ELISA Echinococcosis Dog Feces Antigen kit

Catalog No. LSY-30024

1. Usage

This test kit is used to detect Echinococcosis antigen in dog feces, used to help diagnosis for infection of Echinococcus granulosus in dogs and carnivores.

This test kit use double-antibody sandwich method, monoclonal antibody against Echinococcus granulosus is precoated on microplate. When run the test, add sample, then incubate, if there is Echinococcosis antigen in the sample, it will combine with the pre-coated monoclonal antibody; discard the uncombined antigen after washing, then add enzyme conjugate, forming pre-coated monoclonal antibody-antigen combination forming monoclonal antibody-enzyme conjugate complex; discard the uncombined enzyme conjugate; add substrate, the blue signal formed by the enzyme is proportional to the antigen content in the sample, measure the OD value by ELISA reader with wavelength at 450nm.

2. Reagents and contents

3. Materialrequired notprovided

1) Micropipette: 0.5ul-10ul, 10ul-100ul, 100ul-1000ul.

2) Disposable pipette tips.

3) Graduate: 500ml.

4) Microplate Reader: 96 wells.

5) Distilled water or deionized water.

6) Microplate Washer

4. Washing bufferand Sample treatment solutionpreparation

1) Return 10X Concentrated washing buffer into room temperature before use, if there is salt crystals, shake to make it dissolved, then dilute it at 10 times with distilled water or deionized water. The diluted washing buffer can store at 4℃for about 1 week.

2) Return 5X Sample treatment solution into room temperature before use, if there is salt crystals, shake to make it dissolved, then dilute it at 5 times with distilled water or deionized water. The diluted Sample treatment solution can store at 4℃for about 1 week.

5.  Sample treatment:

Take 1.0 g dog feces sample into a centrifuge tube, add 2.0ml diluted Sample treatment solution, mix it evenly, centrifuge at 2000g for 10min, take up-layer clear liquid for test.

Note: negative and positive control do not need dilute

6. Notes

1) Return all reagents into room temperature before use, put all reagents at room temperature for at least 1 hour. Shake it evenly before use, and store back to 2-8℃after usage.

2) Do not mix use reagents from different kits and different lot no., prevent the reagents been polluted when using.

3) Substrate A and stop solution may have irritation to skin and eyes, be careful to use.

4) Do not expose Substrate to strong light and avoid contact with the oxidant.

5) Antibody coated plates should be sealed and moisture-proof. Put back unused MicroWell plate into dry foil bag and sealed at 2~8 ℃.

6) All wastes should be treated well to avoid pollution before discarding.

7) Strict compliance with the operating instructions can get the best results. Pipetting operation, timing, and washing of the whole process must be precise.

7. Test procedure

1) Take pre-coated micro-well plate (can be opened and used for different times according to sample quantity), add the diluted sample into sample wells, 100ul/well. Meanwhile set 2 wells for positive control, 2 wells for negative control, add negative control, positive control accordingly to its corresponding wells, 100ul/well; Shake gently (do not spill), Cover it with Adhesive Foil, Incubate at 37℃for30minutes.

2) Open the adhesive foil, discard the liquid of the well, add diluted washing buffer to each well, 250ul/well, then discard the liquid, repeat the above step for 6 time, at last flap to dry with the absorbent paper.

3) Add Enzyme Conjugate into each well,100ul/well, cover it with Adhesive Foil,incubateat 37 ℃ for30 minutes;

4) Washing as step 2, remember at last flap to dry with the absorbent paper;

5) Add substrate into each well, 100ul/well, shake gently, cover it with Adhesive Foil,incubateat 37 ℃ for 10 minutes.

6) Add stop solution, 50ul/well into each well to stop the reaction, measure the result with ELISA reader in 10 minutes.

8. Results judgment

Read the OD value at 450nm wavelength (630nm as reference)

For the assay to be valid:

Average OD value of negative control(N) <0.20, meanwhile average OD value of positive control (P) > 0.5.

Calculate method:

S/P value= Sample OD value/ average OD value of positive control

Resultsinterpretation

Negative: S/P value<0.45;

Positive: S/P value ≥ 0.45.

9. Storage and expire date

Store at 2~8℃in dark, no frozen, expiry date: 12 months.