CSFV ELISA Antibody Detection Kit (Indirect Method)

User’s Manual
Item# FZES016-96(96 wells); FZES016-192(192 wells); FZES016-480(480 wells)
【Instruction】
Classical swine fever virus (classical swine fever virus, CSFV) ELISA 4°C for about a week.
antibody detection kit is used to detect the CSF virus antibody in pig
serum and evaluate the immune status or infection status of the swine 【Dilution of samples】
fever vaccine in pig farms. Dilute the serum to be tested 100 times in a serum dilution plate (for
This kit adopts the indirect ELISA method, pre-coating the classical example: add 3 μl of serum to be tested to 297 μl of sample diluent).
swine fever virus antigen on the microwell of the enzyme label strip.
In the test, the diluted serum to be tested is added, and after incubation, Note: Negative and positive controls are not diluted. Tips were
if the sample contains CSFV-specific antibodies, it will bind to the changed after each sample, and the position of each sample on the
antigen epitope coated in the pre-coated plate wells, and the unbound plate was accurately recorded. Each sample should be mixed
antigenic epitopes will be removed by washing. After the antibody and thoroughly before adding to the microwells of the coated plate.
other components, the enzyme-labeled secondary antibody is added to
specifically bind to the antigen-antibody complex on the detection 【Precautions】
plate, and then the unbound enzyme label is removed by washing, and 1. The reagents of the kit should be equilibrated to room temperature
the TMB substrate solution is added to the microwell, and the enzyme before use, and the components of the kit should be placed at room
Catalyze to form a blue product, add stop solution to terminate the temperature for at least 1 hour. Shake well before use, and store at
reaction, and measure the absorbance OD value in the reaction well 2~8°C as soon as possible after use.
with a microplate reader at a wavelength of 450 nm. 2. When drawing and adding samples, each sample should be
replaced with a disposable tip.
【Contents】 3. Reagent components of different products or different batches of
the same product should not be mixed. When using reagents, prevent
Specification X Q’ty cross-contamination of reagents or contamination by samples.
No. Contents
192T/Box 4. The substrate and stop solution may be irritating to the skin and
eyes, so care should be taken when using them.
1 Antigen Pre-Coated Plates 2 Pcs 5. Do not expose TMB (chromogenic solution) to strong light and
2 Enzyme Marker 11ml/Bottle X 2 avoid contact with oxidizing agents.
6. After the test board is unpacked, avoid moisture or water (put the
3 10X Concentrated Washing Liquid 100ml/Bottle X 1 unused test board in a ziplock bag with desiccant and place it at
2~8°C as soon as possible).
4 Chromogenic Liquid 22ml/Bottle X 1 7. All waste should be disposed of properly before discarding to avoid
polluting the environment.
5 Sample Diluent 100ml/Bottle X 1 8. The serum dilution plate is a one-time product and cannot be reused;
the maximum capacity of the serum dilution plate is 300μl/well.
6 Stop Buffer 15ml/Bottle X 1
9. During the operation, the whole process of pipetting, timing and
7 Negative Control 2ml/Bottle X 1 washing should strictly follow the operating instructions to obtain the
best results.
8 Positive Control 1.6ml/Bottle X 1

9 Dilution Plate 2 Pcs 【Procedures】

1. Take the pre-coated test plate (depending on the sample size, it can
10 Membrane 4 Pcs be disassembled and used separately), and add 100μl of the diluted
11 User’s Manual 1 Page
serum to be tested to the well of the test plate. At the same time, set 1
well for negative control and 2 wells for positive control. Add 100μl
of negative control and positive control to the reaction wells
【Equipment and reagents to be prepared】 respectively, and gently shake the samples in the wells (do not
1. Micropipette: 0.5µl~10µl, 10µl~100µl, 100µl~1000µl. overflow).
2. Disposable tips for pipettes. 2. Cover with membrane (can be cut according to actual needs), and
3. Measuring cylinder: 500ml. incubate at 37°C for 30 minutes in the dark.
4. 96-well plate microplate reader. 3. Carefully peel off the membrane, shake off the liquid in the plate
5. Distilled or deionized water. wells, add 250μl of the working concentration washing liquid to each
6. Bottle or plate washer. well, and shake off the liquid in the wells. Repeat the above steps 4
to 6 times, and finally pat dry on clean absorbent paper.
【Sample Preparation】 4. Add 100μl of enzyme marker to each well, cover with membrane,
and incubate at 37°C for 30 minutes in the dark.
Whole blood was taken from animals, and serum was prepared
5. Carefully peel off the membrane, shake off the liquid in the plate
according to conventional methods. The serum was required to be
wells, and wash 4~6 times. The method is the same as 3. Remember
clear and without hemolysis.
to pat dry on clean absorbent paper at the end.
6. Add 100μl of chromogenic liquid to each well, shake and mix
【Preparation of washing liquid】
evenly, cover with membrane and color developing at 37°C for 10
The concentrated washing solution should be returned to room minutes in the dark.
temperature before use. If the salt crystallizes, shake to dissolve the 7. Add 50μl of stop solution to each well to terminate the reaction,
crystallized salt, and then dilute it 10 times with distilled water or and measure the OD value at 450 nm with a microplate reader within
deionized water. The diluted washing solution can be stored at 10 minutes.
Te c h n i c a l S u p p o r t ： 1 3 5 3 02 1 7 3 0 9 E m a i l ： t e c h @ z r- b i o . c o m Te l ： 0 7 5 5 - 2 6 5 8 2 6 3 9 We b s i t e ： w w w. z r- b i o . c o m
【Enzymatic Calibrator Setting】
Read the absorbance OD value with a microplate reader at 450nm
(630nm as a reference wavelength).

Conditions for the establishment of the test:

Negative control (N) OD value < 0.2, while positive control (P) OD
value ≥ 0.6.

Calculation method:
Sample OD value ÷ positive control OD mean = S/P value

【Results Judgment】
S/P≤0.25 is judged as negative;
S/P≥0.3 is judged as positive;

【Storage & Shelf Life】Store at 2~8℃ for 12 months

Instruction Verification Dated: 23 Aug., 2022

Te c h n i c a l S u p p o r t ： 1 3 5 3 02 1 7 3 0 9 E m a i l ： t e c h @ z r- b i o . c o m Te l ： 0 7 5 5 - 2 6 5 8 2 6 3 9 We b s i t e ： w w w. z r- b i o . c o m