OnSite Duo Dengue Ag-IgG/IgM Rapid Test-Cassette (Serum / Plasma / Whole Blood) Page 1 of 2

OnSite™ Duo Dengue Ag-IgG/IgM Rapid Test

7. Wear protective clothing and disposable gloves while handling the kit reagents and clinical
specimens. Wash hands thoroughly after performing the test.
8. Users of this test should follow the US CDC Universal Precautions for prevention of
transmission of HIV, HBV and other blood-borne pathogens.
9. Do not smoke, drink or eat in areas where specimens or kit reagents are being handled.
R0062C 10. Dispose of all specimens and materials used to perform the test as bio-hazardous waste.
11. Handle the negative and positive controls in the same manner as patient specimens.
12. The test results should be read 20-25 minutes after a specimen is applied to the sample well of
INTENDED USE the device. Any results interpreted outside 20-25 minutes should be considered invalid and
The OnSite Duo Dengue Ag-IgG/IgM Rapid Test is a lateral flow immunoassay for the simultaneous must be repeated.
detection and differentiation of IgG anti-dengue virus, IgM anti-dengue virus and dengue NS1 antigen 13. Do not perform the test in a room with strong air flow, i.e. an electric fan or strong
(DEN1, 2, 3, 4) in human serum, plasma or whole blood. It is intended to be used by professionals as a air-conditioning.
screening test and provides a preliminary test result to aid in the diagnosis of infection with dengue virus.
REAGENT PREPARATION AND STORAGE INSTRUCTIONS
Any use or interpretation of this preliminary test result must also rely on other clinical findings and the
All reagents are ready to use as supplied. Store unused test devices unopened at 2-30°C. If stored at
professional judgment of health care providers. Alternative test method(s) should be considered to
2-8°C, ensure that the test device is brought to room temperature before opening. The test device is
confirm the test result obtained by this device.
stable through the expiration date printed on the sealed pouch. Do not freeze the kit or expose the kit to
SUMMARY AND EXPLANATION OF THE TEST temperatures above 30°C.
Dengue virus is an enveloped, single-stranded, positive-sense RNA virus that comprises four related but SPECIMEN COLLECTION AND HANDLING
distinct serotypes (DEN1, 2, 3, and 4). The virus is transmitted by mosquitoes of the daytime-biting
Consider any materials of human origin as infectious and handle them using standard bio-safety
Stegomyia family, principally Aedes aegypti and Aedes albopictus. Today, more than 2.5 billion people
procedures.
living in the areas of tropical Asia, Africa, Australia and the Americas are at risk for dengue infection. An
estimated 100 million cases of dengue fever and 250,000 cases of life-threatening dengue hemorrhagic Plasma
fever occur annually on a worldwide basis1-3. 1. Collect blood specimen into a lavender, blue or green top collection tube (containing EDTA,
citrate or heparin, respectively, in Vacutainer®) by venipuncture.
Serological detection is a common method for the diagnosis of infection with dengue virus. IgM
2. Separate the plasma by centrifugation.
anti-dengue virus starts to appear 3 days after initial exposure and remains in circulation for about 30-60
3. Carefully withdraw the plasma into new pre-labeled tube.
days. IgG anti-dengue virus rises around 7 days, peaks at 2-3 weeks and persists for the duration of
life4-6. Detection of antigens, such as dengue NS1, released during virus replication in the infected Serum
patient show very promising results; it enables diagnosis from the first day after the onset of fever up to 1. Collect blood specimen into a red top collection tube (containing no anticoagulants in
day 9 once the clinical phase of the disease is over, thus, allowing early detection and prompt Vacutainer®) by venipuncture.
treatment7. 2. Allow the blood to clot.
The OnSite Duo Dengue Ag-IgG/IgM Rapid Test detects IgG and IgM anti-dengue virus and circulating 3. Separate the serum by centrifugation.
dengue NS1 antigen (DEN1, 2, 3, 4) in human serum, plasma or whole blood. It can be performed within 4. Carefully withdraw the serum into a new pre-labeled tube.
20-25 minutes by minimally skilled personnel without the use of laboratory equipment. Test specimens as soon as possible after collecting. If not tested immediately, store specimens at 2-8°C
TEST PRINCIPLE for up to 5 days. For longer storage, specimens should be kept frozen at -20°C.

The OnSite Duo Dengue Ag-IgG/IgM Rapid Test contains Avoid multiple freeze-thaw cycles. Prior to testing, bring frozen specimens to room temperature slowly
two test strips (left side: Dengue IgG/IgM test; right side: and mix gently. Specimens containing visible particulate matter should be clarified by centrifugation
Dengue Ag test). before testing. Do not use specimens demonstrating gross lipemia, gross hemolysis or turbidity in order
to avoid interference with result interpretation.
The Dengue IgG/IgM Rapid Test on the left-side is a lateral
flow chromatographic immunoassay. The test strip consists Whole Blood
of: 1) a burgundy colored conjugate pad containing Drops of whole blood can be obtained by either finger tip puncture or venipuncture. Collect blood
recombinant dengue envelope antigens conjugated with specimen into a lavender, blue or green top collection tube (containing EDTA, citrate or heparin,
colloidal gold (dengue Ag conjugates) and a control antibody respectively, in Vacutainer®). Do not use hemolyzed blood for testing.
conjugated with colloidal gold, 2) a nitrocellulose membrane Whole blood specimens should be stored at 2-8°C if not tested immediately. The specimens must be
strip containing two test lines (G and M lines) and a control tested within 24 hours of collection.
line (C line). The G line is pre-coated with antibodies for the
detection of IgG anti-dengue virus, the M line is pre-coated ASSAY PROCEDURE
with antibodies for the detection of IgM anti-dengue virus, Step 1: Bring the specimen and test components to room temperature, if refrigerated or frozen. Once
and the C line is pre-coated with a control line antibody. the specimen is thawed, mix well prior to performing the assay.
The Dengue Ag Rapid Test on the right-side is a lateral flow chromatographic immunoassay. The test Step 2: When ready to test, open the pouch at the notch and remove device. Place the test device on a
strip consists of: 1) a burgundy colored conjugate pad containing antibodies to dengue NS1 antigen clean, flat surface.
conjugated with colloidal gold (dengue Ab conjugates) and a control antibody conjugated with colloidal Step 3: Be sure to label the device with specimen’s ID number.
gold, 2) a nitrocellulose membrane strip containing a test line (T line) and a control line (C line). The T Step 4:
line is pre-coated with antibodies to dengue NS1 antigen, and the C line is pre-coated with a control line For detection of Dengue IgG/IgM For detection of Dengue Ag
antibody. The antibodies to dengue NS1 recognize the antigens from all four dengue virus serotypes. 4.1 Fill the capillary tube with serum/plasma/whole 4.1 Fill the plastic dropper with specimen.
When an adequate volume of specimen is dispensed into the sample well of the test cassette, the blood specimen not to exceed the specimen line 4.2 Holding the dropper vertically, dispense 2
specimen migrates by capillary action across the cassette. Dengue NS1 antigen, if present in the as shown in the images below. drops (about 60 μL) of serum/plasma or 2
specimen, will bind to the dengue Ab conjugates. The immunocomplex is then captured on the 4.2 Holding the capillary tube vertically, dispense the drops of whole blood (about 70 μL) into
membrane by the pre-coated antibodies to dengue NS1 antigen forming a burgundy colored T line. IgG entire specimen (5 µL) into the center of the the center of the sample well (S well),
and/or IgM anti-dengue virus, if present in the specimen, will bind to the dengue Ag conjugates. The sample well (S well) making sure that there are making sure that there are no air bubbles.
immunocomplex is then captured by the pre-coated reagent forming a burgundy colored G and/or M no air bubbles. Immediately add 1 drop (about 30-40 µL)
line, respectively. Immediately add 3 drops (about 90-120 µL) of of sample diluent to the sample well (S
Suggested result interpretation: Ag positive: Early acute primary or secondary infection. IgM positive: Sample Diluent into the buffer well (B well) with well) with the bottle positioned vertically.
acute primary or secondary infection. IgG positive: secondary or past infection. IgM and IgG positive: the bottle positioned vertically.
Late primary or early secondary acute infection.
Absence of any G, M or T lines suggests a negative result. Each test contains an internal control (C
lines) which should exhibit a burgundy colored line of the immunocomplex of the control antibodies in
both the left and right panels, regardless of color development on any of the test lines. If the C line does
not develop in a panel, the test result is invalid and the specimen must be retested with another device. or
or
An invalid result in one panel does not invalidate the test result in the other panel. Specimen Specimen
line line
REAGENTS AND MATERIALS PROVIDED
1. Individually sealed foil pouches containing:
a. One cassette device
5 µL of specimen to S well 2 drops of specimen to S well
b. One desiccant
2. 5 µL capillary tubes (for Dengue IgG/IgM test)
3. Plastic droppers (for Dengue Ag test)
4. Sample diluent (REF SB-R0062, 5mL/bottle)
5. One Package Insert (instruction for use)
MATERIALS MAY BE REQUIRED AND AVAILABLE FOR PURCHASE
1. Positivia Dengue Ag Rapid Test Control Kit (Cat # C0063) contains positive control and negative
control.
MATERIALS REQUIRED BUT NOT PROVIDED
1. Clock or timer
WARNINGS AND PRECAUTIONS 3 drops of sample diluent to B well 1 drop of sample diluent to S well
For in Vitro Diagnostic Use
1. This package insert must be read completely before performing the test. Failure to follow the 20 minutes 20 minutes
insert may give inaccurate test results. Result Result
2. Do not open the sealed pouch unless ready to conduct the assay. Step 5: Set up timer.
3. Do not use expired devices. Step 6: Read results at 20 minutes. Positive results may be visible in as short as 1 minute. Negative
4. Bring all reagents to room temperature (15-30°C) before use. results must be confirmed at the end of the 25 minutes only. However, any results
5. Do not use the components in any other type of test kit as a substitute for the components in this interpreted outside of the 20-25 minute window should be considered invalid and must
kit. be repeated. Discard used device after interpreting the result following local laws
6. Do not use hemolyzed blood specimens for testing. governing the disposal of device.
Copyright 2017 by CTK Biotech, Inc.
OnSite Duo Dengue Ag-IgG/IgM Rapid Test-Cassette (Serum / Plasma / Whole Blood) Page 2 of 2

QUALITY CONTROL 5. Interference

Common substances (such as pain and fever medication, blood components) may affect the
1. Internal Control: This test contains a built-in control feature, the C line. The C line develops
performance of the OnSite Duo Dengue Ag-IgG/IgM Rapid Test. This was studied by spiking
after adding specimen and sample diluent. If the C line does not develop, review the whole
these substances into negative and positive standard controls for dengue NS1 antigen, dengue
procedure and repeat the test with a new device.
IgG and IgM. The results are presented in the following table and demonstrate, at the
2. External Control: Good Laboratory Practice recommends using external controls, positive and concentrations tested, the substances studied do not affect the performance of the OnSite Duo
negative, to assure the proper performance of the assay, particularly under the following Dengue Ag-IgG/IgM Rapid Test.
circumstances:
List of potentially interfering substances and concentrations tested:
a. A new operator uses the kit, prior to performing testing of specimens.
1. Albumin 60 g/L 5. Glucose 5.5 mmol/L
b. A new lot of test kits is used.
2. Bilirubin 20 mg/dL 6. Heparin 3,000 U/L
c. A new shipment of test kits is used.
3. Creatinine 442 µmol/L 7. Sodium citrate 3.8%
d. The temperature during storage of the kit falls outside of 2-30°C.
4. EDTA 3.4 µmol/L 8. Salicylic acid 4.34 mmol/L
e. The temperature of the test area falls outside of 15-30°C.
f. To verify a higher than expected frequency of positive or negative results. LIMITATIONS OF TEST
g. To investigate the cause of repeated invalid results.
1. The Assay Procedure and the Interpretation of Assay Result sections must be followed closely
INTERPRETATION OF ASSAY RESULT when testing for the presence of antibodies to dengue virus and dengue NS1 antigen in serum,
1. NEGATIVE RESULT: 2. INVALID: plasma or whole blood from individual subjects. Failure to follow the procedure may give
If only the C line is present, the absence of If no C line develops, the assay is invalid inaccurate results.
any burgundy color in the G, M or T lines regardless of any burgundy color in the 2. The OnSite Duo Dengue Ag-IgG/IgM Rapid Test is limited to the qualitative detection of
indicates that neither anti-dengue virus G, M or T lines as indicated below. antibodies to dengue virus and dengue NS1 antigen in human serum, plasma or whole blood.
antibodies nor dengue virus NS1 antigen Repeat the assay with a new device. The intensity of the test line does not have a linear correlation with the antibodies and NS1
are detected. The result is negative or antigen titers in the specimen.
non-reactive. 3. Information about the dengue virus serotype(s) present in a specimen cannot be provided from
this test.
4. Serological cross-reactivity with other flaviviruses is common (e.g., Japanese encephalitis, West
Nile virus, yellow fever, etc.). Therefore, it is possible that patients who were exposed to these
viruses may show some level of reactivity with this test.
5. A negative or non-reactive result for an individual subject indicates absence of detectable
dengue virus antibodies or NS1 antigen. However, a negative or non-reactive test result does not
preclude the possibility of exposure to or infection with dengue virus.
6. A negative or non-reactive result can occur if the quantity of antibodies to dengue virus or
dengue NS1 antigen present in the specimen is below the detection limits of the assay or the
antibodies and antigen that are detected are not present during the stage of disease in which a
sample is collected. For example, some patients may not produce detectable levels of IgM
antibodies in early infection or repeat infection.
3. POSITIVE RESULT: 7. Infection may progress rapidly. If the symptoms persist while the result from the OnSite Duo
IgG IgM IgG/IgM Ag Ag/IgM Ag/IgG/IgM Dengue Ag-IgG/IgM Rapid Test is negative or non-reactive, it is recommended to test with an
Positive Positive Positive Positive Positive Positive alternative method, such as PCR or ELISA.
8. Some specimens containing unusually high titers of heterophile antibodies or rheumatoid factor
may affect expected results.
9. The results obtained with this test should only be interpreted in conjunction with other diagnostic
procedures and clinical findings.
REFERENCES
1. Gubler DJ, Clark GG. Dengue/dengue hemorrhagic fever. The emergence of a global health
problem. Emerg Infect Dis.1995;1(2):55–57.
2. Gubler DJ, Trent DW. Emergence of epidemic dengue/dengue hemorrhagic fever as a public
health problem in the Americas. Infect Agents Dis 1993;2:383–393.
Specimens with positive or reactive results should be confirmed with alternative testing method(s) 3. Monath TP. Dengue: the risk to developed and developing countries. Proc Natl Acad Sci U S A
and clinical findings before a diagnosis is made. 1994;91:2395-400.
4. Price DD, Wilson SR, "Severe Dengue Infection." Medscape Reference Drugs, Diseases &
PERFORMANCE CHARACTERISTICS
Procedures, May 2011. Web. http://www.emedicine.com/EMERG/topic124.htm.
1. Limit of Detection 5. Innis BL, and Nisalak A, et al: An enzyme-linked immunosorbent assay to characterize dengue
The OnSite Duo Dengue Ag-IgG/IgM Rapid Test was found to detect NS1 protein in all 4 types of infections where denude and Japanese encephalitis co-circulate. Am J Trop Med Hygiene 1989:
dengue virus lysate I, II, III, and IV. The limit of detection is 0.25 ng/mL as determined on 40: 418-27.
recombinant dengue NS1 antigen from serotype 2 (DEN2). 6. Anonymous. Dengue hemorrhagic fever: diagnosis, treatment, prevention and control. 2nd ed.
2. Clinical Performance for Ag Test Geneva: World Health Organization, 1997.
A total of 100 specimens were collected from susceptible subjects and normal healthy control 7. Alcon S, Talarmin A., Debruyne M., et al: Enzyme-linked immunosorbent assay specific to
subjects, and tested by the OnSite Duo Dengue Ag-IgG/IgM Rapid Test and by a commercial Dengue virus type 1 nonstructural protein NS1 reveals circulation of the antigen in the blood
Dengue Ag ELISA. Comparison for all subjects is shown in the following table: during the acute phase of disease in patients experiencing primary or secondary infections. J
Clin Microbiol, 2002, 40: 376–81.
OnSite Duo Dengue Ag-IgG/IgM Rapid Test
IgM EIA Test Positive Negative Total Index of CE Symbols
Positive 20 0 20
Consult For in vitro
Negative 1 79 80 instructions for use diagnostic use only Use by
Total 21 79 100
Relative Sensitivity: 100%, Relative Specificity: 98.75%, Overall Agreement: 99.0% REF Catalog # Lot Number N Tests per kit
3. Clinical Performance for IgG Test
Store between Authorized
2-30°C Representative Do not reuse
A total of 326 specimens were collected from susceptible subjects, and tested with the OnSite
Duo Dengue Ag-IgG/IgM Rapid Test and by a commercial EIA. Comparison for all subjects is
shown in the following table: Manufacturer Date of manufacture
OnSite Duo Dengue Ag-IgG/IgM Rapid Test
IgM EIA Test Positive Negative Total
Positive 36 1 37
MDSS GmbH
S IMPLIFYING DIAGNOSTICS

Negative 2 287 289 CTK Biotech, Inc. Schiffgraben 41

Total 38 288 326 10110 Mesa Rim Road 30175 Hannover, Germany
Relative Sensitivity: 97.3%, Relative Specificity: 99.3%, Overall Agreement: 99.1% San Diego, CA 92121, USA
Tel: 858-457-8698
4. Clinical Performance for IgM Test Fax: 858-535-1739
A total of 314 specimens were collected from susceptible subjects and tested with the OnSite E-mail: [email protected]
Duo Dengue Ag-IgG/IgM Rapid Test and by a commercial EIA. Comparison for all subjects is
shown in the following table: PI-R0062C Rev. J
Date released: 2017-05-22
OnSite Duo Dengue Ag-IgG/IgM Rapid Test
English version
IgG EIA Test Positive Negative Total
Positive 31 1 32 For Export Only, Not For Re-sale In the USA
Negative 3 279 282
Total 34 280 314
Relative Sensitivity: 96.9%, Relative Specificity: 98.9%, Overall Agreement: 98.7%
4. Cross Reactivity
Specimens from other infectious diseases were tested for cross-reactivity with the OnSite Duo
Dengue Ag-IgG/IgM Rapid Test according to the standard procedure. The results showed that
the following specimens (n=1-10) did not cross-react with the OnSite Duo Dengue Ag-IgG/IgM
Rapid Test.
Chikungunya CMV HAV HBV HCV
HIV hCG H.pylori TB T. gondii
Typhoid Rubella ANA HAMA RF (up to 8,400 IU/mL)

Copyright 2017 by CTK Biotech, Inc.