Nebraska Perfect Lab Corp.

Pathology Department Urinalysis Section Omaha, NE

Human Chorionic Gonadotropin (hCG) in Urine Specimens Rapid Chromatographic Immunoassay Method AUTHOR: Nancy Strong REPLACES:
DATE TECHNOLOGIST IN CHARGE SUPERVISOR LABORATORY DIRECTOR

ADOPTED REVIEWED/REVISED REVIEWED/REVISED REVIEWED/REVISED REVIEWED/REVISED

9/21/09

N. Strong

J. Richards

Dr. Williams

PRINCIPLE The SP Brand Rapid Test hCG Cassette is a Rapid chromatographic immunoassay for the qualitative detection of human chorionic gonadotropin (hCG) in urine to aid in the early detection of pregnancy. The SP Brand Rapid Test hCG Cassette utilizes a combination of mouse monoclonal anti-hCG antibodies and goat polyclonal anti-hCG antibodies to selectively detect elevated levels of hCG in urine. The assay is conducted by adding a urine specimen to the specimen well of the test cassette and observing the formation of colored lines. The specimen migrates via capillary action along the membrane to react with the colored conjugate. Positive specimens react with the specific colored antibody conjugates and form a colored line at the test line region of the membrane. Absence of this colored line suggests a negative result. To serve as a procedural control, a colored line will always appear at the control line region if the test has been performed properly. The test qualitatively detects the presence of hCG in urine specimens at the sensitivity of 25 mlU/ml. At the levels of claimed sensitivity the SP Brand Test hCG Cassette shows no crossreactivity interference from the structurally related glycoprotein hormones hFSH, hLH and hTSH at high physiological levels.

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CLINICAL SIGNIFICANCE Human chorionic gonadotropin (hCG) is a glycoprotein hormone produced by the developing placenta shortly after fertilization. After fertilization, the concentration of hCG rapidly rises in both the urine and serum of pregnant women. In normal pregnancy, hCG can be detected in both urine and serum as early as 7 to 10 days after conception. hCG levels continue to rise very rapidly, frequently exceeding 100 mlU/ml by the first missed menstrual period, and peaking in the 100,000-200,000 mlU/ml range about 10-12 weeks into pregnancy. The appearance of hCG in both the urine and serum soon after conception, and its subsequent rapid rise in concentration during early gestational growth, make it an excellent marker for the early detection of pregnancy. SPECIMEN Patient Preparation: None Type: Urine (first morning urine specimen preferred) Amount required: Optimum: 1 ml Minimum: 0.5 ml

Collection: Collect urine specimens in a clean container. Stability: Specimens may be stored at 2-8 C for up to 48 hours prior to testing. For prolonged storage, specimens are to be frozen and stored below -20 C (-4 F). Frozen specimens should be thawed and mixed before testing. Unacceptable specimens: Specimens from patients who have received preparations of Mouse Monoclonal Antibodies for diagnosis or therapy may contain Human Anti-Mouse Antibodies (HAMA). Such specimens may demonstrate either falsely elevated or falsely depressed results when tested with assay kits which employ Mouse Monoclonal Antibodies. These specimens should not be tested with the SP Brand Rapid Test hCG Cassette Urine Test Device. REAGENTS/CALIBRATORS/CONTROLS/MATERIALS CAUTION: This product contains human source material and/or potentially infectious components. Follow Laboratory Standard Precautions at all times when handling these materials. Reagents: Individually packaged test cassettes (30): Mouse monoclonal anti-hCG antibodies and goat polyclonal anti-hCG antibodies. Each package contains a disposable pipette. Test cassettes are stable until the expiration printed on the outer box if stored at room temperature (15-30 C) and out of direct sunlight.
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Calibrators: None Controls: Sure-Vue hCG Urine Control Set: One negative control: contains 0.2% sodium azide One 25 mlU/mL hCG Positive Urine Control: contains normal human urine and 0.2% sodium azide One 250 mlU/mL hCG Positive Urine Control: contains normal human urine and 0.2% sodium azide The Control Set should be stored at 2-8 C. Observe expiration date on vials. CAUTION: The control set contains sodium azide. Sodium azide may react with lead or copper plumbing to form potentially explosive metal azides. Copious quantities of water should be used to flush the control set down a sink. If solution contacts the skin or eye, flush with copious amounts of water.

CALIBRATION: None QUALITY CONTROL Frequency/Use: 1) External controls: Sure-Vue hcG urine controls (two positive and a negative) shall be performed upon opening a new kit. 2) Internal Controls: a. Positive internal procedural control: -The development of the red procedural control line next to the letter "C". -It confirms sufficient specimen volume and correct procedural technique. -If not present, test result is invalid. b. Negative internal procedural control: -The absence of interfering background color. -If the test is working properly, the background in the result area should be white to light pink and not interfere with the ability to read the test result. -If background color appears in the result window which interferes with your ability to read the test result, the result is invalid. Preparation: Sure-Vue hCG Urine Control Set should be brought to room temperature prior to testing.

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Acceptance Limits/Corrective Action: If the valid procedural controls do not appear, the patient result is invalid. If this occurs: -Review the test procedure to ensure sufficient sample was added. -Test with a new SP Brand Rapid Test cassette. -If still invalid, consult technical coordinator. Recording: External Controls: 1. Record the result of each level of control on the manual UA/Coag log sheet. 2. Mark the appropriate space with controls okay, your initials and the date. 3. Enter results into Cerner using the appropriate QC code for each level of control as found on the UA/Coag log sheet. 4. On the outside of the test kit record the date that QC was performed and your initials. Internal Controls: Record on the manual UA/Coag log sheet that internal controls were okay. This must be recorded for each test that is performed. PROCEDURE CAUTION: When handling biohazardous materials, gloves and lab coats/gowns must be worn. 1. Remove the test cassette and pipette from the pouch and place on a flat, dry surface. Label the cassette. (Cassette must be used without delay.) 2. Use the provided pipette for testing. Hold the dropper vertically and transfer 3 full drops of urine (approx 100 ul) to the specimen well of the test cassette and then start the timer. 3. The test results should be read at three minutes. Wait for the red line(s) to appear. It is important that the background is clear before the result is read. NOTE: A low hCG concentration might result in a weak line appearing in the test region (T) after an extended period of time. Do not interpret results after 10 minutes. CALCULATIONS: None REPORTING RESULTS Positive: Two distinct red lines appear. A red line in the procedural control region (C) and a red or pink line in the test region (T) in the result window at three minutes. Record result as Positive. NOTE: The intensity of the red color in the test line region (T) will vary depending on the concentration of hCG present in the specimen. However, neither the quantitative value nor the rate of increase in hCG can be determined by this qualitative test.
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Negative: One red line appears in the control region. A red line in the procedural control region (C) and no apparent red or pink line appear in the test region (T) in the result window at three minutes. Record result as "Negative". Invalid: Absence of a red line in the procedural control region (C).

A red procedural Control Line must always appear next to the control region (C) in the Results Window. If no red Procedural Control Line appears, the test is invalid, and the specimen must be retested. NOTE: 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 cassette. If the problem persists, discontinue using the test kit immediately and notify a Tech Coordinator or charge tech. Call Technical services at 1-866-211-7853. Recording and Reporting Results: 1. Place a patient sticker or manually record the patient information in the "Name/Accession #" space on the manual UA/Coag log sheet. 2. Record the patient result in the appropriate space on the log sheet. 3. Mark the appropriate space on the log sheet with controls okay, your initials and the date. 4. Enter results into Cerner using the Accession Result Entry application. Reference Ranges: Negative Panic/Alert/Critical Values: None LIMITATIONS Linearity: N/A Detection Limits: The SP Brand Rapid Test hCG Cassette has a sensitivity of 25 mlU/mL, and is capable of detecting pregnancy as early as 1 day after the first missed menses.
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Dilution Protocol: None Interferences: Interfering substances may falsely depress or falsely elevate results. These interfering substances may cause false results over the entire range of the assay, not just at low levels, and may indicate the presence of hCG when there is none. As with any immunochemical reaction, unknown interferences from medications or endogenous substances may affect results. There are no interfering substances listed in the package insert. PROCEDURE NOTES 1. For diagnostic purposes, hCG test results should always be used in conjunction with other methods and in the context of the patients clinical information (e.g., medical history, symptoms, results of other tests, clinical impression, etc.). Ectopic pregnancy cannot be distinguished from normal pregnancy by hCG measurements alone. 2. If the hCG level is inconsistent with, or unsupported by, clinical evidence, results should also be confirmed by an alternative hCG method. The absence of urinary hCG may suggest a falsely elevated serum result. Additionally, results may be confirmed by performing serial dilutions of the sample as usually, but not always, samples that contain interfering substances exhibit nonlinear results when diluted. Test results should be confirmed using a quantitative hCG assay prior to the performance of any critical medical procedure. 3. Interfering substances may falsely depress or falsely elevate results. These interfering substances may cause false results over the entire range of the assay, not just at low levels, and may indicate the presence of hCG when there is none. As with any immunochemical reaction, unknown interferences from medications or endogenous substances may affect results. There are no interfering substances listed in the package insert. 4. Infrequently, hCG levels may appear consistently elevated and could be due to, but not limited to, the presence of the following: -Heterophilic antibodies: Patients routinely exposed to animals or to animal serum products, can be prone to this interference and anomalous values may be observed. -Trophoblastic or nontrophoblastic neoplasms: abnormal physiological states that may falsely elevate hCG levels. This test should not be used in the diagnosis of these conditions. -Nonspecific protein binding -hCG like substances REFERENCES: SP Brand Rapid Test hCG Cassette procedure booklet, January 2007.

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