
BIO
BIO
BIO
281
281
281
281
281
1:20
1:20
1:20
1:20
1:20
VHS 903 (Fi13)
VHS 922 DF72/94 (Isolate Wi)
VHS 917 DK-3592 «Voldbjerg»
VHS 958 07/71 (France)
VHS 919 1P8 (Isolate from herring)
EPC
EPC
EPC
EPC
EPC
+
+
+
+
+
IO 281
B
BIO 281
1:20
1:20
IHN 265 (Isolate 332 from Germany)
SVC 1228 RVC 56/70 (FIJAN)
RTG-2
RTG-2
0
0
IO 281
B
BIO 281
1:20
1:20
IHN 265 (Isolate 332 from Germany)
SVC 1228 RVC 56/70 (FIJAN)
EPC
EPC
0
0
Bio-X Diagnostics
Site du complexe des Postes 49, rue Joseph Wauters BE-5580 Jemelle BELGIUM (Europe)
Tel: +32 (0)84.32.23.77 Fax: +32 (0)84.31.52.63 E-mail: [email protected] http://www.biox.com
www.biox.com
Dilutions
Strains / Virus
Cells Results
-------------------------------------------------------------------------------------BIO 281 1:20
VHS903 (Fi13)
RTG-2
+
BIO 281 1:40
VHS903 (Fi13)
RTG-2
+
BIO 281 1:80
VHS903 (Fi13)
RTG-2
+/-