Typhoid

Typhoid Rapid Test Cassette
(Whole Blood/Serum/Plasma)
Package Insert

TYPO01 - English

A rapid test for the qualitative detection of IgG and IgM antibodies to Salmonella typhi (S. typhi) in human whole blood, serum or plasma specimen.
For professional in vitro diagnostic use only.

[INTENDED USE]
The Typhoid Rapid Test Cassette is a rapid chromatographic immunoassay for the simultaneous detection and differentiation of IgG and IgM types of antibodies against Salmonella typhi (S. typhi) in human whole blood, serum or plasma. It is intended to be used as a screening test as an aid in the diagnosis of infection with S. typhi. Any reactive specimen with the Typhoid rapid test cassette needs to be confirmed with alternative testing method.

[SUMMARY]
Typhoid fever is caused by S. typhi, a Gram-negative bacterium. World-wide an estimated 17 million cases and 600,000 associated deaths occur annually¹. Patients who are infected with HIV are at significantly increased risk of clinical infection with S. typhi². Evidence of H. pylori infection also presents an increase risk of acquiring typhoid fever. 1-5% of patients become chronic carrier harboring S. typhi in the gallbladder.
The clinical diagnosis of typhoid fever depends on the isolation of S. typhi from blood, bone marrow or a specific anatomic lesion in the facilities that cannot afford to perform this complicated and time consuming procedure, Widal test (also referred as Weil-Felix Test) is used to facilitate the diagnosis. However, many limitations lead to difficulties in the interpretation of the Widal test ^{3, 4}.
In contrast, the Typhoid Rapid Test Cassette is a simple and rapid laboratory test. The test simultaneously detects and differentiates the IgG and the IgM antibodies to S. typhi specific antigen,⁵ in whole blood, serum or plasma thus aid in the determination of current or previous exposure the S. typhi.

[PRINCIPLE]
The Typhoid Rapid Test Cassette is a qualitative, membrane based immunoassay for the detection of antibodies (IgG and IgM) to Salmonella typhi (S. typhi) in human whole blood, serum or plasma. The diagnostic test cassette consists of two components: an IgG component and an IgM component. The IgG line region is pre-coated with reagents for the detection of anti-S. typhi (IgG). The IgM line region is pre-coated with monoclonal anti-human IgM for detection of anti-S. typhi (IgM).
During testing, specimen dispensed into the sample well of the test cassette binds with Typhoid conjugates impregnated in the reagent area, if the specimen contains anti-Typhoid antibodies. The immunocomplex thus formed migrates by capillary action. If the present antibodies in specimen are of IgG types, the immunocomplex is then captured by the pre-coated reagents on the membrane, forming a coloured IgG line, indicating a S. typhi IgG positive test result. If the present antibodies in the specimen are of IgM type, the immunocomplex would be captured on the membrane by the pre- coated anti-human IgM antibody, forming a coloured IgM line, indicating a S. typhi IgM positive test result.
Absence of any T lines (IgM and IgG) indicates a negative result. A coloured control line (C) should always appear in case of a positive or a negative result. Its absence indicates invalid test results.

[REAGENTS]
The test contains mouse anti-human IgM, mouse anti-human IgG and Typhoid antigen. A goat antibody is employed in the control line system.

[PRECAUTIONS]

1.	For in vitro diagnostic use only. Do not use after the expiration date.
2.	Do not smoke, drink, or eat in areas where specimens or kit reagents are handled.
3.	Dispose of all specimens and materials used to perform the test as bio-hazardous waste.
4.	This package insert must be read completely before performing the test.
5.	Bring all reagents to room temperature (15°C-40°C) before use.
6.	Do not interchange the buffer and test cassettes of different lots.
7.	Do not use hemolyzed blood specimens for testing.
[STORAGE AND STABILITY] Store as packaged in the sealed pouch at room temperature or refrigerated (2-30°C). The test is stable through the expiration date printed on the sealed pouch. The test must remain in the sealed pouch until use. DO NOT FREEZE. Do not use beyond the expiration date. [SPECIMEN COLLECTION AND PREPARATION]

*	The Typhoid Rapid Test Cassette (Whole Blood/Serum/Plasma) can be performed using whole blood (from venipuncture or fingerstick), serum or plasma.
*	To collect Fingerstick Whole Blood specimens:
	•	Wash the patient's hand with soap and warm water or clean with an alcohol swab. Allow to dry.
	•	Massage the hand without touching the puncture site by rubbing down the hand towards the fingertip of the middle or ring finger.
	•	Puncture the skin with a sterile lancet. Wipe away the first sign of blood.
	•	Gently rub the hand from wrist to palm to finger to form a rounded drop of blood over the puncture site.
	•	Add the Fingerstick Whole Blood specimen to the test by using the dropper:
		°	Touch the end of the dropper to the drop of blood and draw up approximately 40 µL (1 drop). Avoid air bubbles.
		°	Then squeeze the bulb to dispense the whole blood into the specimen well of the test cassette.
*	Seperate the serum or plasma from blood as soon as possible to avoid hemolysis. Only clear non- hemolyzed specimens can be used.
*	Testing should be performed immediately after the specimens have been collected. Do not leave the specimens at room temperature for prolonged periods. Serum and plasma specimens may be stored at 2-8°C for up to 3 days. For long term storage, specimens should be kept below -20°C. Whole blood collected by venipuncture should be stored at 2-8°C if the test is to be run within 2 days of collection. Do not freeze whole blood specimens. Whole blood collected by fingerstick should be tested immediately.
*	Bring specimens to room temperature prior to testing. Frozen specimens must be completely thawed and mixed well prior to testing. Specimens should not be frozen and thawed repeatedly.
*	If specimens are to be shipped, they should be packed in compliance with regulations for transportation of etiologic agents.
[MATERIALS ] Materials provided • Test cassettes • Sample droppers • Buffer • Lancets • Package insert • Alcohol Swabs Materials required but not provided • Specimen collection containers • Centrifuge • Timer

[DIRECTIONS FOR USE] Allow test cassette, specimen, and/or controls to equilibrate to room temperature (15-30°C) prior to testing.
1.	Bring the pouch to room temperature before opening it. Remove the test cassette and dropper from the sealed pouch and use it as soon as possible.
2.	Place the cassette on a clean and level surface. For Serum or Plasma specimen: Hold the dropper vertically and transfer 1 drop of serum or plasma (approximately 40µL) to the specimen well, then add 2 drops of buffer (approximately 80µL),and start the timer, see illustration below. For Whole Blood specimen: Hold the dropper vertically and transfer 1 drop of whole blood (approximately 40µL) to the specimen well, then add 2 drops of buffer (approximately 80µL), and start the timer. See illustration below.
3.	Wait for the coloured line(s) to appear. Read the result at 15 minutes, do not interpret the result after 20 minutes
[INTERPRETATION OF RESULTS] (Please refer to the illustration above) *POSITIVE: Two or three lines appear.** One coloured line should always appear in the control line region (C) and another one or two apparent coloured line(s) should be in the test line region(s) (IgM and/or IgG).
	IgM Positive: Along with line in Control region (C), a line appears in IgM region. It indicates a positive Test result for antibodies to S. typhi (Isotype IgM) IgG Positive: Along with line in Control region (C), a line appears in IgG region. It indicates a positive Test result for antibodies to S. typhi (Isotype IgG)
NOTE: The intensity of the colour in the test line regions (IgM and IgG) may vary depending on the concentration of Typhoid antibodies present in the specimen. Therefore, any shade of colour in the test line region (IgM and/or IgG) should be considered positive. NEGATIVE: One coloured line appears in the control line region (C).* No line appears in the test line regions (IgM and IgG). 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. [QUALITY CONTROL] Internal procedural controls are included in the test. A coloured line appearing in the control region (C) is an internal positive procedural control. It confirms sufficient specimen volume and correct procedural technique. [LIMITATIONS]
1.	The assay procedure and the test result interpretation must be followed closely when performing the assay. Failure to follow the procedure may give inaccurate results.
2.	The Typhoid Rapid Test Cassette is for qualitative detection of antibodies to S. typhi in human whole blood, serum or plasma. The intensity of the test band has not linear correlation with the antibody titer in the specimen.
3.	A negative result only indicates absence of anti-S. typhi antibodies above detectable levels. A negative test result does not preclude the possibility of exposure to S. typhi as a negative result can occur if the quantity of anti-S typhi antibodies present in the specimen is below the detection limit of the assay.
4.	Specimens containing unusually high titer of heterophile antibodies or rheumatoid factor may affect expected results.
5.	The results obtained with this test should only be interpreted in conjunction with other diagnostic procedures and clinical findings.
[EXPECTED VALUES] The Typhoid Rapid Test Cassette (Whole Blood/Serum/Plasma) has been compared with a leading commercial Typhoid ELISA test. The correlation between these two systems is over 98%. [PERFORMANCE CHARACTERISTICS] Sensitivity and Specificity A clinical evaluation was conducted comparing the results obtained using the Typhoid Rapid Test Cassette to Typhoid IgG/IgM ELISA Testing. The study included 15 IgG specimens and 33 IgM specimens. For the IgG specimens both assays identified 298 negative and 1 positive result. For the IgM specimens both assays identified 298 negative and 3 positive results. IgM Results Sensitivity: 93.9% (95%CI: 79.8%~99.2%) Specificity: 99.0% (95%CI: 97.1%~99.8%) Accuracy: 98.5% (95%CI: 96.5%~99.5%) Confidence Intervals IgG Results Sensitivity: 86.7% (95%CI: 59.5%~89.3%) Specificity: 99.7% (95%CI: 98.2%~99.9%) Accuracy: 99.0% (95%CI: 97.2%~99.8%) Confidence Intervals Precision Intra-Assay Within-run precision has been determined by using 10 replicates of three specimens: a negative, a low positive, and a high positive. The negative, low positive, and high positive values were correctly identified >99% of the time. Inter-Assay Between-run precision has been determined by 10 independent assays on the same three specimens: a negative, a low positive, and a high positive. Three different lots of the Typhoid Rapid Test cassette (Whole Blood/Serum/Plasma) have been tested over a 3-day period using negative, low positive, and high positive specimens. The specimens were correctly identified >99% of the time. Cross-reactivity The Typhoid Rapid Test Cassette (Whole Blood/Serum/Plasma) has been tested for HBsAg, HBsAb, HbeAg, HBeAb, HBcAb, HCV, HIV, Syphilis, H. Pylori, CMV, Rubella and Toxo positive specimens. The results showed no cross-reactivity. Interfering Substances The following potentially interfering substances were added to Typhoid negative and positive specimens.

Acetaminophen: 20 mg/dL	Caffeine: 20 mg/dL
Acetylsalicylic Acid: 20 mg/dL	Gentisic Acid: 20 mg/dL
Ascorbic Acid: 2g/dL	Albumin: 2 g/dL
Bilirubin: 1g/dL	Oxalic Acid: 600mg/dL

None of the substances at the concentration tested interfered in the assay. [BIBLIOGRAPHY]
1.	Ivanoff BN, Leivne MM, and Lambert PH. Vaccination against typhoid fever: Present status. Bulletin of the World Health Organization 1994; 72:957-71
2.	Gotuzzo E, Frisancho O, Sanchez J, Liendo G, Carillo C, Black RE, Morris JG. Salmonella typhi or Salmonella paratyphi in an endemic typhoid area. Archives of Internal Medicine 1991;151:381-2
3.	Clegg A, Passey M, Omena MK,et al. Re-evaluation of the Widal agglutination test in response to the changing pattern of typhoid fever in the highlands of Papua New Guinea. Acta Tropica 1994;57:255-63
4.	Pang T. False positive Wiadal test I non-typhoid Salmonella infection. Southeast Asian Journal of Tropical Medicine and Public Health 1989;20:163-4
5.	Ismail A, Hai OK, Kader ZA. Demonstration of an antigenic protein specific for Salmonella typhi,Biochem Biophys Res Commun.1991;181(1):301-5

	ICT INTERNATIONAL 271 De Goed Hoop Estate Noordhoek Cape Town South Africa www.ictdiagnostics.co.za Tel: +27 82 441 1922 Email: russellg@icon.co.za
Effective date: 3rd April, 2019