hc Step Serum/Urine, hc Step Detector and HumaPreg Serum/Urine

Transcription

1 Design Verification hc Step Serum/Urine, hc Step Detector and HumaPreg Serum/Urine 1 Function Sensitivity and Dynamic Range Analytical Sensitivity Validation of Efficacy Clinical Evaluation Summary Cross-Reactivity, Interferences Description of Control Materials Results Reproducibility Within-Run Performance / Lot-to-Lot Stability Real Time Stability... 4 Form Rev. 006 valid of /5

2 1 Function HEXAGON hcg 1-STEP Serum/Urine, HEXAGON hcg 1-STEP Detector and HumaPreg Serum/Urine tests have been developed for the rapid qualitative detection of the hormone human chorionic gonadotropin (hcg) in human serum and urine. The tests are based on a membrane immunochromatographic method featuring a combination of monoclonal and polyclonal antibodies and red-violet coloured reagent. The coloured reagent, the antibodies and the nature of the membrane have been specially selected for the application with serum and urine samples. Specimen being brought into contact with the test strip/test device causes re-dissolution of the reagents which are impregnated on the strip or the inner part of the test device. Results appear typically within a short time. Specimens with hcg concentrations near the detection limit show a positive result within 4 minutes with serum as well as with urine samples. Positive results are interpreted by the appearance of two red coloured lines. The test strips used for HEXAGON hcg 1-STEP Serum/Urine, HEXAGON hcg 1-STEP Detector and for HumaPreg Serum/ Urine are identical (difference only in one cover foil). Accordingly, evaluation results obtained with test strips can be applied to test devices. 2 Sensitivity and Dynamic Range 2.1 Analytical Sensitivity Standard material (calibrated to the 4 th International Standard for chorionic gonadotropin - 75/589) was used to determine the analytical sensitivity. For this, hcg negative urine was spiked with the standard material. The results are summarised in the following table: No. hcg Concentration in IU/l Test line 1 600, , , , , , Intensity grades: 4+ (very strong intensity); 3+ (strong intensity); 2+ (medium intensity); + (visible); ± (very weakly positive); - (no intensity) To determine the sensitivity with both, serum and urine specimens with hcg concentrations close to the cut-off, standard material (Sigma, code CG5 traceable to the 4 th International Standard for chorionic gonadotropin - 75/589) was reconstituted with 10 ml of distilled water. This stock solution was diluted with a confirmed hcg negative pool serum and with hcg negative urine. The results are summarized in the following table: hcg IU/l Results with urine Results with serum Test line Test line ± ± 15 ± ± Intensity grades: 4+ (very strong intensity); 3+ (strong intensity); 2+ (medium intensity); + (visible); ± (very weakly positive); - (no intensity) Conclusion: HEXAGON hcg 1-STEP Serum/Urine, HEXAGON hcg 1-STEP Detector and HumaPreg Serum/Urine shows with serum as well as with urine a sensitivity of at least 25 IU/l hcg. Rev /5

3 3 Validation of Efficacy 3.1 Clinical Evaluation For a clinical evaluation 146 female patients of child-bearing age were selected by four different hospitals. The eligibility criterion for participation was a predicted ovulation date 7 days prior to the collection of a first blood sample. Further samples were taken in intervals of 3-7 days (i.e., 10 or 14 days after ovulation). The diagnosis pregnancy was confirmed or excluded by ultrasonography. Results of the clinical evaluation Hospital No. of patients Results with HEXAGON hcg 1-STEP Serum/Urine Negative Positive at day 7 Positive at day10 Positive at day14 (days after the predicted ovulation date) A not tested 12 B C D Total of 38* 26 of 26** 38 of 38* * confirmed pregnancies, ** confirmed pregnancies only in hospital B, C and D. 3.2 Summary HEXAGON hcg 1-STEP Serum/Urine showed with serum samples from pregnant women an excellent sensitivity. A positive test result was with 33 serum samples already detected at day 7 after the predicted ovulation date. In all 38 confirmed pregnancies a positive result was obtained the latest at day 14 after ovulation. This confirms that HEXAGON hcg 1-STEP Serum/Urine is well suited to detect an early pregnancy. 4 Cross-Reactivity, Interferences 4.1 Description of Control Materials Cross-reactivity has been tested with hcg negative specimens spiked with LH, FSH and TSH preparations to obtain the following concentrations: 1,000 IU/l TSH, 300 IU/l LH, 1,000 IU/l FSH. Potentially interfering substances (such as bilirubin, haemoglobin, drugs, vitamins, etc.) have been added to hcg negative urine specimens and to specimens with a hcg concentration adjusted to 50 IU/l. 4.2 Results Substance Concentration, Result with negative urine Result with positive mg/dl specimens specimens (50 IU/l hcg) Bilirubin 2 negative positive Haemoglobin 24 negative positive Creatinine 200 negative positive Albumin 2,000 negative positive Human serum proteins 2,000 negative positive Urea 3,900 negative positive Uric acid 10 negative positive Acetaminophen 20 negative positive Acetylsalicylic acid 20 negative positive Ascorbic acid 40 negative positive Caffeine 40 negative positive Gentisic acid 40 negative positive Glucose 2,000 negative positive Ampicillin 40 negative positive Tetracycline 40 negative positive Ethanol 5,000 negative positive Amobarbital 10 µg/dl negative positive Codeine 6 µg/dl negative positive Methamphetamine 10 µg/dl negative positive Morphine 6 µg/dl negative positive Rev /5

4 Summary: No cross-reactivity has been observed with the tested concentrations of LH, FSH and TSH. None of the above listed, potentially interfering substances showed any influence on the expected result. 5 Reproducibility 5.1 Within-Run Performance / Lot-to-Lot Two different specimens were prepared for reproducibility testing: hcg negative serum and serum containing hcg adjusted to a concentration of 25 IU/l. These specimens were tested with HEXAGON hcg 1-STEP Serum/Urine test strips according to the instruction for use. Results with HEXAGON hcg 1-STEP Serum/Urine hcg Negative Serum Positive Serum 25 IU/l hcg Lot X2001, expiry date: Lot X3002, expiry Lot X1201, expiry Conclusion: The study has shown for visually interpreted HEXAGON hcg 1-STEP Serum/Urine tests a 100% reproducibility of negative and weakly positive results (within-run performance and lot-to-lot testing). 6 Stability 6.1 Real Time Stability Samples from three different lots were stored at room temperature. Testing at each time interval consisted of testing negative serum and a positive serum specimen with a hcg concentration adjusted to the cut-off of 25 IU/l. The results are summarised in the following table. Results: Lot X2001 Expiry date: Lot X3002 Expiry date: Lot X1201 Expiry date: Month 0 IU/l 25 IU/l 0 IU/l 25 IU/l 0 IU/l 25 IU/l 0 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 1 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 2 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 3 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 4 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 5 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 6 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 7 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 8 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 9 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 10 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 11 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 12 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 13 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 14 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] Rev /5

5 15 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 16 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 17 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 18 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 19 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 20 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 21 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 22 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 23 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 24 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 25 neg. [-] pos. [+] neg. [-] pos. [+] neg. [-] pos. [+] 26 neg. [-] pos. [±] neg. [-] pos. [±] neg. [-] pos. [±] + = clearly visible test line; ± = weak positive test line; - = no test line visible The three tested lots revealed an excellent stability when stored at room temperature. The obtained results support the claimed shelf life of 24 months from the date of manufacturing. Rev /5