Peste
des petits ruminants virus (PPRV) antibody ELISA kit
Catalog No. LSY-30035
1. Usage
It
is used to detect PPRV antibody in serum of sheep and goat qualitatively,
evaluate the immune status of Peste des petits ruminants vaccine and assisted
serological diagnosis of infected animals.
2. Principle
This
kit use competition ELISA method, PPRV antigen is pre-coated on enzyme
micro-well strips. When testing, add serum sample and Monoclonal Enzyme
conjugate, after incubation, if there is PPR virus specific antibody, it will
bind to the PPRV antigen on the coating plate and prevent the enzyme-labeled
monoclonal antibody from binding to the antigen on the plate; conversely, if
the sample does not contain PPRV-specific antibody, it will not bind to the
coating plate. After washing to remove the unbound antibody and other
components, add substrate to the microwells to form a blue product through
enzymatic catalysis. After adding stop solution to terminate the reaction, use
a microplate reader at 450nm/630nm double wavelength to measure the absorbance
A value in the reaction well.
3. The kit components
|
1
|
PPRV
antigen coated microplate
|
96T
X 2
|
|
2
|
Enzyme
conjugate
|
12ml
|
yellow
lid
|
|
3
|
Sample
dilution
|
12ml
|
transparent
lid
|
|
4
|
PPRV-IgG
Negative control serum
|
1.5ml
|
green
lid
|
|
5
|
PPRV-IgG
Positive control serum
|
1.5ml
|
red
lid
|
|
6
|
Substrate
|
12ml
X2
|
orange
lid
|
|
7
|
Stop
solution
|
12ml
|
blue
lid
|
|
8
|
10×concentrated washing buffer
|
50ml
|
white
lid
|
|
9
|
Adhesive
Foil
|
2
pieces
|
|
|
10
|
Instruction
|
1
piece
|
|
4. Material required not provided
1)
Microplate Reader (double-wave length: 450/630
nm).
2)
Precise micropipette (single-channel 10-100ul、0.5-10ul、multi-channel 30-300ul)
3)
Constant temperature box or water bath box.
4)
Oscillator.
5)
Disposable tips (10ul, 200ul)
6)
Deionized water
5. Sample requirement
1)
The samples are sheep and goat serum, which
should be collected with no bacteria. The storage time should be less than 1
week at 2-8 ℃, if for long
term, it should be kept at -20℃.
2)
Avoid to use the samples with severe hemolysis,
precipitate, contaminated by bacteria or protein suspension.
6. Preparation
1) Bring ELISA reagents
to the room temperature (20-25 ℃)
for 30 min to get best results. Microplate should return to room temperature
and dry before open package.
2)
Washing solution preparation: Dilute the 10×concentrated washing buffer with
deionized water at 10 times. (10ml 10×concentrated washing buffer + 90ml deionized
water ) It is normal if there is crystallization in the 10×concentrated washing buffer, put at 37℃ until completely dissolved.
7. Test procedure
1) Take the pre-coated microplate (according to the number of samples, it can be
disassembled and used separately), set 2 wells for positive control, 2 wells
for negative control , add 50µl positive control serum or negative control
serum to it’s well accordingly; others are sample well, firstly add 40µl sample
dilution, then add 10µl serum sample into each well. At last, add 50µl Enzyme
conjugate to each well, shake gently to mix it evenly, cover the plate
with adhesive foil, incubate at
37 ℃ for 30 minutes;
2)
Open the adhesive foil, discard the liquid of the well, add diluted washing buffer to each
well, 300ul/well, be static for 30s, then discard the liquid, repeat the above
step for 5 times, at last flap to dry with the absorbent paper;
3) Add substrate
solution,100ul/well, mix it evenly then cover it with Adhesive Foil, incubate at 37 ℃ in dark for 15 minutes;
4)
Add stop solution 50ul/well to stop the reaction, measure the result in 10
minutes.
8. Results judgement
Read
the OD value with ELISA Reader at 450nm (630nm as reference).
For the
assay to be valid:
OD value of Negative control (N) ≥0.60,
OD value of Positive control (P) ＜0.40;
Calculate
method:
Sample OD value/average OD value of
Negative control (N)= S/N value
Results interpretation
S/N value≥0.5: Negative
S/N value<0.5: Positive
9. Precautions and warnings for users
1). Read the Manual carefully before use.
2). Do not use reagents expired, do not mix reagents from
different lots.
3).
Experiment rubbish should be dealt with high pressure steam sterilization at 121
℃ for 30
minutes, or treated with 5.0g/L sodium hypochlorite disinfectant for 30 minutes,
then discard.
4).
MicroWell plate removed from the refrigerated environment should be balanced
moisture to dry at room temperature, then can be opened. Put back unused
MicroWell plate into dry foil bag and sealed at 4 ℃. Unused liquid reagent should cover
caps, store at 2-8 ℃ in dark with other group components.
5).
Should use Micropipettor to add sample and reagents, and often proof its
accuracy.
6).
When adding washing buffer, should be full but no overflow, avoid appearing
free enzyme at mouth of well or cross pollution between wells.
7).
Stop solution is corrosive, use large amount of water to wash immediately when
touch the skin or clothes.
10.Storage and expire date
Store at 2~8℃ in dark, no frozen,
expiry date: 12 months.
Shenzhen Lvshiyuan Biotechnology Co., Ltd
D Building,
National Biological Industrial Park of Marinelife, No.2 Binhai Road, Dapeng,
Shenzhen, 518120 China
Tel.
86-755-28438788 Fax 86-755-28938800
Email: info@lsybt.com
www.lsybt.com