Human Anti-Hepatitis A Virus ELISA kit

Sample

Serum

Species Reactivity

Human

Detection Method

cELISA

Intended Use

Anti-HAV ELISA is an enzyme immunoassay for the detection of antibodies against Hepatitis A virus in Human Serum for research use.

Contents of Kit

1.Microtiter plate, ready for use, coated with inactivated HAV- antigen. Wells are separately breakable, (8×12) wells.
2.Mouse anti-HAV antibody (100×), 100-fold concentrated solution, 1×120 μL.
3.Conjugate, ready for use, polyclonal antibody specific for mouse antibody, 1×12 mL.
4.Positive Control (PC), anti-HAV positive control, lyophilized powder.
5.Negative Control (NC), anti-HAV negative control, lyophilized powder.
6.Dilution Buffer, ready for use, 1×50 mL
7.Wash Buffer, 20-fold concentrated solution, 1×50 mL
8.Substrate, ready for use, 2×6 mL
9.Stop Solution, ready for use, 1×7 mL
10.Serum Standard (100 mIU), anti-HAV standard 100 mIU, lyophilized powder. For semi-quantitative test.

Storage

After you receive the kit, all the components should be stored in the refrigerator (4-8°C) also up to 1 year. Long term storage, improper storage conditions and large temperature fluctuation cycles may cause precipitates in the TMB solution. These precipitates should not affect the assay noticeably. Nevertheless, if you observe such precipitates, we recommend to avoid them by allowing them to sink to the bottom.

Precision

Intra-CV < 15%
Inter-CV < 15%

Sensitivity

The Detection Threshold for the assay is 0.65 mIU/mL.

General Description

After infection with the Hepatitis A-Virus, neutralizing antibodies appear at the same time of Anti- HAV of IgG-Class formation. The Anti-HAV titers 3 to 6 months after naturally occurring infections are very high, within the range of 100 to more than 300 International Units per ml (IU/ml). Even after more than 10 years the titers usually remain at more than 1 to 10 IU/ml.
A value of 10 - 30 (mIU/ml) milli International Units can be considered the minimal protective level. Since the incidence of HAV infection in children has diminished in Northern Europe in recent years, children and juveniles are predominantly Anti-HAV negative. In an adult population could be observed, that the percentage of anti-HAV positive individuals raises with increasing age: only 3.9 % of the young people (18-24 years) show antibody against HAV, in older population (≥50 years) HAV-Antibody was found in 40.3 % of the population.
In countries with less favorable sanitary conditions around the Mediterranean area, Africa or Asia, the incidence is very high. Higher risk is also given for clinical staff, especially in the pediatric.
Since a vaccine against Hepatitis A virus infections is available, vaccinations are recommended for people travelling to countries where a high risk of HAV infections exists and for health care employees.

Standard Curve

A more accurate determination of serum titer can be carried out in semi- quantitative test method.
These samples are used to estimate the anti-HAV titer based on a calibration curve. The absorbance values of each serum standard (S1 - S3) and the negative control (NC) as standard with the concentration 0 mIU/mL are given on the y-axis against the concentration (mIU/mL) of the anti- HAV antibody on the x-axis and the regression line is placed through the points.
For computational evaluation various regression methods are useful. The method with the best curve fit (at least R2 > 0.99) should be selected. Ideally, the calculation of the antibody content of the sample is performed by means of an evaluation program. An exemplary standard curve determined by 4- parameter logistics is shown in Figure.

Figure: Exemplary standard curve with negative control as standard with the concentration of 0 mIU/mL.
In the table, exemplary results of anti-HAV antibody ELISA are shown.
Table: An example of an extinction with the semi-quantitative method when using the CD standards including negative control as standard with 0 mIU/mL. The sample dilution >1:20 was carried out in dilution buffer. The calculation of the antibody content of the sample was performed using an evaluation program.