hCG Test for the early detection of Pregnancy

Sandwich immunoassay in cassette format, individually wrapped, for the fast detection of human chorionic gonadotropin (hCG) in urine. In vitro diagnostic agent for professional use.

INTENDED USE The ultimed hCG Cassette Pregnancy Test is a rapid chromatograph immunoassay for the qualitative detection of hCG in urine to aid in the early detection of Pregnancy The test indicates if a defined cut-off value is exceeded or not.

SUMMARY The human chorionic gonadotropin hormone (hCG) is a glucoprotein consisting of an  and a -subunit, which is passed initially by the fertilised ovum and after implantation of the ovum in the mucous membrane of the uterus by this into the maternal blood circulation. In the normal course of pregnancy, hCG can be detected in the serum around seven days after fertilisation of the ovum (1 – 4). The hCG concentration has frequently risen above 100 mIU/ml 14 days after fertilisation of the ovum, therefore at the time when the otherwise expected period is missed. Concentration peaks of 200,000 mIU/ml and more can be observed in the 10th to 12th week of pregnancy (2 – 4). The early rise of the hCG concentration even before the period is missed qualifies the hCG hormone for the early detection of pregnancy.

The ultimed hCG pregnancy test is set to a sensitivity of 25 mIU/ml. Both subunits are acquired by complex formation in the course of the test. Monoclonal antibodies, which because of their immunological specificity do not interact with the structurally related human follicle-stimulating hormone (hFSH), human luteinising hormone (hLH) and human thyroid-stimulating hormone (TSH) if these are present in physiological concentrations, are used in the test for selective detection of hCG concentrations in urine samples.

TEST PRINCIPLES The ultimed hCG pregnancy test is a one-step sandwich immunoassay for the fast detection of hCG in urine. The special membrane is placed in front of a reaction pad which contains colloidal gold particles coated with monoclonal anti-hCG antibodies. If a sample is applied, the colloidal gold particles dissolve in the liquid sample. If the sample contains the hCG hormone, this is bonded to the monoclonal antibodies marked with colloidal gold particles. The dissolved gold particles are transported through the membrane due to the capillary forces effective in the special membrane. In the area of the T-line, the anti-hCG antibodies immobilised there form the complex of hCG and colloidal gold. A more or less intensive red line is formed depending on the hCG concentration. The surplus colloidal gold particles are then bonded in the area of the C-line by the antibodies immobilised there, so that a red line also becomes visible in this area. This line serves as an internal functional check and must be formed in every test. If no, or only very little, hCG is contained in the sample (< 25 mIU/ml), the hormone-gold particle complex is not formed or it is formed in insufficient amount to generate a visible red line in the area of the T-line.

STORAGE AND STABILITY The ultimed hCG pregnancy test can be stored at room temperature (2° to 30°C). The test can be used up to the time printed on the packaging. The test cassette must be kept in the packaging foil until immediately before use.

NOTES For in-vitro diagnostics use only Single use only Do not use after expiry date. Do not use if the protective foil is damaged. The test cassette must reach room temperature before use.

MATERIALS PROVIDED 1. Test cassette with coated special membrane and sample dropper pipette in sealed protective foil. 2. Instructions for use.

MATERIALS REQUIRED BUT NOT PROVIDED 1. Sample collection cup ( available from Omega Diagnostics Irl.) 2. Timer

URINE SPECIMEN COLLECTION

As the highest hormone concentrations are present in the morning , morning urine should be used if possible for early detection of pregnancy.

Note: urine specimens and all materials that have been in contact with urine should be considered, treated and disposed off as infectious material. A contact with the skin is to be avoided by wearing protective gloves and proper protective clothing.

1 DIRECTIONS FOR USE

The urine specimen, test cassette and any controls should be at room temperature before the test is performed. Open the protective foil shortly before performing the test, as the test membrane is sensitive to humidity. Use the dropper /pipette enclosed in each pouch for applying the sample to the cassette.

1. Mark the test card with the relevant patient id number directly after it is taken out of the protective foil. 2. Using the enclosed disposable sample dropper, drop 3 drops of sample (~ 120 µl) into the sample opening (S) of the test card. 3. Wait until the chromatography has finished i.e. that the liquid front has passed the evaluation window. 4. Evaluate the test after 3-5 minutes. The confirmation of a negative result should be done at 10 minutes. 5. It is important that the background is clear when the result is read.

Note: The ultimed hCG pregnancy test is checked continuously with regard to quality, but it is recommended that an internal Quality control is made with a control solution.The necessary control solutions for internal quality control ( Urinalysis Controls) are available from Omega Diagnostics ([email protected]) as required.

INTERPRETATION OF THE TEST RESULTS

positive negative invalid invalid

Reproductions may vary from original! Positive (hCG > 25 mIU/ml) Reddish discolouration is visible both in the area of the T-line and in the area of the C-line. The intensity of the T-line depends on the concentration of the hCG and is usually less than that of the C-line. The different intensities of the T-line are not suitable for quantitative measurement of the hormone concentration.

Negative (hCG < 25 mIU/ml) Red discolouration is present only in the area of the C-line.

Invalid If, after 10 minutes. a red line can be seen neither in the T area nor in the C area or only in the test area, then the test must be classified as invalid and it must be repeated with a new test cassette. It may be necessary to check the affected test batch with a control solution.(Urine Control available at [email protected])

NOTE ON INTERPRETATION OF THE TEST RESULTS

In the case of an expected pregnancy, a negative result should be checked 2 - 3 days later with a new urine specimen or checked with a quantitative Laboratory test. Borderline cases, in which only a weak reddish (T- line) appears, should be checked 2 - 3 days later with a new urine specimen. If on repeated testing you receive a definite negative result, this could indicate a miscarriage

LIMITATIONS OF THE TEST METHOD A rise in hCG concentration is not attributable to pregnancy in all cases. Neoplastic processes can also bring about a rise in the hCG concentration. Urine samples with low specific gravity can contain too little hormone, so that repetition 2 to 3 days later with morning urine is recommended. The ultimed hCG pregnancy test is a diagnostic test, the result should be confirmed by further clinical examination results by the attending physician. Medication taken in the course of antibody therapy can falsify the result. At very high hCG concentrations (~ 200,000 mIU/ml) the free hCG can block the formation of a red line in the T area. In such cases, repeat the test with a diluted sample (1:10 or 1:20). Heterophilic antibodies as well as non-specific protein binding can lead to false positive results. If the test result does not fit in the clinical picture, then the test should be repeated with another method.

2 EXPECTD VALUES The ultimed hCG pregnancy test produces a negative result (hCG < 25 mIU/ml in healthy, not pregnant women and healthy men. At the beginning of pregnancy the hCG concentration doubles on average every 2 - 3 days. Concentrations around 20 mIU/ml can be observed after 8 - 10 days. The highest concentration is reached in the 8th to 10th week and the concentration then drops, so that only slightly elevated values are present at the end of pregnancy. QUALITY CONTROL

The ultimed hCG pregnancy test contains an internal control, the C-line, which indicates the effectiveness of the reagents. In addition, the function of the special membrane can be checked by reference to the background colour. If the membrane has a clear white background after the test is performed, then chromatography has occurred without errors. The ultimed hCG pregnancy test is checked continuously with regard to its quality, but it is recommended that internal laboratory control is performed using a quality control solution when a new batch is used or if the test is performed for the first time. Liquid urine quality controls containing hCG are available from Omega Diagnostics (Ireland) Ltd.

SPECIFICITY The possible interactions and therefore false positive results have been checked with the structurally similar hormones like human follicle-stimulating hormone (hFSH), human luteinizing hormone (hLH) and human thyroid-stimulating hormone (hTSH). At the below mentioned concentrations there was no discolouration observed at the indicated concentrations in the test region. hLH 300 no hTSH 1000 no hFSH 1000 no

CROSS REACTIONS The following substances were added to both positive and negative urine:

Acetaminophene 20 mg/ml Caffeine 20 mg/ml Acetyl salicylic acid 20 mg/ml Gentisic acid 20 mg/ml Ascorbic acid 20 mg/ml Glucose 2 g/dl Atropine 20 mg/ml Haemoglobin 1 mg/dl Bilirubin 2 mg/dl

No cross reactions were observed.

BIBLIOGRAPHY

1. Batzer FR. “Hormonal evaluation of early pregnancy”, Fertil. Steril. 1980; 34(1): 1-13 2. Catt KJ, ML Dufau, JL Vaitukaitis “Appearance of hCG in pregnancy plasma following the initiation of implantation of the blastocyte”, J. Clin. Endocrinol. Metab. 1975; 40(3): 537-540 3. Braunstein GD, J Rasor, H. Danzer, D Adler, ME Wade “Serum human chorionic gonadotropin levels throughout normal pregnancy”, Am. J. Obstet. Gynecol. 1976; 126(6): 678-681 4. Lenton EA, LM Neal, R Sulaiman “Plasma concentration of human chorionic gonadotropin from the time of implantation until the second week of pregnancy”, Fertil. Steril. 1982; 37(6): 773-778 5. Steier JA, P Bergsjo, OL Myking “Human chorionic gonadotropin in maternal plasma after induced abortion, spontaneous abortion and removed ectopic pregnancy”, Obstet. Gynecol. 1984; 64(3): 391-394 6. Dawood MY, BB Saxena, R Landesman “Human chorionic gonadotropin and its subunits in hydatidiform mole and choriocarcinoma”, Obstet. Gynecol. 1977; 50(2): 172-181 7. Braunstein GD, JL Vaitukaitis, PP Carbone, GT Ross “Ectopic production of human chorionic gonadotropin by neoplasms”, Ann. Intern Med. 1973; 78(1): 39-45

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