COVID-19 Rapid Antigen Test Protocol

What is COVID-19?

SARS-CoV-2 is the causative viral agent of the disease COVID-19. It is primarily transmitted through respiratory droplets, and enters into cells via the ACE2 receptor found on alveolar epithelial cells (among other cell types). Symptoms of COVID-19 may appear as early as 2 days after initial infection, or as late as 14 days. There is variation in the severity of symptoms and in their duration and time of onset.

Currently, the patients infected by the novel coronavirus are the main source of infection; asymptomatic infected people can also be an infectious source. The main manifestations include fever, fatigue and dry cough. Nasal congestion, runny nose, sore throat, myalgia and diarrhea are found in a few cases.

Intended use of COVID-19 Rapid Antigen Test

The COVID-19 Antigen Rapid Test (Nasopharyngeal Swab) is a rapid chromatographic immunoassay for the qualitative detection of SARS-CoV-2 antigens in nasopharyngeal swab specimens from individuals with suspected SARS-CoV-2 infection in conjunction with clinical presentation and the results of other laboratory tests.

Results are for the detection of SARS-CoV-2 Antigens. An antigen is generally detectable in upper respiratory specimens during the acute phase of infection. Positive results indicate the presence of viral antigens, but clinical correlation with patient history and other diagnostic information is necessary to determine infection status. Positive results do not rule out bacterial infection or co-infection with other viruses. The agent detected may not be the definite cause of disease.

Negative results do not preclude SARS-CoV-2 infection and should not be used as the sole basis for treatment or patient management decisions. Negative results should be treated as presumptive and confirmed with a molecular assay, if necessary for patient management. Negative results should be considered in the context of a patient’s recent exposures, history and the presence of clinical signs and symptoms consistent with COVID-19.

The COVID-19 Antigen Rapid Test is intended for use by trained clinical laboratory personnel.

Principle

The COVID-19 Antigen Rapid Test (Nasopharyngeal Swab) is a qualitative membrane-based immunoassay for the detection of SARS-CoV-2 Antigens in human nasopharyngeal swab specimen.

SARS-CoV-2 antibody is coated in the test line region. During testing, the specimen reacts with SARS-CoV-2 antibody-coated particles in the test. The mixture then migrates upward on the membrane by capillary action and reacts with the SARS-CoV-2 antibody in test line region. If the specimen contains SARS-CoV-2 Antigens, a colored line will appear in test line region as a result of this. If the specimen does not contain antigens to SARS-CoV-2, no colored line will appear in the test line region, indicating a negative result.

To serve as a procedural control, a colored line will always appear in the control line region, indicating that the proper volume of specimen has been added and membrane wicking has occurred.

Specimen Collection, Transport and Storage

Specimen collection

Step	Procedure
1.	Insert a sterile swab into the nostril of the patient, reaching the surface of the posterior nasopharynx.
2.	Swab over the surface of the posterior nasopharynx.
3.	Withdraw the sterile swab from the nasal cavity.

Specimen transport and storage

Specimens should be tested as soon as possible after collection. If swabs are not processed immediately, it is highly recommended to place the swab into a dry, sterile, and tightly sealed plastic tube for storage. The swab specimen in dry and sterile conditions is stable for up to 8 hours at room temperature and 24 hours at 2-8°C.

If transport of samples with viral transport media (VTM) is required, the VTM should not contain guanidinium (for example Guanidine Hydrochloride and Guanidine isothiocyanate) and minimal dilution of the sample is recommended, as dilution may result in decreased test sensitivity. Whenever possible, 1 milliliter or less is best to avoid excessive dilution of the patient sample. Nasopharyngeal swabs in VTM are stable for up to 8 hours at room temperature and 24 hours at 2-8°C.

Specimen Preparation

Only the extraction buffer and tubes provided in the kit is to be used for swab specimen preparation.

Preparation with Extraction buffer with Integrated Extraction Tube

Step	Procedure
1.	Unscrew the cap of the specimen Extraction tube with Extraction buffer.
2.	Insert the swab specimen into the specimen Extraction tube. Press against the inner wall of the tube and stir the swab for approximately 10 seconds while pressing the swab head against the inner wall of the tube to release the antigens in the extraction tube.
3.	Remove the swab while squeezing the sides of the tube to extract the liquid from the swab.
4.	Tighten the cap onto the specimen extraction tube.

Figure 2: Schematic representation of Specimen Preparation

Preparation with Extraction buffer with Non-Integrated Extraction Tube

Step	Procedure
1.	Place the Extraction tube in the workstation. Add Approx. 350μl Extraction Buffer to the Extraction tube. See illustration 1.
2.	Place the swab specimen in the Extraction tube. Rotate the swab for approximately 10 seconds while pressing the head against the inside of the tube to release the antigen in the swab. See illustration 2.
3.	Remove the swab while squeezing the swab head against the inside of the Extraction Tube as you remove it to expel as much liquid as possible from the swab. Discard the swab in accordance with your biohazard waste disposal protocol. See illustration 3.
4.	Fit the dropper tip on top of the extraction tube.

Figure 3: Schematic representation of Specimen Preparation with Non-Integrated Extraction Tube

Preparation for VTM specimen

When using viral transport media (VTM), it is important to ensure that the VTM containing the sample has balanced to room temperature (15-30°C). Using a pipette, transfer 350μl of specimen solution from the VTM to the Specimen Extraction Tubes with Extraction Buffer, mix well by shaking. *NOTE: The storage of the specimen after extraction is stable for 2 hours at room temperature or 24 hours at 2-8°C.

Materials

Test cassettes
Package insert
Extraction Buffer
Extraction tubes and tips (Optional)
Workstation

Materials required but not provided

Timer
Sterile swabs

Directions for use

Allow the test, extracted specimen and/or controls to equilibrate to room temperature (15-30°C) prior to testing.

Step	Procedure
1.	Remove the test cassette from the sealed foil pouch and use it within one hour. Best results will be obtained if the test is performed immediately after opening the foil pouch.
2.	Invert the specimen extraction tube and add 3 drops of extracted specimen (approx.100μl) to the specimen well(S) and then start the timer.
3.	Wait for the colored line(s) to appear. Read the result at 15 minutes. Do not interpret the result after 20 minutes.

Figure 4: Schematic representation of the interpretation of results from the COVID-19 Test Cassette

Interpretations of results

Result	Description
POSITIVE COVID-19: Two distinct colored lines appear.	One colored line should be in the control region (C) and another colored line should be in the Test region (T). Positive result in the Test region indicates detection of SARS-COV-2 antigens in the sample. *NOTE: The intensity of the color in the test line region (T) will vary based on the amount of SARS-COV-2 antigen present in the sample. So any shade of color in the test region (T) should be considered positive.
NEGATIVE: One colored line appears in the control region (C).	No apparent colored line appears in the test line region (T).
INVALID: Control line fails to appear.	Insufficient specimen volume or incorrect procedural techniques are the most likely reasons for control line failure. Review the procedure and repeat the test with a new test. If the problem persists, discontinue using the test kit immediately and contact your local distributor.

Limitations

No.	Description
1.	The test Procedure and the Interpretation of test Result must be followed closely when testing for the presence of SARS-CoV-2 antigens in the human nasopharyngeal specimens from suspected individuals. For optimal test performance, proper sample collection is critical. Failure to follow the procedure may give inaccurate results.
2.	The performance of the COVID-19 Antigen Rapid Test (Nasopharyngeal swab) was evaluated using the procedures provided in this product insert only. Modifications to these procedures may alter the performance of the test. Viral Transport Media (VTM) may affect the test result; extracted specimens for PCR tests cannot be used for the test.
3.	The COVID-19 Antigen Rapid Test (Nasopharyngeal swab) is for in vitro diagnostic use only. This test should be used for detection of SARS-CoV-2 Antigens in human nasopharyngeal specimens as an aid in the diagnosis of patients with suspected SARS-CoV-2 infection in conjunction with clinical presentation and the results of other laboratory tests. Neither the quantitative value nor the rate of increase in the concentration of SARS-CoV-2 antigens can be determined by this qualitative test.
4.	The COVID-19 Antigen Rapid Test (Nasopharyngeal Swab) will only indicate the presence of SARS-CoV-2 Antigens in the specimen and should not be used as the sole criteria for the diagnosis of SARS-CoV-2 infections.
5.	The results obtained with the test should be considered with other clinical findings from other laboratory tests and evaluations.
6.	If the test result is negative or non-reactive and clinical symptoms persist. It is recommended to re-sample the patient a few days later and test again or test with a molecular diagnostic device to rule out infection in these individuals.
7.	The test will show negative results under the following conditions: The concentration of the novel coronavirus antigens in the sample is lower than the minimum detection limit of the test.
8.	Negative results do not rule out SARS-CoV-2 infection, particularly in those who have been in contact with the virus. Follow-up testing with a molecular diagnostic should be considered to rule out infection in these individuals.
9.	Excess blood or mucin on the swab specimen may interfere with test performance and may yield a false positive result.
10.	The accuracy of the test depends on the quality of the swab sample. False negatives may result from improper sample collection or storage.
11.	Positive results of COVID-19 may be due to infection with non-SARS-CoV-2 coronavirus strains or other interference factors.

Rapid Antigen Test Protocol