Pullorum Disease (PD) and Fowl Typhoid (FT) Antibody ELISA kit for poultry diagnostics

Pullorum Disease (PD) and Fowl Typhoid (FT) Antibody ELISA kit

Catalog No. LSY-30027

1. Usage

This kit is used to detect Pullorum Disease (PD) and Fowl Typhoid (FT) antibody in chicken serum, to assess antibody condition by Pullorum Disease (PD) and Fowl Typhoid (FT) vaccine in chicken farm and assist diagnosis of serological infected chicken.

2. Principle

The Pullorum Disease (PD) and Fowl Typhoid (FT) antibody ELISA kit is based on an indirect enzymatic immunoassay (Indirect ELISA).The antigen is coated on plates. When a sample serum contains specific antibodies against virus, they will bind to the antigen on plates. Wash the unbound antibodies and other components. Then add a specific enzyme conjugate. After incubation and washing, add the TMB substrate. A colorimetric reaction will appear, measured by a spectrophotometer (450 nm).

3. Reagents

4. Materials required but not provided

1) Micropipettors and disposable tips: 0.5μL~10μL,10μL~100μL,100μL~1000μL

2) 37 ℃ Incubator

3) Measuring cylinder: 500 ml

4) 96 wells microplate reader

5) Distilled water or deionied water

6) Bottle or microplate washing machine

5. Sample preparation

Take animal whole blood, make serum according to regular methods, the serum should be clear, without hemolysis.

6. Preparation of washing buffer

Return washing buffer to room temperature before use, if there is salty crystals, shake to make the crystals dissolve, then use distilled water or deionied water to diluent it at 10 times. The diluent washing buffer can store for 1 week at 4 ℃.

7. Sample dilution

At serum diluent plate, dilute serum at 1:100 with sample diluent

Notice: Negative control serum and Positive control serum do not need dilute. Exchange tip after taking sample every time, record the situation of the sample on plate accurately. Shake the sample evenly before adding it.

8. Notes

1) All reagents should be adjusted to the room temperature and shake evenly before using, store at 2-8 ℃ after using

2) Do not exchange the reagents from the kits of different lot numbers to use. Avoid reagent pollution when using.

3) Substrate and stop solution may have excitant to skin and eyes, pay attention when using.

4) Do not expose TMB (Substrate B) to light and avoid it contact with antioxidants.

5) The wells should avoid damp or touching water after unsealing (Put the un-using microplate back to bag with dehydrator in 2~8 ℃ soon )

6) Deal all waste reasonable before dumping to avoid pollution.

7) Strictly adhere to instruction to get best result. All procedure including pipetting, timing and washing etc. must be accurate.

8) Serum diluent plate is disposable, do not use for second time; the MAX volume of it is 300μL/well.

9. ELISA procedure

1) Take pre-coated microplate (Can unseal for several time use as per sample quantity), add 100μL diluted serum to the sample well, meanwhile set 1 well for Negative control serum, 2 wells for Positive control serum separately. Add 100 μL Negative/Positive control serum to its wells. S

2) Shake softly and cover the microplate with adhesive film and incubate at 37℃ for 30 min.

3) Open the adhesive film, pour the liquid out of the wells, add 250 μL diluted washing buffer to each well, pour out. Repeat 4-6 times, then pat to dry on absorbent paper.

3) Add 100 μL Enzyme conjugate to each well, cover the microplate with adhesive film and incubate at 37℃ for 30 min.

4) Open the adhesive film, pour the liquid out of the wells, add 250 μL diluted washing buffer to each well, pour out. Repeat 4-6 times, then pat to dry on absorbent paper.

5) Add 100 μL substrate solution to each well, mix properly, react for 10 min at 37℃ in dark.

6. Add stop solution 50 μL in each well, and measure the result within 10 min.

10. Results

Read the OD value by microplate-reader at wavelength of OD450nm (630 nm as reference).

The test to be valid :

OD value of Negative control (N)<0.2, meanwhile OD value of Positive control >0.4

Calculation method:

OD value of Sample/Average OD value of Positive Control=S/P value

Result judgement:

S/P<0.35: Negative;

S/P≥0.35: Positive.

Specifications: 96*2 wells/kit.

Expiry date: 12 months.

Storage: Storing at 2-8℃, in the dark.

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