Differential diagnosis of lymphocytosis in routine laboratory practice: Contribution of lymphocyte parameters using the Sysmex-XN9000 haematology analyzer.

INTRODUCTION: This study investigates the potential contribution of structural and dispersion parameters, as well as alarms provided by Sysmex XN9000 haematology analyzer. The objective was to assess the need for a microscopic examination in the context of lymphocytosis. It also aims to contribute in differentiating rapidly lymphoproliferative disorders such as chronic lymphocytic leukaemia (CLL), non-chronic lymphocytic leukaemia (NON-CLL) and non-infectious reactive lymphocytosis (REAC). METHODS: We prospectively assessed lymphocyte parameters (Ly-X, Ly-Y, Ly-Z, Ly-WX, Ly-WY, Ly-WZ) provided by the Sysmex XN9000 analyzer; they were measured in the white blood cell differential (WDF) channel which also provides alarms via the precursor/pathological cellular channel (WPC). Blood samples from 71 subjects with CLL, NON-CLL lymphoproliferative and REAC non-infectious reactive lymphocytosis, as well as a control (NORM) group of 12 subjects without any abnormalities were analyzed. RESULTS: The most discriminating parameters to distinguish the different groups were Ly-X, Ly-Z and Ly-WZ. The lymphoid structural parameters Ly-X and Ly-Z significantly discriminated the CLL group from the other groups (p < 0.001), and the CLL group from the REAC group (p < 0.01), respectively. The Ly-WZ parameter discriminated the CLL group from the NON-CLL, REAC (p < 0.001) and NORM (p < 0.01) groups. Alarms were higher in all study groups compared to the NORM group. An algorithm integrating these structural and alarm parameters is proposed. CONCLUSION: This study demonstrated that Ly-X, Ly-Z, and Ly-WZ lymphocyte parameters are useful for detecting morphological changes in lymphocytes; they provide useful information for the differential diagnosis of lymphocytosis, and this before the examination of the blood smear. An algorithm combining the WDF (parameters) and the WPC (alarms) makes it possible to decide whether or not to use a microscopic examination or flow cytometry immunophenotyping.

Evaluation of maternal serum biomarkers in predicting outcome of successful expectant management of tubal ectopic pregnancies.

OBJECTIVE: To assess the value of multiple serum biomarkers for the prediction of successful outcome of expectant management in women with tubal ectopic pregnancy (TEP). STUDY DESIGN: Women with a conclusive ultrasound diagnosis of TEP had a blood test to measure ß-human chorionic gonadotropin (ß-hCG), progesterone, inhibin A, activin A and high sensitivity C-reactive protein (hsCRP) at the initial visit. Women presenting with pain, serum ß-hCG ≥ 1500 IU, evidence of a live ectopic pregnancy or a significant haemoperitoneum were advised to have emergency surgery. Women eligible for expectant management were followed-up prospectively until serum ß-hCG declined to non-pregnant level or surgical treatment was required. RESULTS: A total of 93 women with a TEP were included in the final cohort. Emergency surgery was carried out in 42/93 (45 %) of women whilst 51/93 (55 %) were managed expectantly. Of the latter group, 42/51 (82 %) had successful expectant management and 9/51(18 %) required surgical procedure after a period of follow up. On multi-variable analysis, only higher values of serum ß-hCG and progesterone at the initial visit were associated with a lower chance of successful expectant management of TEP. A one-unit increase in either variable on the log-scale was associated with an approximate 20-fold reduction in the odds of a successful outcome. CONCLUSION(S): Serum ß-hCG and progesterone were significantly lower in women who had successful expectant management of TEP. Other biomarkers under consideration were not significantly different in women with successful and failed expectant management.

Baseline hepcidin measurement in the differential diagnosis of anaemia for elderly patients and its correlation with the increment of transferrin saturation following an oral iron absorption test.

Background Anaemia is often multifactorial in the elderly, with a frequent association between iron deficiency anaemia (IDA) and anaemia of chronic disease (ACD). The primary objective of our study was to investigate whether baseline hepcidin measurement could be useful for identifying iron deficiency (ID) in anaemic elderly patients. The secondary objective was to assess whether baseline hepcidin concentrations correlated with the relative increase of transferrin saturation (TS) after an oral iron absorption test (OIAT). Methods Blood samples were collected between 7:30 am and 10:00 am in 328 geriatric outpatients, 102 underwent the OIAT. Types of anaemia were classified according biochemical and clinical criteria. TS and hepcidin were measured at baseline and 4 h after the iron dose. The ability of baseline hepcidin measurement to highlight ID in elderly anaemic patients was assessed using a receiver operator curve (ROC) analysis. Correlations between baseline hepcidin levels and the increment of TS following the OIAT were investigated using the Spearman coefficient. Results Among 328 included patients, 78 (23.8%) suffered from anaemia; 13 (4.0%), 19 (5.8%), 27 (8.2%) and 19 (5.8%) patients fulfilled criteria for IDA, IDA/ACD, ACD and unexplained anaemia, respectively. By multivariable analysis, creatinine, C-reactive protein, ferritin, Delta TS and Delta hepcidin were independently associated with baseline hepcidin concentrations. The area under the ROC curve (95% confidence interval) was 0.900 (0.830-0.970) for baseline hepcidin measurement. Baseline hepcidin levels correlated negatively with the relative increase in TS with a Spearman coefficient of -0.742. Conclusions Baseline hepcidin levels could be a useful tool to identify ID in anaemic elderly patients and may predict acute iron response following OIAT.

Neonatal Screening for Sickle Cell Disease in Belgium for More than 20 Years: An Experience for Comprehensive Care Improvement.

Our previous results reported that compared to sickle cell patients who were not screened at birth, those who benefited from it had a lower incidence of a first bacteremia and a reduced number and days of hospitalizations. In this context, this article reviews the Belgian experience on neonatal screening for sickle cell disease (SCD). It gives an update on the two regional neonatal screening programs for SCD in Belgium and their impact on initiatives to improve clinical care for sickle cell patients. Neonatal screening in Brussels and Liège Regions began in 1994 and 2002, respectively. Compiled results for the 2009 to 2017 period demonstrated a birth prevalence of sickle cell disorder above 1:2000. In parallel, to improve clinical care, (1) a committee of health care providers dedicated to non-malignant hematological diseases has been created within the Belgian Haematology Society; (2) a clinical registry was implemented in 2008 and has been updated in 2018; (3) a plan of action has been proposed to the Belgian national health authority. To date, neonatal screening is not integrated into the respective Belgian regional neonatal screening programs, the ongoing initiatives in Brussels and Liège Regions are not any further funded and better management of the disease through the implementation of specific actions is not yet perceived as a public health priority in Belgium.

Comparison of two assays measuring human chorionic gonadotrophin serum concentrations in pregnancy termination and trophoblastic or non-trophoblastic tumours.

BACKGROUND: Differences in human chorionic gonadotrophin (hCG) results provided by the commercial immunoassays reflect the heterogeneity of antibodies and the use of suboptimal standards. As a consequence, the principal forms of hCG and metabolites are differentially detected and the hCG tests are not suited for the same clinical applications. Conflicting results are available in the literature regarding which hCG variants are recognized by the Roche Elecsys hCG + ß test. The aim of our study was to compare the hCG concentrations provided by the Siemens Immulite 2000 test and the Roche test as well as to assess the concordance between both assays. METHODS: In this purpose, 152 samples obtained from women and 44 samples from men were analysed by both tests during the follow-up of pregnancy termination, gestational trophoblastic disease and malignancies. The intermediate precision of the Roche test was also investigated on a pool with a low hCG concentration. RESULTS AND CONCLUSIONS: The hCG concentrations measured with the Roche test were slightly lower compared with the Siemens assay; mean biases of -34.2% and -8% were respectively obtained for hCG values ≤100 UI/L and higher than 100 UI/L. The overall agreement between both assays was 96.1% for women and 97.7% for men. By using an upper reference limit of 3.2 UI/L for women and 1.6 UI/L for men, the Roche test demonstrated a respective concordance of 98.7% and 100%. This test also yielded an excellent precision with a coefficient of variation of 2.8% at a mean hCG concentration of 7 UI/L.

Maternal serum markers in predicting successful outcome in expectant management of missed miscarriage.

The aim of this study was to evaluate the use of biological serum markers, available routinely in most hospital clinical laboratories, in predicting successful outcomes of expectant management in women presenting with a missed miscarriage. This is a single centre observational prospective study over a 16-month period. Among the 490 women who consented to the study protocol, 83 presented with missed miscarriage during the first trimester of pregnancy and opted for expectant management. The mean gestation sac diameter and volume of the gestation sac were recorded during ultrasound examination. Maternal serum samples were obtained in each case and assayed for human chorionic gonadotrophin, progesterone, pregnancy associated plasma protein A (PAPP-A) and high-sensitivity C-reactive protein using commercial assays. When examined individually, maternal age (P = 0.01), progesterone (P = 0.03) and PAPP-A (P = 0.02) were all significantly associated with successful expectant management. Increased maternal age was associated with an increased chance of success with the odds of success increased by around 75% for a 5-year increase in age. Higher values of progesterone and PAPP-A were associated with a reduced chance of successful management. Low maternal serum progesterone concentration was the strongest parameter associated with a successful spontaneous completion of miscarriage.

Survival among children and adults with sickle cell disease in Belgium: Benefit from hydroxyurea treatment.

OBJECTIVE: To evaluate the survival of patients with sickle cell disease (SCD) recorded in the Belgian SCD Registry and to assess the impact of disease-modifying treatments (DMT). METHOD: The Registry created in 2008 included patients of eight centers. All available data in 2008 were retrospectively encoded in the database. After 2008 and until 2012, all data were recorded prospectively for already registered patients as well as newly diagnosed subjects. Data were registered from neonatal screening or from diagnosis (first contact) until last follow-up or death. Data included diagnosis, demography, and outcome data. RESULTS: We collected data from 469 patients over a 5,110 patient years (PY) follow-up period. The global mortality rate was low (0.25/100 PY), although 13 patients died (2.8%) and was similar between children, adolescents (10-18 years), and young adults (P = 0.76). Out of the cohort, 185 patients received hydroxyurea at last follow-up (median duration of treatment: 10.3 years), 90 underwent hematopoietic stem cell transplantation (HSCT), 24 were chronically transfused, and 170 had never had any DMT. Hydroxyurea showed significant benefit on patients outcome as reflected by a lower mortality rate compared to transplanted individuals or people without DMT (0.14, 0.36, and 0.38 per 100 PY, respectively) and by higher Kaplan-Meier estimates of 15 year survival (99.4%) compared to HSCT (93.8%; P = 0.01) or no DMT groups (95.4%; P = 0.04). CONCLUSION: SCD mortality in Belgium is low with no increase observed in young adults. Patients treated with hydroxyurea demonstrate a significant benefit in survival when compared to those without DMT or transplanted.

Evaluation of the role of maternal serum high-sensitivity C-reactive protein in predicting early pregnancy failure.

Maternal serum high-sensitivity C-reactive protein (HSCRP) was evaluated in predicting spontaneous abortion in spontaneous pregnancies presenting with threatened spontaneous abortion. Seventy-one cases of threatened spontaneous abortion (group A) and 71 asymptomatic controls (group B), matched for gestational and maternal age, body mass index and smoking status, were included. Maternal serum samples were evaluated for HCG, progesterone, pregnancy-associated plasma protein-A (PAPP-A) and HSCRP using standard bio-assays. No difference was observed in ultrasound measurements, and median progesterone maternal serum level was significantly lower (P < 0.05) in group A compared with group B. In group A, the median of all ultrasound and maternal serum parameters was significantly lower (P < 0.01) compared with group B. The median gestational sac diameter, volume and median HSCRP and PAPP-A levels were significantly increased (P < 0.05) in group A, with a normal outcome compared with group B, probably owing to the inflammatory reaction associated with intrauterine bleeding. In group A patients destined to abortion, the gestational sac development and corresponding protein synthesis fell before the fetal heart activity stopped; in spontaneous pregnancies, maternal serum HSCRP did not provide additional information for the management of threatened spontaneous abortion but warrants further research in assisted reproduction pregnancies.

Automated reticulocyte parameters for hereditary spherocytosis screening.

The laboratory diagnosis of hereditary spherocytosis (HS) is based on several screening and confirmatory tests; our algorithm includes clinical features, red blood cell morphology analysis and cryohaemolysis test, and, in case of positive screening, sodium dodecyl sulphate polyacrylamide gel electrophoresis as a diagnostic test. Using the UniCel DxH800 (Beckman Coulter) haematology analyser, we investigated automated reticulocyte parameters as HS screening tool, i.e. mean reticulocyte volume (MRV), immature reticulocyte fraction (IRF) and mean sphered cell volume (MSCV). A total of 410 samples were screened. Gel electrophoresis was applied to 159 samples that were positive for the screening tests. A total of 48 patients were diagnosed as HS, and seven were diagnosed as acquired autoimmune haemolytic anaemia (AIHA). Some other 31 anaemic conditions were also studied. From the receiver operating characteristic (ROC) curve analysis, both delta (mean cell volume (MCV)-MSCV) and MRV presented an area under the curve (AUC) of 0.98. At the diagnostic cut-off of 100 % sensitivity, MRV showed the best specificity of 88 % and a positive likelihood ratio of 8.7. The parameters IRF, MRV and MSCV discriminated HS not only from controls and other tested pathologies but also from AIHA contrary to the cryohaemolysis test. In conclusion, automated reticulocyte parameters might be helpful for haemolytic anaemia diagnostic orientation even for general laboratories. In combination with cryohaemolysis, they ensure an effective and time-saving screening for HS for more specialised laboratories.

Marked 25-hydroxyvitamin D deficiency is associated with poor prognosis in patients with alcoholic liver disease.

BACKGROUND & AIMS: Vitamin D deficiency has been frequently reported in advanced liver disease. However, its influence on alcoholic liver disease (ALD) has been poorly elucidated. We investigated the association of vitamin D with clinical, biological, and histological parameters and survival in ALD patients. Furthermore, we explored the effect of vitamin D treatment on ALD patient peripheral blood mononuclear cells (PBMCs), and in a murine experimental model of ALD. METHODS: Serum levels of 25-hydroxyvitamin D [25(OH)D] were determined in 324 Caucasian ALD patients and 201 healthy controls. In vitro experiments on vitamin D pre-treated PBMCs evaluated TNFα production by ELISA in culture supernatants. Mice were submitted to an ethanol-fed diet and some of them were orally supplemented three times per week with 1,25(OH)2D. RESULTS: Severe deficiency in 25(OH)D (<10 ng/ml) was significantly associated with higher aspartate aminotransferase levels (p=1.00 × 10(-3)), increased hepatic venous pressure gradient (p=5.80 × 10(-6)), MELD (p=2.50 × 10(-4)), and Child-Pugh scores (p=8.50 × 10(-7)). Furthermore, in multivariable analysis, a low 25(OH)D concentration was associated with cirrhosis (OR=2.13, 95% CI=1.18-3.84, p=0.013) and mortality (HR=4.33, 95% CI=1.47-12.78, p=7.94 × 10(-3)) at one year. In addition, in vitro, 1,25(OH)2D pretreatment decreased TNFα production by stimulated PBMCs of ALD patients (p=3.00 × 10(-3)), while in vivo, it decreased hepatic TNFα expression in ethanol-fed mice (p=0.04). CONCLUSIONS: Low 25(OH)D levels are associated with increased liver damage and mortality in ALD. Our results suggest that vitamin D might be both a biomarker of severity and a potential therapeutic target in ALD.

Limitations of serum values to estimate glomerular filtration rate during exercise.

OBJECTIVE: Glomerular filtration rate (GFR) is part of routine medical practice for clinical assessment of kidney function in health and disease conditions, and is determined by measuring the clearance of creatinine (Cl-Crn) or estimated (eGFR) from equations using serum creatinine (Crn) or cystatin C (Cyst C). Crn and Cyst C methods obviate the need for urine collection but their reliability under non-resting conditions is uncertain. This study compared GFR determined by Cl-Crn, Crn and Cyst C methods under the conditions of rest and after exercise. METHODS: Twelve young male subjects performed a 30 min treadmill exercise at 80% of the maximal oxygen capacity. Venous blood samples and urine collections were collected before and after exercise and after recovery period. GFR rates were calculated from serum Crn and Cyst C equations, and Cl-Crn measured from serum and urine Crn output. Albumin was also determined for all samples. RESULTS: Under resting conditions, eGFR from Crn and Cyst C did not differ from Cl-Crn (p=0.39). Immediately after exercise, GFR decreased significantly, regardless of the method, but more so for Cl-Crn (-30.0%; p<0.05) compared with Crn (-18.2%) and Cyst C (-19.8%). After the recovery period, GFR determined by Cl-Crn was returned to initial values whereas Crn and Cyst C remained reduced. Although eGFR methods accurately estimate GFR at rest, those methods underestimated the change in GFR after acute exercise. CONCLUSIONS: These results indicate that exercise-induced changes in GFR should be determined by Cl-Crn method.

Reliability of mycophenolic acid monitoring by an enzyme multiplied immunoassay technique.

BACKGROUND: For mycophenolic acid (MPA), substantial inter- and intra-individual variability and drug interactions have been observed and therapeutic drug monitoring is now recommended. In this study, a MPA commercial Enzyme Multiplied Immunoassay Technique (EMIT) was evaluated and compared with the HPLC-UV reference method which is easily practicable in a routine laboratory. METHODS: Plasma samples (n = 117) were collected from adult renal graft patients treated by mycophenolate in combination with either cyclosporin A (CyA) (n = 32) or tacrolimus (n = 85). RESULTS: Considering all samples, correlation was excellent (p < 0.0001). However, significant MPA overestimation was observed with EMIT in the early post-transplant period (30%, n = 32) or when combined with cyclosporin (45%). CONCLUSIONS: In the early post-transplant period, or in cases where CyA is used in combination with MPA, the EMIT cannot be recommended. HPLC or LC/MS are here the method of choice.

Evaluation of an automated capillary electrophoresis system in the screening for hemoglobinopathies.

BACKGROUND: The biochemical diagnosis of hemoglobinopathies (thalassemias and hemoglobin (Hb) variants) is based on the separation and the quantification of Hb fractions. HPLC is the most commonly used method but capillary electrophoresis (CE) methods have also been developed successfully. The Capillarys II system is the first fully automated CE system that allows the quantification of Hb A2 and Hb F and the separation of Hb variants. We evaluated the ability of this system to separate and identify Hb variants and to quantify Hb A2 and Hb F. MATERIAL AND METHODS: The separation of 18 different Hb variants was studied and the imprecision on migration times was calculated for the three most frequent ones. The total imprecision on Hb A2 and Hb F quantification was determined. The results obtained for 44 patients were compared with those given by HPLC. The interference on Hb A2 measurement due to the presence of Hb S was studied. RESULTS: Fourteen out of the 18 variants tested, including all variants of clinical importance, were separated from Hb A. Imprecision on migration times was less than 1%. For Hb A2 quantification, imprecision was less than 3.5% and for Hb F, less than 7.0%. The comparison with HPLC showed an acceptable agreement between both methods but a systematic negative bias for Hb A2 and both proportional and systematic biases for Hb F. No interference from the presence of Hb S on the quantification of Hb A2 was observed. CONCLUSIONS: The fully automated Capillarys Hemoglobin method allows the detection and the separation of the most common Hb variants. It provides also a precise, quick, and very easy quantification of Hb F and Hb A2, even in the presence of Hb S. It is very suitable for routine investigation of hemoglopinopathies.

Plasma hydroxyurea determined by gas chromatography-mass spectrometry.

Hydroxyurea treatment is efficiently used to ameliorate the clinical course of patients affected with sickle cell disease. To understand the patient's wide variation in the clinical response to that drug and monitor its plasma levels, a new method was developed and validated. Fifty microL plasmatic samples containing hydroxyurea are added with internal standard, deproteinized, evaporated to dryness, silanized, and analyzed by gas chromatography-mass spectrometry, which operates in the selected ion mode after electron impact fragmentation. Linearity was found to extend to at least 100mg/L. Over a 1-25mg/L concentration range, coefficients of variation for intra-day and inter-day precision are 5.3% and 7.7%, respectively. Plasma blank-samples reveal endogenous hydroxyurea at a level

Transfer of folic acid inside the first-trimester gestational sac and the effect of maternal smoking.

OBJECTIVE: The objective of the study was to investigate the transfer pathways of folic acid inside the first-trimester gestational sac and to evaluate the impact of maternal smoking. STUDY DESIGN: Folate and cotinine levels were evaluated in maternal serum (n = 125) and coelomic fluid (n = 42), and immunostaining was used to identify folate receptors in placental villi, decidual tissue, and secondary yolk sacs from normal pregnancies at 6-12 weeks' gestation. RESULTS: Folate receptors-alpha were found on the apical surface of syncytiotrophoblast, uterine glandular epithelial cells, and mesothelial layer of the yolk sac. Significantly higher (P < .05) median folate levels were found in maternal serum than in coelomic fluid. The median folate maternal serum and coelomic levels were significantly (P < .0001 and P < .005) lower in smokers, compared with nonsmokers. CONCLUSION: Uterine glands and the secondary yolk sac play key roles in supplying folic acid to the developing fetus before the placental circulations are established, and maternal smoking does not seem to impair directly its transfer mechanism in early pregnancy.

Neonatal haemoglobinopathy screening: review of a 10-year programme in Brussels.

Since 1994, a neonatal screening programme for major haemoglobinopathies has been conducted in Brussels. We performed a 10-year re-evaluation of the incidence of haemoglobinopathies in Brussels and found that of the 118,366 newborns screened, 64 were diagnosed with a sickle cell syndrome, six had beta-thalassaemia major, four had a haemoglobin C disease and three had a haemoglobin H disease. Of the 64 babies with a sickle cell disease, two died before the age of two years and two did not present at the first neonatal visit. Of the six babies suffering from beta-thalassaemia major, all are alive and two have undergone a haematopoietic stem cell transplantation. The universal neonatal screening programme for haemoglobinopathies should be maintained in Brussels.

Neonatal screening for sickle cell disorders in Ouagadougou, Burkina Faso: a pilot study.

OBJECTIVES: To determine the incidence of sickle cell disorders (SCDs) and the feasibility of a neonatal screening programme in Ouagadougou. METHODS: During 2000, 2003 and 2004, 2341 cord blood samples obtained in five maternity hospitals in Ouagadougou were screened for SCDs using an isoelectric focusing technique. The feasibility of a neonatal screening programme was evaluated. RESULTS: The incidence of SCD was 1:57; 14 neonates were homozygous for haemoglobin (Hb)S and 27 were compound heterozygotes for HbSC. Thirty-two neonates were homozygous for HbC. The incidence of the HbC trait was 1:6; incidence of the HbS trait was 1:14. A centralized laboratory for neonatal screening of SCDs was established. CONCLUSIONS: SCDs should be considered a major public health problem in Ouagadougou. A neonatal screening programme should be implemented, but to be effective it requires strategies adapted to the local situation.

The place of capillary electrophoresis techniques in screening for haemoglobinopathies.

BACKGROUND: Capillary zone electrophoresis (CZE) methods are new laboratory diagnostic tools. The screening of haemoglobin (Hb) variants by a battery of two automated CZE methods at alkaline and acid pH, followed by micellar electrokinetic capillary chromatography (MECC), has been evaluated. METHODS: Three hundred and ninety-two patients' samples with an abnormal haemoglobin fraction, detected either by isoeletric focusing (IEF) or by automated cation-exchange exchange high-performance liquid chromatography (automated HPLC), were tested by both CZE methods. Their performances were compared with IEF and automated HPLC techniques. The place of MECC has been evaluated. RESULTS: Using both CZE methods, the clinically relevant variants (HbS, HbC, etc.) as well as 15 of 20 clinically silent variants tested were separated from HbA. Using the alkaline CZE method, a presumptive identification of Hb Bart's, HbH and Hb Constant Spring was obtained. Complementary testing by MECC has demonstrated it to be an aid in distinguishing the globin chain mutations before confirmation by DNA or protein structure analysis. CONCLUSION: Using only one device, alkaline CZE, acid CZE and MECC methods offer high resolution, automated sampling and rapid analysis. They are good tools for screening of haemoglobinopathies and can replace conventional techniques.

Relationship among maternal serum endocrinology, placental karyotype, and intervillous circulation in early pregnancy failure.

OBJECTIVE: To evaluate the relationship among maternal serum endocrinology, placental karyotype, and intervillous blood flow in missed miscarriage. DESIGN: Cross-sectional study of maternal serum, transvaginal ultrasound/Doppler, and placental cytogenetic and immunohistochemical investigations. SETTING: Tertiary care academic hospital. PATIENT(S): One hundred fifty-two women with missed miscarriage between 7 and 13 weeks of gestation. INTERVENTION(S): Ultrasound features, placental intervillous circulation findings on color Doppler imaging, and maternal serum level of alpha-fetoprotein (AFP), beta-hCG, E(2), P, and inhibin A were compared retrospectively with placenta karyotype and hCG immunochemistry. MAIN OUTCOME MEASURES: Data were analyzed according to karyotype results, presence or absence of an intervillous circulation, and delay between fetal demise and evacuation. RESULT(S): The presence of intervillous blood flow and serum concentrations of the different hormones were independent of placental karyotype. Serum beta-hCG and P were significantly higher in cases with intervillous blood flow. No difference in immunostaining for beta-hCG was found between placental tissues from normal pregnancies and missed miscarriages, but significantly higher villous beta-hCG content was found on Western blotting in miscarriage with a recent fetal demise. CONCLUSION(S): The excessive entry of maternal blood inside the placenta in the early stage of most miscarriages is unrelated to conceptus karyotype, and hCG features may reflect a temporary attempt of the trophoblast to stabilize after the initial oxidative insult.