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0032 (0)16 41 44 07
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Česká republika Praha
US New York
0032 (0)16 41 44 07
Catalog No. 130BQ 033G
NAME AND INTENDED USE
The BI ANA Screen ELISA is intended for use in
the evaluation of patients with suspected autoimmune diseases.
SUMMARY AND EXPLANATION OF THE TEST
Antinuclear antibodies (ANA) are frequently present in patients with systemic
lupus erythematosus (SLE) and, less commonly, in other autoimmune
diseases Rheumatoid arthritis, Collagen vascular diseases, chronic liver
diseases and systemic sclerosis (scleroderma). ANA bind to several nuclear
antigens including DsDNA, SSDNA, RNP, Sm, SSA and SSB. ANA
frequency increases with age in apparently healthy people, especially
women after the age of 45 years.
ANA ELISA is widely used as a screening procedure for different
PRINCIPLE OF THE TEST
Diluted patient serum is added to wells coated with purified ANA antigen.
ANA IgG specific antibody, if present, binds to the antigen. All unbound
materials are washed away and the enzyme conjugate is added to bind to
the antibody-antigen complex, if present. Excess enzyme conjugate is
washed off and substrate is added. The plate is incubated to allow the
hydrolysis of the substrate by the enzyme. The intensity of the color
generated is proportional to the amount of IgG specific antibody in the
1. Microwell Strips: Nuclear Antigen coated wells (12 x 8 x 1 wells)
2. Sample Diluent: 1 bottle (22 mL). Ready to use.
3. Calibrator: Yellow Cap. (1.50 mL/vial). Ready to use.
4. Positive Control: Red Cap. (1.50 mL/vial). Ready to use.
5. Negative Control: Blue Cap. (1.50 mL/vial). Ready to use.
6. Enzyme Conjugate: 1 bottle (12 mL). Ready to use.
7. TMB Substrate: 1 bottle (12 mL). Ready to use.
8. Stop Solution: 1N H2SO4; 1 bottle (12 mL). Ready to use.
9. Wash Concentrate: 1 bottle (50 mL), 20X concentrate.
STORAGE AND STABILITY
1. Store the kit at 2 – 8° C.
2. Keep microwells sealed in a dry bag with desiccants.
3. The reagents are stable until expiration of the kit.
4. Do not expose test reagents to heat, sun or strong light during storage
WARNINGS AND PRECAUTIONS
1. Potential biohazardous materials:
The calibrator and controls contain human source components, which
have been tested and found non-reactive for hepatitis B surface antigen
as well as HIV antibody with FDA licensed reagents. However, as
there is no test method that can offer complete assurance that HIV,
Hepatitis B virus or other infectious agents are absent, these reagents
should be handled at the Biosafety Level 2, as recommended in the
Centers for Disease Control/National Institutes of Health manual,
"Biosafety in Microbiological and Biomedical Laboratories." 1984
2. Optimal results will be obtained by strict adherence to the test protocol.
Precise pipetting as well as following the exact time and temperature
requirements is essential.
3. Do not pipette by mouth. Do not smoke, eat, or drink in the areas in
which specimens or kit reagents are handled.
4. The components in this kit are intended for use as an integral unit. The
components of different lots should not be mixed.
5. This product contains components preserved with sodium azide.
Sodium azide may react with lead and copper plumbing to form
explosive metal azide. On disposal, flush with a large volume of water.
SPECIMEN COLLECTION AND HANDLING
1. Collect blood specimens and separate the serum.
2. Specimens may be refrigerated at 2–8°C for up to seven days or frozen
for up to six months. Avoid repetitive freezing and thawing of serum
PREPARATION FOR ASSAY
1. Bring all specimens and kit reagents to room temperature (20-25°C)
and gently mix.
2. Prepare washing buffer by adding the contents of the bottle (50 mL,
20X Wash concentrate) to 950 mL of distilled or deionized water in oneliter
container. Store at room temperature.
1. Place the desired number of coated strips into the holder.
2. Negative control, positive control, and calibrator are ready to use.
Prepare 1:21 dilution of test samples, by adding 10 μL of the sample to
200 μL of sample diluent. Mix well.
3. Dispense 100 μL of diluted sera, calibrator, and control into the
appropriate wells. For the reagent blank, dispense 100 μL sample
diluent in 1A well position. Tap the holder to remove air bubbles from
the liquid and mix well. Incubate for 20 minutes at room temperature.
4. Remove liquid from all wells. Repeat washing three times with washing
5. Dispense 100 μL of enzyme conjugate to each well and incubate for 20
minutes at room temperature.
6. Remove enzyme conjugate from all wells. Repeat washing three times
with washing buffer.
7. Dispense 100 μL of TMB substrate solution and incubate for 10
minutes at room temperature.
8. Add 100 μL of 1N H2SO4 to stop reaction.
9. Read O.D. on ELISA reader at 450 nm within 30 minutes after adding
CALCULATION OF RESULTS
1. Check Calibrator Factor (CF) value on the calibrator bottle. This value
might vary from lot to lot. Make sure you check the value on every kit.
2. Calculate the cut-off value: Calibrator OD x Calibrator Factor (CF).
3. Calculate the Ab (Antibody) Index of each determination by dividing the
O.D. value of each sample by cut-off value.
Example of typical results:
Calibrator mean OD = 0.8
Calibrator Factor (CF) = 0.5
Cut-off Value = 0.8 x 0.5= 0.400
Positive control O.D. = 1.2
Ab Index = 1.2 / 0.4 = 3
Patient sample O.D. = 1.6
Ab Index = 1.6 / 0.4 = 4.0
The test run may be considered valid provided the following criteria are met:
1. If the O.D. of the Calibrator should be greater than 0.250.
2. The Ab index for Negative control should be less than 0.9.
3. The Ab index for Positive control should be greater than 1.2.