Immunological detection of pregnancy: Evidence for systemic immune modulation during early pregnancy in ruminants.

Mammalian pregnancy creates unique challenges for immune systems highly evolved to detect and eliminate invading pathogens. Recognition of the challenges created by gestating a semi-allogeneic fetus evolved from the discipline of transplantation biology and were informed by studies on the unique natural parabiosis that occurs when female calves are gestated with twin male fetuses. These pregnancies typically result in an intersex female termed a freemartin, which revealed insights into development of the male and female reproductive tracts. However, they also uncovered important clues on immune tolerance with wide-ranging implications to reproductive biology, transplantation biology and autoimmune disease. Many studies focused on identifying mechanisms through which the fetus evades maternal immune detection and elimination. These included studies characterizing immune interactions between the fetus and mother at the nourishing interface of the placenta and uterine endometrium. This immunological forbearance only occurs under high concentrations of circulating progesterone. Beyond the requirement for progesterone, there has been considerable progress towards understanding the effects of conceptus signals on maternal immune function. One common theme is that pregnancy induces a T helper 2 immune bias as shown in several mammalian species, including domestic ruminants. However, a growing body of evidence shows that the fetus not only evades, but also provokes immune responses locally in the uterus and in peripheral tissues. This is perhaps most dramatically illustrated by domestic ruminants where the conceptus secretes a unique interferon in the opening salvo of hormonal communication with the maternal immune system. The role of interferon tau in regulating expression of genes of the innate immune system in the uterus has been extensively studied. More recently, it was determined that these same genes are also induced in peripheral immune cells and other tissues throughout the body. In addition to interferon tau and progesterone, pregnancy associate glycoproteins and chaperonin 10 (aka Early Pregnancy Factor) are implicated in altering immune function both locally and systemically during pregnancy. While it is tempting to speculate that this activation of innate immunity is designed to counteract selective immunosuppression, knowledge of the importance of local and systemic immune activation to the success of pregnancy remains incomplete. This area remains fertile ground for developing better approaches to diagnose and treat infertility in domestic farm species and humans alike.

Comparison of Result Times Between Urine and Whole Blood Point-of-care Pregnancy Testing.

INTRODUCTION: Point-of-care (POC) pregnancy testing is commonly performed in the emergency department (ED). One prior study demonstrated equivalent accuracy between urine and whole blood for one common brand of POC pregnancy testing. Our study sought to determine the difference in result times when comparing whole blood versus urine for the same brand of POC pregnancy testing. METHODS: We conducted a prospective, observational study at an urban, academic, tertiary care hospital comparing the turnaround time between order and result for urine and whole blood pregnancy tests collected according to standard protocol without intervention from the investigators. After the blood was collected, the nurse would place three drops onto a Beckman Coulter ICON 25 Rapid HCG bedside pregnancy test and set a timer for 10 minutes. At the end of the 10 minutes, the result and time were recorded on an encoded data sheet and not used clinically. The same make and model analyzer was also used for urine tests in the lab located within the ED. The primary outcome was the difference in mean turnaround time between whole blood in the ED and urine testing in the adjacent lab results. Concordance between samples was assessed as a secondary outcome. RESULTS: 265 total patients were included in the study. The use of whole blood resulted in a mean time savings of 21 minutes (95% CI 16-25 minutes) when compared with urine (p<0.001). There was 99.6% concordance between results, with one false negative urine specimen with a quantitative HCG level of 81 mIU/L. CONCLUSION: Our results suggest that the use of whole blood in place of urine for bedside pregnancy testing may reduce the total result turnaround time without significant changes in accuracy in this single-center study.

Quantitative analysis of total ß-subunit of human chorionic gonadotropin concentration in urine by immunomagnetic reduction to assist in the diagnosis of ectopic pregnancy.

BACKGROUND: The initial diagnosis of ectopic pregnancy depends on physical examination, ultrasound, and serial measurements of total ß-subunit of human chorionic gonadotropin (hCGß) concentrations in serum. The aim of this study was to explore the possibility of using quantitative analysis of total hCGß in urine rather than in serum by immunomagnetic reduction (IMR) assay as an alternative method to diagnose an ectopic pregnancy. METHODS: We established a standard calibration curve of IMR intensity against total hCGß concentration based on standard hCGß samples, and used an IMR assay to detect total hCGß concentrations in the urine of pregnant women with lower abdominal pain and/or vaginal bleeding. The final diagnosis of ectopic pregnancy was based on ultrasound scans, operative findings, and pathology reports. In this prospective study, ten clinical samples were used to analyze the relationship of total hCGß IMR signals between urine and serum. Furthermore, 20 clinical samples were used to analyze the relationship between urine IMR signals and serum levels of total hCGß. RESULTS: The calibration curve extended from 0.01 ng/mL to 10,000 ng/mL with an excellent correlation (R(2)=0.999). In addition, an excellent correlation of total hCGß IMR signals between urine and serum was noted (R(2)=0.994). Furthermore, a high correlation between urine IMR signals and serum levels of total hCGß was noted (R(2)=0.862). CONCLUSION: An IMR assay can quantitatively analyze total hCGß concentrations in urine, and is a potential candidate for point-of-care testing to assist in the diagnosis of ectopic pregnancy.

Purification of pregnancy-associated glycoproteins from late-pregnancy Bubalus bubalis placentas and development of a radioimmunoassay for pregnancy diagnosis in water buffalo females.

BACKGROUND: Pregnancy-associated glycoproteins (PAGs) were first described as placental antigens present in the blood serum of the mother soon after implantation. Here, we describe the purification of several pregnancy-associated glycoproteins from water buffalo placenta (wbPAGs). A specific radioimmunoassay (RIA) was developed for early pregnancy diagnosis in buffalo species. RESULTS: Amino-terminal microsequencing of immunoreactive placental proteins allowed the identification of eleven wbPAGs sequences [Swiss-Prot accession numbers: P86369 to P86379]. Three polyclonal antisera (AS#858, AS#859 and AS#860) were raised in rabbits against distinct wbPAG fractions. A new RIA (RIA-860) was developed and used to distinguish between pregnant (n=33) and non-pregnant (n=26) water buffalo females. CONCLUSIONS: Our results confirmed the multiplicity of PAG expression in buffalo placenta. In addition, the RIA-860 system was shown to be sensitive, linear, reproducible, accurate and specific in measuring PAG concentrations in buffalo plasma samples from Day 37 of gestation onwards.

Substituting whole blood for urine in a bedside pregnancy test.

BACKGROUND: Point-of-care testing for rapid detection of pregnancy in women of reproductive age is common practice in the emergency department. Commercially available rapid human chorionic gonadotropin (hCG) immunoassays are validated for use with urine and serum, but not whole blood. STUDY OBJECTIVES: We assessed the validity of using whole blood to detect pregnancy using a point-of-care hCG assay by comparing it to a laboratory quantitative serum hCG assay as the criterion standard. METHODS: A convenience sample of female patients of reproductive age (18-51 years) submitted 5mL of whole blood, from which two drops were immediately applied to a point-of-care hCG kit, with results recorded at 10min. The remainder of each whole blood specimen was sent to the hospital laboratory for the criterion-standard quantitative serum hCG assay. The criterion standard for a positive pregnancy test was defined as quantitative serum hCG≥5 mIU/mL. Investigators performing the whole blood test and laboratory technicians performing the quantitative serum assay were blinded to one another's results. RESULTS: There were 633 patients enrolled, with a mean age of 30 years (± 7.7 years); 34% of the patients were pregnant. Overall, the whole blood pregnancy test was 95.8% sensitive (negative predictive value 97.9%), whereas the urine test was 95.3% sensitive (negative predictive value 97.6%); the specificity and positive predictive value of both tests was 100%. CONCLUSION: Using a standard point-of-care qualitative hCG immunoassay kit, whole blood may be used for rapid detection of pregnancy with similar, or greater, accuracy than urine.

Incorporation of a rapid pregnancy-associated glycoprotein ELISA into a CIDR-Ovsynch resynchronization program for a 28 day re-insemination interval.

The objective was to compare two resynchronization programs; one that used a blood-based ELISA for pregnancy-associated glycoproteins (PAG) for pregnancy diagnosis so that non-pregnant cows were re-inseminated at 28 d after first TAI, and another that used transrectal ultrasonography for pregnancy diagnosis so that non-pregnant cows were re-inseminated at 35 d after first TAI. The PAG_resynch cows (n = 103) began CIDR-Ovsynch resynchronization on Day 18 after first TAI (Day 0). On Day 25, the CIDR was removed and pregnancy diagnosis with a PAG ELISA was performed. If a cow was not pregnant on Day 25, she was treated with PGF(2α), treated with GnRH 2 d later (Day 27), and TAI on Day 28. Control cows (n = 99) were observed for estrus until Day 25, when they began an identical CIDR-Ovsynch program with pregnancy diagnosis by transrectal ultrasonography on Day 32. If a cow was not pregnant on Day 32, then she was treated with PGF(2α), treated with GnRH 2 d later (Day 34), and TAI on Day 35. There was no difference in pregnancy per AI (P/AI) for either group at first or second insemination. For cows without pregnancy loss, the interval between first and second (P < 0.001) or second and third (P < 0.016) TAI was shorter for PAG_resynch cows compared with Control cows. The interval between first and second or second and third TAI was not different if pregnancy loss cows were included in the analysis. Plasma progesterone concentrations were similar at PGF(2α) treatment, and plasma estradiol concentrations increased similarly after PGF(2α) treatment for PAG_resynch and Control cows. In conclusion, the 28 d CIDR-Ovsynch resynchronization protocol was comparable to a 35 d CIDR-Ovsynch resynchronization protocol that also included estrus detection. Shortened resynchronization protocols that do not require estrus detection may improve reproductive efficiency in dairy cattle.

Field experiences with early pregnancy diagnosis by progesterone-based ELISA in sows.

In four Kenyan pig breeding units the pregnancy diagnosis of sows has been carried out in two groups: Group 1 (n = 1911): the sows were transrectaly pregnancy tested between Days 17-22 post-mating by ultrasound. Sows testing non-pregnant immediately received one dose of 400 IU pregnant mare serum gonadotropin (PMSG) (equine chorion gonadotropin, eCG) and 200 IU human chorion gonadotropin (hCG). On showing signs of oestrous, the animals were subsequently artificially inseminated (AI). Group 2 (n = 1923): sows were pregnancy tested by serum progesterone (P4)-based enzyme-linked immunosorbent assay (ELISA) on Day 17 post-breeding. P4 concentrations were categorized as positive (> 5 ng/ml) or negative (< 5 ng/ml). Sows testing nonpregnant immediately received one dose of 400 IU PMSG and 200 IU hCG by injection, and were subsequently artificially inseminated. The following parameters were evaluated: sows diagnosed non-pregnant, days from first post-weaning insemination until the sows were inseminated at their first return to oestrus; farrowing rate and total piglets born and number of live-born piglets in litters. The percentage of sows diagnosed non-pregnant in the two groups, as well as the totals of born piglets and of live-born piglets in litters did not differ significantly between the two groups. The number of days from the first post-weaning mating until the sows were artificially inseminated at their first return to oestrus and the administration of eCG and hCG was shorter (P < 0.01) and farrowing rate was higher (P< 0.01) in the ELISA-tested sows.

Concentration of bovine pregnancy associated glycoprotein in plasma and milk: its application for pregnancy diagnosis in cows.

Pregnancy diagnosis is an important part in reproduction management of ruminants. The aim of the study was to use a new method for evaluating the bPAG and cPAG in milk and blood bPAG and compare this results with the other method for pregnancy diagnosis in the cows. The study was carried out in 220 Holstein Frisian cows. Heparinised blood samples were taken from the jugular vein and stored at -20 degrees C until PAG assay by RIA. For bPAG and cPAG, RIA test, milk samples were homogenized. Pure bPAG was used as a standard tracer described by Zoli et al. (1992). The cows were diagnosed as pregnant by means of USG (Aloka SSD 210) and by rectal palpation. bPAG and cPAG concentration in milk increased after 28 day of pregnancy and showed the rapid increase near the parturition. The same results of bPAG concentration we obtained in the blood samples. The decline of bPAG concentration was faster in the milk than in the blood. The data showed that the RIA method is precise enough to measure PAG concentrations in the maternal blood and milk of cows. The data indicate that milk samples can be used for pregnancy diagnosis in cows. The sensitivity and specificity of RIA measurement of PAG are very high.

Evaluation of early conception factor lateral flow test to determine nonpregnancy in dairy cattle.

The early conception factor (ECF) lateral flow test was evaluated for its ability to accurately determine nonpregnant status in dairy cattle. Results of 2 field trials involving 191 cows and 832 tests indicated the probability that a cow can be correctly diagnosed as nonpregnant by using the ECF test is only about 50%. Agreement of test results between milk and serum obtained from the same cow was 57.5%. The ECF test was not consistent in identifying nonpregnancy when the same cows were tested repeatedly over a period of 4 weeks. We conclude that the ECF lateral flow test does not accurately identify nonpregnancy in dairy cattle.

Quantification of urinary chorionic gonadotropin in spontaneous abortion of pre-clinically recognized pregnancy: method development and analytical validation.

Determination of environmental impacts on reproductive health and specifically on the incidence of early spontaneous abortion requires accurate estimates of the latter. This negative reproductive outcome can be detected by the pattern of elevation and decline of human chorionic gonadotropin (hCG) levels near and shortly beyond the expected time of implantation, requiring daily biomonitoring of hCG levels during the relevant period of the menstrual cycle. Prospective pregnancy studies to assess effects of potentially toxic exposures on human reproductive outcomes can involve up to three menstrual cycles and a huge number of samples in each, for the quantification of the inherently very low hCG levels usually can be determined only in serum. The invasive nature of blood collection, the number of samples needed for the development of prospective studies, and the lack of quantitative methods for the determination of low hCG levels in urine point to the need for collecting urine rather than blood and make it imperative to develop suitable quantitative methods for biomonitoring of very low levels of hCG in urine. This paper describes the development and validation procedures of an automated solid-phase two-site chemiluminescent immunometric assay for the quantification of urinary hCG in early pregnancy and early pregnancy loss. For the validation, both undiluted and diluted urine and control samples have been prepared. From the results, it can be concluded that the assay has a calibration range that extends to 5000 mIU/ml, with a detection limit of approximately 1.2 mIU/ml, practically identical to that found by the IMMULITE 2000 manufacturer's validation study. The intra- and inter-assay precision ranges up to a maximum of around 7%, meaning that the practical limit for functional sensitivity can be established as low as 10%. This means that the immunoassay from DPC can identify, with relatively high confidence, non-pregnant women and the typical "rise and fall" pattern of early pregnancy loss through analysis of urine samples. Results also lead to the conclusion that there is a very good agreement between expected and observed urinary hCG levels indicative of good immunoassay accuracy for the studied range of hCG concentrations. In terms of analyte stability, it can be concluded that urinary hCG is stable under the expected conditions required for ongoing investigations that include temperatures of 2-8 degrees C for up to 48 h and temperatures of around -20 degrees C for longer periods that can extend to over 3 months.

Uso adecuado del ensayo de gonadotrofina coriónica humana en el diagnóstico de embarazo. ¿Sangre u orina? / Adequate use of the human chorionic gonadotropin in the pregnancy diagnosis. Blood or urine?

Introducción: Los niveles de Gonadotrofina Coriónica humana (hCG) pueden detectarse en suero o en orina con métodos de distinta sensibilidad y permiten no solamente el diagnóstico del embarazo, sino también su seguimiento, la posibilidad de detección de embarazos ectópicos y las amenazas de aborto. Objetivos: 1) Demostrar la concordancia entre las pruebas para diagnóstico precoz de embarazo en muestras simultáneas de sangre y orina a niveles iguales o superiores a 25 mUI/ml. 2) Estimar el grado de sobreutilización del enzimoinmunoensayo en sangre y su repercusión sobre los costos. Material y métodos: Sesenta muestras consecutivasde suero y sus correspondientes de orina recolectadas el mismo día de la extracción de sangre. La determinación cuantitativa de hCG b total en sangre se efectuó por el método ELISA (MEIA) (Abbott. AXSYM System). En orina se utilizó un inmunoensayo cromatográfico cualitativo en un solo paso (Acon).

Uso adecuado del ensayo de gonadotrofina coriónica humana en el diagnóstico de embarazo. ¿Sangre u orina? / Adequate use of the human chorionic gonadotropin in the pregnancy diagnosis. Blood or urine?

Introducción: Los niveles de Gonadotrofina Coriónica humana (hCG) pueden detectarse en suero o en orina con métodos de distinta sensibilidad y permiten no solamente el diagnóstico del embarazo, sino también su seguimiento, la posibilidad de detección de embarazos ectópicos y las amenazas de aborto. Objetivos: 1) Demostrar la concordancia entre las pruebas para diagnóstico precoz de embarazo en muestras simultáneas de sangre y orina a niveles iguales o superiores a 25 mUI/ml. 2) Estimar el grado de sobreutilización del enzimoinmunoensayo en sangre y su repercusión sobre los costos. Material y métodos: Sesenta muestras consecutivasde suero y sus correspondientes de orina recolectadas el mismo día de la extracción de sangre. La determinación cuantitativa de hCG b total en sangre se efectuó por el método ELISA (MEIA) (Abbott. AXSYM System). En orina se utilizó un inmunoensayo cromatográfico cualitativo en un solo paso (Acon).(AU)

A single serum test for measuring early pregnancy outcome with high predictive value.

OBJECTIVES: Current testing to determine a failing pregnancy requires two separate clinic visits to measure the hCG doubling rate. Diagnosing a failing pregnancy is often done in emergency departments where simplified and accelerated testing methods are needed. Here, we investigated hyperglycosylated hCG (hCG-H) for predicting pregnancy failure. DESIGN AND METHODS: We studied two independent sets of patient samples collected in the early weeks of gestation. One set was urine samples, and the other was serum samples. In all cases, hCG and hCG-H were measured using automated chemiluminescence immunoassays. Concentrations of hCG and hCG-H were plotted on a scattergram, and levels in failing pregnancies were compared to those in continuing pregnancies. RESULTS: Data indicated that a threshold level of hCG-H (13 microg/L) in both serum and urine samples defined the concentration below where pregnancies were likely to fail. This cut-off corresponded to 73% detection of failures at a 2.9% false positive rate using serum and 75% detection at a 15% false positive rate using urine. Using an hCG cut-off that corresponded to the same false positive rates, hCG detected only 42% of failures using serum and 43% of failures using urine. CONCLUSIONS: Our data indicate that hCG-H provides a much more accurate single test than hCG for assessing pregnancy outcome. Compatible with the use of serum or urine samples, a single hCG-H test might provide simpler, faster, and more accurate results for predicting the progress of a pregnancy than standard hCG testing.

Comparación de dos métodos para la evaluación de la gonadotrofina coriónica en suero / Comparison of two methods for the evaluation of the chorionic gonadotrophin in serum

El uso de los anticuerpos (Ac) monoclonales específicos dirigidos contra la sub-unidad beta de la gonadotrofina coriónica (hCG-beta) es un prerrequisito para los tests de embarazo con un alto nivel de sensibilidad. Sin embargo, el diseño de inmunoensayos cada vez más sensibles y específicos no se ha visto reflejado en la comparabilidad de los resultados de los distintos métodos. Objetivos: a) Principal: evaluar la concordancia entre dos inmunoensayos (EIA VIDAS Y AxSYM) para la determinación cuantitativa de la hormona gonadotrofina coriónica humana (hCG) sub-unidad beta en suero. b) Secundario: evaluar la correlación e intercambiabilidad de ambos ensayos y c) plantear las dificultades inherentes a su utilización a través de nuestra experiencia. Material y métodos: se recolectaron 106 muestras consecutivas de suero de pacientes que concurrieron al Consultorio de Procreación Responsable y embarazadas de edades gestacionales entre 3 y 12 semanas extraídas para la determinación de hCG o hCG-beta. Las muestras fueron analizadas por dos metodologías de EIA: 1. AxSYM betahCG total (Lab. Abbott). 2. Vidas hCG (Lab. Biomerieux). Conclusiones: a pesar de la complejidad de la bioquímica y la fisiopatología de la hCG, los recientes avances han mejorado la estandarización del ensayo de esta hormona. Para el caso específico del diagnóstico precoz del embarazo, (donde los resultados obtenidos son cercanos a cero o bajos) la concordancia entre los métodos permite el uso indistinto de uno u otro. Para valores por encima de 20.000 mU/ml, los resultados obtenidos están demasiado separados entre sí y, por consiguiente, los métodos no serían intercambiables, lo que implica la necesidad de realizar las mediciones seriadas de una misma paciente con un mismo equipo de diagnóstico.

Comparación de dos métodos para la evaluación de la gonadotrofina coriónica en suero / Comparison of two methods for the evaluation of the chorionic gonadotrophin in serum

El uso de los anticuerpos (Ac) monoclonales específicos dirigidos contra la sub-unidad beta de la gonadotrofina coriónica (hCG-beta) es un prerrequisito para los tests de embarazo con un alto nivel de sensibilidad. Sin embargo, el diseño de inmunoensayos cada vez más sensibles y específicos no se ha visto reflejado en la comparabilidad de los resultados de los distintos métodos. Objetivos: a) Principal: evaluar la concordancia entre dos inmunoensayos (EIA VIDAS Y AxSYM) para la determinación cuantitativa de la hormona gonadotrofina coriónica humana (hCG) sub-unidad beta en suero. b) Secundario: evaluar la correlación e intercambiabilidad de ambos ensayos y c) plantear las dificultades inherentes a su utilización a través de nuestra experiencia. Material y métodos: se recolectaron 106 muestras consecutivas de suero de pacientes que concurrieron al Consultorio de Procreación Responsable y embarazadas de edades gestacionales entre 3 y 12 semanas extraídas para la determinación de hCG o hCG-beta. Las muestras fueron analizadas por dos metodologías de EIA: 1. AxSYM betahCG total (Lab. Abbott). 2. Vidas hCG (Lab. Biomerieux). Conclusiones: a pesar de la complejidad de la bioquímica y la fisiopatología de la hCG, los recientes avances han mejorado la estandarización del ensayo de esta hormona. Para el caso específico del diagnóstico precoz del embarazo, (donde los resultados obtenidos son cercanos a cero o bajos) la concordancia entre los métodos permite el uso indistinto de uno u otro. Para valores por encima de 20.000 mU/ml, los resultados obtenidos están demasiado separados entre sí y, por consiguiente, los métodos no serían intercambiables, lo que implica la necesidad de realizar las mediciones seriadas de una misma paciente con un mismo equipo de diagnóstico. (AU)

Frequent false positive beta human chorionic gonadotropin tests in immunoglobulin A deficiency.

A patient with IgA deficiency had a series of positive serum pregnancy tests which led to medical and surgical procedures for suspected molar pregnancy. These tests were found to be falsely positive due to heterophile antibody. The aim of this study was to determine the frequency of false positive betahCG assays in sera of IgA deficient patients. Sera from a panel of IgA deficient (IgA < 7 mg/dl) patients were tested for the presence of betaHCG using three different assays, and also for IgG anti-goat and anti-mouse antibodies. Patients were seen at Mount Sinai Medical Center and included 54 patients (ages 1-80 years, 32 females, 22 males) with IgA deficiency. Thirty percent of 54 IgA deficient patient sera yielded positive pregnancy tests by one or more of the three betahCG assays, however, none of the patients were pregnant. In comparison to sera of normal controls, 39% of the patient sera contained significant amounts of anti-goat antibody and 18% contained significant amounts of anti-mouse antibody. While heterophile antibodies are common in IgA deficient serum, false positive assays for betahCG in IgA deficient serum have not been previously reported. The possibility of false positive test results should be considered prior to invasive procedures in IgA deficient patients.

Quantitative automated human chorionic gonadotropin measurement in urine using the Modular Analytics E170 module (Roche).

Ongoing demands on laboratory performance require optimization of processes. An obvious way to achieve this is to reduce manual labor in favor of automated methods. We describe the validation of an automated quantitative urine human chorionic gonadotropin (hCG) analysis on the Roche Modular E170 analyzer to replace the manual qualitative pregnancy test in urine. At urine hCG concentrations of 476, 45 and 11 U/L, we found inter-assay variation of 4.3%, 4.3% and 6.8% and average intra-assay variation of 3.0%, 2.6% and 3.0%, respectively. The analytical detection limit was 0.7 U/L. We did not detect any loss (due to degradation or adsorption) during a storage period of 5 days at 4 degrees C or at -20 degrees C. Recoveries of hCG in urine of a pregnant woman diluted with urine of a pre-menopausal non-pregnant woman (concentration range between 6 and 800 mU/L) were between 93% and 112% (y=0.997x-3.843, r 2 =0.999). Diluting a serum sample (hCG 42,000 U/L) with urine (negative for hCG) up to 8000-fold yielded a completely linear hCG response, indicating that the assay was not affected by the urine matrix. In a correlation study with 60 urine samples (of which 10 were of male origin), we did not find any discrepancies between results for the manual pregnancy test and the hCG test on the Roche Modular E170 (using a cutoff value of 50 U/L).