Bioneovan HEV-IgG ELISA is an in vitro enzyme linked immunoassay supplied for the detection of HEV-IgG in human serum or plasma. It is intended to be used as an aid in supplementary diagnosis to acute hepatitis E infection and prevalence studies among the population.

This kit employs solid phase, indirect ELISA method for detection of IgG antibodies to HEV (anti-HEV) in serum or plasma with two-step incubation procedure. Polystyrene micros well are pre-coated with purified activated recombinant HEV antigen. The HRP conjugated mouse anti human IgG (r chain) monoclonal antibody serves as tracer. TMB is substrate for HRP. The enzyme reaction with substrate TMB produces a color change, and the intensity of the absorbance at 450 nm indicates the presence or absence of Anti-HEV antibodies IgM in the sample.  The test is specific, sensitive, reproducible and easy to operate. It is for blood screen of HEV infection.

1. Antigen Coated Microwell Plate

2. Specimen Diluent

3. Negative Control

4. Positive Control

5.20 X Wash Buffer (dilution prior to use)

6. Enzyme Conjugant (anti human IgG -HRP)

7. Substrate A

8. Substrate B

9. Stop Solution (2M H₂SO₄)

10. Plastic Bag

11. Seal Paper

12. Manual

Blood serum samples are routinely prepared form vein. Blood plasma samples are routinely prepared with routine amount of anticoagulant such as heparin or sodium citrate. Sample can be stored at 4°C if tested within two days. Specimens not required within 3 days should be stored at -20°C. Avoid hemolysis and repetitive freeze and thaw of samples. Samples with cloud or precipitation should be centrifugated or filtered before test. Prevent serum from bacterium contamination during collecting and storing.

1. Bring ELISA Kit for Antibody IgG to Hepatitis E Virus(all reagents), and Specimens to room temperature before use (approximately 30 minutes).

2. Dilute concentrated wash buffer 1:19 with ddH₂O.

3. For each test, set one blank, two positive and three negative controls. Add 100μl positive and negative control serum into positive and negative control wells respectively.

4. Add 100 μl sample diluents in each other test wells, then at 10 μl test serum into test wells. Pipet up and down to mix the samples well.

5. Cover wells with seal paper, and then incubate 30 minutes at 37°C.

6. Discard the liquid in all wells and fill the wells with wash solution. Lay aside for 15 seconds, discard the liquid in all wells and fill the wells with wash solution. Repeat 5 times and dry wells after last wash.

7. Add 100 μl Enzyme Conjugant in each well except the blank.

8. Cover wells with seal paper, and then incubate 30 minutes at 37°C.

9. Repeat step 6.

10. Add 50μl substrate A and B respectively to each well including the blank well. Mix gently, protected from light and incubates 15 minutes at 37°C.

11. Add 50μl of stop solution into each well to stop the reaction, including blank well.

12. Measure the absorbance at 450 nm against the blank, or measure the absorbance at 450 nm/630-690 nm.

Colorimetric Method

Cut off Value calculation:

COV = 0.10 + the average OD of negative controls (If the absorbance of negative controls is below 0.05, calculate it as 0.05.If the absorbance of negative controls is above 0.05; calculate it as its original value.)

Positive OD₄₅₀ of sample ≥ COV

Negative  OD₄₅₀ of sample < COV