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Mononucleosis
Test
A Rapid Visual Test for the Qualitative Detection
of Infectious Mononucleosis Heterophile Antibody.
This test is NOT AVAILABLE FOR RETAIL SALE.
FOR HEALTHCARE PROFESSIONAL PURCHASES ONLY.
INTENDED USE
The Mono Test is a rapid test for the visual, qualitative detection
of Heterophile antibodies specific to Infectious Mononucleosis (IM)
in human serum, plasma or whole blood. This test kit is intended
as an aid in the diagnosis of IM in patients with characteristic
clinical symptoms, and is intended for profes-sional laboratory
use only. This information can be used by the physician and the
patient for disease management.
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SUMMARY
Infectious Mononucleosis is an acute, self-limiting disease caused
by the Epstein-Barr virus (EBV). Infection with EBV in early life
usually is asymptomatic. However, up to 50% of infection occurring
in young adulthood and adolescence will develop clinical manifestations
associated with IM. Diagnosis of IM is based on the evaluation of
characteristic clini-cal symptoms and serological changes. Serological
diagnosis of IM has been demonstrated by the detection of heterophile
and EBV specific antibodies. The Heterophile antibody is detectable
at some point during IM in most adults. It is a widely accepted
practice among physicians to use the detection of het-erophile antibodies
as an aid in the diagnosis of IM. The Mono Test utilizes bovine
erythrocyte extract which has a higher sensitivity and specificity
than extracts from other species.
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TEST PRINCIPLE
The Mono Test has been designed to detect IM through visual interpretation
of color development in the test device, which is a sandwich solid
phase gold conjugate immunoassay. The test device contains a membrane
strip which is pre-coated with heterophile antigens on the test
band region and goat anti-mouse antibody on the control band region.
The anti-human IgM antibody-colloidal gold conjugate pad is placed
at the end of the membrane. A mix-ture of colloidal gold conjugate
together with the sample and developer buffer will move along the
membrane chromato-graphically by capillary action. When the IM heterophile
anti-bodies are present in the patient sample, the mixture will
migrate to the test band region and form a visible line as the antibody
complexes with the heterophile antigen. When IM heterophile antibodies
are absent from the sample, no visible color band will form on test
line region. Therefore, the presence of a colored band on the test
line region indicates a positive result. A colored band will always
appear at the control region. This control band serves as a procedural
indicator for the proper performance of the test and the device.
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STORAGE AND STABILITY
The test kit is to be stored at refrigerated (2-8°C) or at
room tem-perature (up to 30°C) in the sealed pouch for the duration
of the shelf-life.
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PRECAUTIONS
- FOR IN-VITRO DIAGNOSTIC USE ONLY.
- Do not use test kit beyond expiration date.
- Do not mix reagents from different lots.
- All patient samples should be treated as if capable of transmitting
disease.
- Do not use whole blood stored for more than three days.
- Grossly hemolyzed samples should not be used.
- Developer Buffer and Controls contain sodium azide. Sodium
azide may react with lead or copper plumbing to form potentially
explosive metal azides. When disposing of these solutions, always
flush with copious amounts of water to prevent azide buildup.
- Warning: Potential Biohazardous Material Each donor unit of
human plasma or serum used in the preparation of the Positive
and Negative Controls was tested by FDA-approved methods for the
presence of anti-HIV-1/HIV-2, HBsAg and anti-HCV, and found to
be negative. However, caution should be used when handling and
disposing of these items at biosafety level 2, as recommended
in the Center for Disease Control/National Institutes of Health
Manual, Biosafety in Micobiological and BiomedicalLaboratories,
1984.
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REAGENTS AND MATERIALS SUPPLIED
- 25 individually wrapped test devices. Each test cassette contains
one test strip with heterophile antigen coated membrane and colored
antibody pad.
- Developer Buffer (8 ml): 0.1 M Phosphate Buffered Saline, with
additives and 0.1% sodium azide.
- Mono Negative Control (0.2 ml): Normal human plasma or serum
diluted in saline solution with 0.2% sodium azide.
- Mono Positive Control (0.2 ml): IM heterophile antibody positive
human plasma or serum diluted in saline solution with 0.2% sodium
azide.
- 25 disposable Transfer Pipets.
- One Instruction Sheet.
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SPECIMEN COLLECTION AND HANDLING
Fingerstick
1. Clean the area to be lanced with an alcohol swab.
2. Squeeze the end of the fingertip and pierce with a sterile
lancet.
3. Wipe away the first drop of blood with sterile gauze or cotton.
4. Allow the second drop to flow directly into the sample well
of the test device or use the pipette provided to obtain fresh
blood. Add 2 drops (about 40 µl) into the sample well.
Whole Blood
1. A certified phlebotomist should collect whole blood into a
purple, blue or green top collection tube (containing EDTA, citrate
or heparin, respectively) by venipuncture.
2. The whole blood may be used for testing immediately or may
be stored at 2-8°C up to three days.
Plasma
1. A certified phlebotomist should collect whole blood into a
purple, blue or green top collection tube (containing EDTA, citrate
or heparin, respectively) by venipuncture.
2. Separate the plasma by centrifugation.
3. Carefully withdraw the plasma for testing or label and store
at 2-8°C for up to two weeks. Plasma may be frozen at -20°C
for up to one year.
Serum
1. A certified phlebotomist should collect whole blood into a
red top collection tube (containing no anticoagulants) by venipuncture.
2. Allow the blood to clot and separate serum by centrifugation.
3. Carefully withdraw the serum for testing, or label and store
at 2-8°C for up to two weeks. Serum may be frozen at -20°C
for up to one year.
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TEST PROCEDURE
Assay Procedure
1. Review specimen collection instructions.
Test device, Developer Buffer, patient's samples, controls should
be brought to room temperature (20 to 30°C) prior to testing.
Do not open pouches until ready to perform the assay. Remove the
test device from its protective pouch. (Bring the device to room
temperature before opening to avoid condensation of moisture on
the membrane). Label the device with patient or control identification.
2. Add specimen to sample well:
a). For fingertip blood: Allow the second drop to flow directly
into the sample well of the test device or use the pipet provided
to obtain fresh blood. Add 2 drops (about 40 µl) into the
sample well.
b). For whole blood samples in collection tubes: add two drops (about
40 µl) into the sample well, holding the provided transfer
pipet in a vertical position.
c). For plasma or serum samples: add 1 drop (about 20 µl)
of serum or plasma into the sample well, holding the provided transfer
pipet in a vertical position.
3. Immediately add 3 - 4 drops of Developer Buffer.
4. After the addition of the Developer Buffer wait for the pink-red
colored bands to appear. Depending on the concentration of heterophile
antibodies present, positive results may be observed within 3 minutes.
However, to confirm a negative result, the complete reaction time
of 5 minutes is required. Do not read test results after 8 minutes.
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INTERPRETATION OF RESULTS
1. POSITIVE: Two pink-red colored bands appear. One in the control
region (C) and one pink-red colored band in the test region (T).
When testing with strong positive samples, the intensity of the
control band may be lighter than expected. It is not recommended
to compare the intensity of the lines.
2. NEGATIVE: Only one pink-red colored band appears in the control
line region. No apparent faint pink or red colored band on the test
line region (T).
3. INVALID: A total absence of pink colored bands in both regions
is an indication of procedural error or that test reagent deterioration
has occurred.
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QUALITY CONTROL
Internal Procedural Control
1. A procedural control is included in the test. A colored band
appearing in the control region (C) is considered an internal procedural
control, indicating proper performance and reactive reagents.
2. A clear background in the result window is considered an internal
negative control. However, when whole blood sam-ples are tested,
the background may appear slightly reddish due to the low level
hemolysis of some red blood cells. This is acceptable as long as
it does not interfere with the reading of the test. The test is
invalid if the background fails to clear and obscures the reading
of the test result.
External Quality Control
1. Positive Control: Add 1 drop (about 20 µl) of Positive
Control into the sample well using the (provided) transfer pipet
by holding the pipet in a vertical position. Immediately add 3 to
4 drops of the Developer Buffer. A positive signal is indicated
by the development of two pink to red colored bands in the test
region (T) and control region (C).
2. Negative Control: Add 1 drop (about 20 µl) of Negative
Control into the sample well using the (provided) transfer pipet
by holding the pipet in a vertical position. Immediately add 3 to
4 drops of the Developer Buffer. A negative signal is indicated
by the development of only one pink to red colored band in control
region (C).
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LIMITATIONS
1. This test kit is to be used for the qualitative detection of
IgM antibodies to IM heterophile antigen. A positive result suggests
the presence of IgM antibodies to heterophile antigen.
2. This test kit should be used for symptomatic individuals suspected
of having IM. Diagnosis of IM should be made by confirmation with
other clinical findings.
3. A negative result does not rule out the possibility of IM because
the antibodies to heterophile antigen may be absent or may not be
present in sufficient quantity to be detected.
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EXPECTED RESULTS
1. During the acute phase of IM, heterophile antibodies are detectable
in 80-85% of patients. Heterophile antibodies are detectable during
first month of illness and decrease rapidly after week four.
2. Positive results may be persistent for months or even years.
3. Some segment of the population who contract IM do not produce
measurable level of heterophile antibodies. Approximately 50% of
children under 4 years old who have IM may test negative.
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PERFORMANCE CHARACTERISTICS
A. Accuracy
The accuracy of the Mono Test was evaluated in com-parison to a
commercially available qualitative color immuno-chromatographic
assay for the detection of IM IgM heterophile antibodies in human
serum, plasma or whole blood. Four hun-dred and six human serum,
plasma and whole blood samples (288 serum, 103 plasma and 15 whole
blood) were used in the comparison study. The comparison results
are summarized in Table 1.
The results indicated that the Mono Test, demonstrated a Sensitivity
of 98.6% (146/148), Specificity of 98.8% (255/258), and a total
agreement of 98.8% (401/406).
B. Precision/ Site Study
The precision of the Mono Test has been evaluated at ABI and at
two other sites, including a physician's office and an independent
clinical laboratory. Out of 27 positive samples with different levels,
all (27) results were positive. Out of 33 negative samples, all
(33) results were negative. The results obtained from all site studies
demonstrated 100% agreement with the expected result. In addition,
one posi-tive and one negative control were tested each day. Results
showed 100% agreement for control specimens.
C. Sensitivity:
Since there is no sensitivity standard established for IM heterophile
anti-bodies, the following dilution (test sensitivity) studies were
per-formed for comparison purposes. Two mono positive human sera
purchased from suppliers were used for the serial dilution in a
mono negative human serum. The results of the test sensitivity study
are summarized in Tables 2a and 2b.
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