Preoperative lifestyle modifications combined with a structured exercise protocol on patient outcomes following total knee replacement surgery: A systematic review.

BACKGROUND: Medical guidelines recommend structured prehabilitation protocols consisting of lifestyle modifications and exercise to enhance post-operative outcomes for patients undergoing a total knee replacement (TKR). However, current research showing effectiveness is limited and has primarily focused on outcomes of exercise-based prehabilitation. OBJECTIVES: To investigate whether a structured prehabilitation protocol consisting of exercise and lifestyle modifications improves physical function and patient-reported outcomes following TKR surgery compared with usual care. DESIGN: Systematic review. METHODS: Five databases were searched to identify randomised controlled trials comparing structured prehabilitation programs consisting of lifestyle modifications and exercise, with usual care, for those undergoing a TKR. Methodological quality of included studies was assessed via the RoB 2.0 tool and results synthesis via a Grading of Recommendation Assessment, Development and Evaluation approach was performed to determine the certainty evidence for each outcome. RESULTS/FINDINGS: Four studies were included in this review. Despite a positive trend supporting the inclusion of a structured prehabilitation protocol, additional improvements in post-operative pain, physical function and self-reported function were only seen in one study. Reductions in hospital length of stay were also seen in one study. No additional improvements in post-operative quality of life following prehabilitation were reported. CONCLUSION: Limited evidence supporting prehabilitation reported in our review is likely attributed to the intervention type, intensity, and delivery model of included studies. However, there remains to be strong evidence supporting the use of a structured prehabilitation protocol consisting of lifestyle modifications and exercise to improve post-operative outcome.

Harmonizing Definitions for Diagnostic Criteria and Prognostic Assessment of Transplantation-Associated Thrombotic Microangiopathy: A Report on Behalf of the European Society for Blood and Marrow Transplantation, American Society for Transplantation and Cellular Therapy, Asia-Pacific Blood and Marrow Transplantation Group, and Center for International Blood and Marrow Transplant Research.

Transplantation-associated thrombotic microangiopathy (TA-TMA) is an increasingly recognized complication of hematopoietic cell transplantation (HCT) associated with significant morbidity and mortality. However, TA-TMA is a clinical diagnosis, and multiple criteria have been proposed without universal application. Although some patients have a self-resolving disease, others progress to multiorgan failure and/or death. Poor prognostic features also are not uniformly accepted. The lack of harmonization of diagnostic and prognostic markers has precluded multi-institutional studies to better understand incidence and outcomes. Even current interventional trials use different criteria, making it challenging to interpret the data. To address this urgent need, the American Society for Transplantation and Cellular Therapy, Center for International Bone Marrow Transplant Research, Asia-Pacific Blood and Marrow Transplantation, and European Society for Blood and Marrow Transplantation nominated representatives for an expert panel tasked with reaching consensus on diagnostic and prognostic criteria. The panel reviewed literature, generated consensus statements regarding diagnostic and prognostic features of TA-TMA using the Delphi method, and identified future directions of investigation. Consensus was reached on 4 key concepts: (1) TA-TMA can be diagnosed using clinical and laboratory criteria or tissue biopsy of kidney or gastrointestinal tissue; however, biopsy is not required; (2) consensus diagnostic criteria are proposed using the modified Jodele criteria with additional definitions of anemia and thrombocytopenia. TA-TMA is diagnosed when ≥4 of the following 7 features occur twice within 14 days: anemia, defined as failure to achieve transfusion independence despite neutrophil engraftment; hemoglobin decline by ≥1 g/dL or new-onset transfusion dependence; thrombocytopenia, defined as failure to achieve platelet engraftment, higher-than-expected transfusion needs, refractory to platelet transfusions, or ≥50% reduction in baseline platelet count after full platelet engraftment; lactate dehydrogenase (LDH) exceeding the upper limit of normal (ULN); schistocytes; hypertension; soluble C5b-9 (sC5b-9) exceeding the ULN; and proteinuria (≥1 mg/mg random urine protein-to-creatinine ratio [rUPCR]); (3) patients with any of the following features are at increased risk of nonrelapse mortality and should be stratified as high-risk TA-TMA: elevated sC5b-9, LDH ≥2 times the ULN, rUPCR ≥1 mg/mg, multiorgan dysfunction, concurrent grade II-IV acute graft-versus-host disease (GVHD), or infection (bacterial or viral); and (4) all allogeneic and pediatric autologous HCT recipients with neuroblastoma should be screened weekly for TA-TMA during the first 100 days post-HCT. Patients diagnosed with TA-TMA should be risk-stratified, and those with high-risk disease should be offered participation in a clinical trial for TA-TMA-directed therapy if available. We propose that these criteria and risk stratification features be used in data registries, prospective studies, and clinical practice across international settings. This harmonization will facilitate the investigation of TA-TMA across populations diverse in race, ethnicity, age, disease indications, and transplantation characteristics. As these criteria are widely used, we expect continued refinement as necessary. Efforts to identify more specific diagnostic and prognostic biomarkers are a top priority of the field. Finally, an investigation of the impact of TA-TMA-directed treatment, particularly in the setting of concurrent highly morbid complications, such as steroid-refractory GVHD and infection, is critically needed.

Cell-free DNA analysis in current cancer clinical trials: a review.

Cell-free DNA (cfDNA) analysis represents a promising method for the diagnosis, treatment selection and clinical follow-up of cancer patients. Although its general methodological feasibility and usefulness has been demonstrated, several issues related to standardisation and technical validation must be addressed for its routine clinical application in cancer. In this regard, most cfDNA clinical applications are still limited to clinical trials, proving its value in several settings. In this paper, we review the current clinical trials involving cfDNA/ctDNA analysis and highlight those where it has been useful for patient stratification, treatment follow-up or development of novel approaches for early diagnosis. Our query included clinical trials, including the terms 'cfDNA', 'ctDNA', 'liquid biopsy' AND 'cancer OR neoplasm' in the FDA and EMA public databases. We identified 1370 clinical trials (FDA = 1129, EMA = 241) involving liquid-biopsy analysis in cancer. These clinical trials show promising results for the early detection of cancer and confirm cfDNA as a tool for real-time monitoring of acquired therapy resistance, accurate disease-progression surveillance and improvement of treatment, situations that result in a better quality of life and extended overall survival for cancer patients.

The high pressure gas capabilities at Oak Ridge National Laboratory's neutron facilities.

The study of samples subjected to high pressure gas is an important asset in materials research and has consequently been a priority of the sample environment development at the Oak Ridge National Laboratory's (ORNL) neutron program. Such effort has resulted in the availability of an extensive combination of pressure cells and gas intensifiers (both commercially available and custom made). These resources are available across both neutron facilities at ORNL: the Spallation Neutron Source and the High Flux Isotope Reactor. Current capabilities include, for example, in situ measurements up to 6 kbar and a 3 kbar hydrogen-capable intensifier with a gas recovery feature. In this communication, we will review the existing suite of high pressure gas capabilities, with special emphasis on recent in-house developments. A number of examples will be presented to illustrate how such capabilities are being deployed on neutron beamlines to enable frontier science.

Public health clinic-based hepatitis C testing and linkage to care in Baltimore.

Testing and linkage to care are important determinants of hepatitis C virus (HCV) treatment effectiveness. Public health clinics serve populations at high risk of HCV. We investigated their potential to serve as sites for HCV testing, initiation of and linkage to HCV care. Cross-sectional study of patients accessing sexually transmitted infection (STI) care at the Baltimore City Health Department (BCHD) STI clinics, from June 2013 through April 2014 was conducted. Logistic regression was used to assess factors associated with HCV infection and specialist linkage to care. Between 24 June 2013 and 15 April 2014, 2681 patients were screened for HCV infection. Overall, 189 (7%) were anti-HCV positive, of whom 185 (98%) received follow-up HCV RNA testing, with 155 (84%) testing RNA positive. Of 155 RNA-positive individuals, 138 (89%) returned to the STI clinic for HCV RNA results and initial HCV care including counselling regarding transmission and harm reduction in alcohol, and 132 (85%) were referred to a specialist for HCV care. With provision of patient navigation services, 81 (52%) attended an offsite HCV specialist appointment. Alcohol use and lack of insurance coverage were associated with lower rates of specialist linkage (OR 0.4 [95% CI 0.1-0.9] and OR 0.4 [95% CI 0.1-0.9], respectively). We identified a high prevalence of HCV infection in BCHD STI clinics. With availability of patient navigation services, a large proportion of HCV-infected patients linked to off-site specialist care.

NEOCENT: a randomised feasibility and translational study comparing neoadjuvant endocrine therapy with chemotherapy in ER-rich postmenopausal primary breast cancer.

Neoadjuvant endocrine therapy is an alternative to chemotherapy for women with oestrogen receptor (ER)-positive early breast cancer (BC). We aimed to assess feasibility of recruiting patients to a study comparing chemotherapy versus endocrine therapy in postmenopausal women with ER-rich primary BC, and response as well as translational endpoints were assessed. Patients requiring neoadjuvant therapy were randomised to chemotherapy: 6 × 3-weekly cycles FE100C or endocrine therapy: letrozole 2.5 mg, daily for 18-23 weeks. Primary endpoints were recruitment feasibility and tissue collection. Secondary endpoints included clinical, radiological and pathological response rates, quality of life and translational endpoints. 63/80 patients approached were eligible, of those 44 (70, 95% CI 57-81) were randomised. 12 (54.5, 95% CI 32.2-75.6) chemotherapy patients showed radiological objective response compared with 13 (59.1, 95% CI 36.4-79.3) letrozole patients. Compared with baseline, mean Ki-67 levels fell in both groups at days 2-4 and at surgery [fold change: 0.24 (95% CI 0.12-0.51) and 0.24; (95% CI 0.15-0.37), respectively]. Plasma total cfDNA levels rose from baseline to week 8 [fold change: chemotherapy 2.10 (95% CI 1.47-3.00), letrozole 1.47(95% CI 0.98-2.20)], and were maintained at surgery in the chemotherapy group [chemotherapy 2.63; 95% CI 1.56-4.41), letrozole 0.95 (95% CI 0.71-1.26)]. An increase in plasma let-7a miRNA was seen at surgery for patients with objective radiological response to chemotherapy. Recruitment and tissue collection endpoints were met; however, a larger trial was deemed unfeasible due to slow accrual. Both regimens were equally efficacious. Dynamic changes were seen in Ki-67 and circulating biomarkers in both groups with increases in cfDNA and let-7a miRNA persisting until surgery for chemotherapy patients.

Dendritic cell-derived tumor necrosis factor α modifies airway epithelial cell responses.

Mucosal dendritic cells (DC) are intimately associated with the airway epithelium and thus are ideally situated to be first responders to pathogens. We hypothesize that DC drive innate immune responses through early release of tumor necrosis factor (TNF) α, which drives airway epithelial cell responses. In a mouse model, TNFα release was significantly increased following a single exposure to German cockroach (GC) frass, an event independent of neutrophil recruitment into the airways. While lung epithelial cells and alveolar macrophages failed to release TNFα following GC frass exposure, bone marrow-derived DC (BMDC) produced substantial amounts of TNFα suggesting their importance as early responding cells. This was confirmed by flow cytometry of pulmonary myeloid DC. Addition of GC frass-pulsed BMDC or conditioned media from GC frass-pulsed BMDC to primary mouse tracheal epithelial cells (MTEC) or MLE-15 cells induced chemokine (C-C) motif ligand (CCL) 20 and granulocyte macrophage (GM) colony-stimulating factor (CSF), both of which are important for DC recruitment, survival and differentiation. Importantly, DC do not produce CCL20 or GM-CSF following allergen exposure. Blocking TNFα receptor 1 (TNFR1) completely abolished chemokine production, suggesting that BMDC-derived TNFα induced airway epithelial cell activation and enhancement of the innate immune response. Lastly, blocking TNFR1 in vivo resulted in significantly decreased CCL20 and GM-CSF production in the lungs of mice. Together, our data strongly suggest that DC-derived TNFα plays a crucial role in the initiation of innate immune responses through the modification of airway epithelial cell responses.

The presence of disseminated tumour cells in the bone marrow is inversely related to circulating free DNA in plasma in breast cancer dormancy.

BACKGROUND: The aim of this study was to gain insight into breast cancer dormancy by examining different measures of minimal residual disease (MRD) over time in relation to known prognostic factors. METHODS: Sixty-four primary breast cancer patients on follow-up (a median of 8.3 years post surgery) who were disease free had sequential bone marrow aspirates and blood samples taken for the measurement of disseminated tumour cells (DTCs), circulating tumour cells (CTCs) by CellSearch and qPCR measurement of overlapping (96-bp and 291-bp) amplicons in circulating free DNA (cfDNA). RESULTS: The presence of CTCs was correlated with the presence of DTCs measured by immunocytochemistry (P=0.01) but both were infrequently detected. Increasing cfDNA concentration correlated with ER, HER2 and triple-negative tumours and high tumour grade, and the 291-bp amplicon was inversely correlated with DTCs measured by CK19 qRT-PCR (P=0.047). CONCLUSION: Our results show that breast cancer patients have evidence of MRD for many years after diagnosis despite there being no overt evidence of disease. The inverse relationship between bone marrow CK19 mRNA and the 291-bp amplicon in cfDNA suggests that an inverse relationship between a measure of cell viability in the bone marrow (DTCs) and cell death in the plasma occurs during the dormancy phase of breast cancer.

Detection of HER2 amplification in circulating free DNA in patients with breast cancer.

BACKGROUND: Human epidermal growth factor receptor 2 (HER2) is amplified and overexpressed in 20-25% of breast cancers. This study investigated circulating free DNA (cfDNA) for detection of HER2 gene amplification in patients with breast cancer. METHODS: Circulating free DNA was extracted from plasma of unselected patients with primary breast cancer (22 before surgery and 68 following treatment), 30 metastatic patients and 98 female controls using the QIAamp Blood DNA Mini Kit (Qiagen). The ratio of HER2 to an unamplified reference gene (contactin-associated protein 1 (CNTNAP1)) was measured in cfDNA samples by quantitative PCR (qPCR) using SK-BR-3 cell line DNA as a positive control. RESULTS: We validated the qPCR assay with DNA extracted from 23 HER2 3+ and 40 HER2-negative tumour tissue samples; the results agreed for 60 of 63 (95.2%) tumours. Amplification was detected in cfDNA for 8 of 68 patients following primary breast cancer treatment and 5 of 30 metastatic patients, but was undetected in 22 patients with primary breast cancer and 98 healthy female controls. Of the patients with amplification in cfDNA, 10 had HER2 3+ tumour status by immunohistochemistry. CONCLUSIONS: The results demonstrate for the first time the existence of amplified HER2 in cfDNA in the follow-up of breast cancer patients who are otherwise disease free. This approach could potentially provide a marker in patients with HER2-positive breast cancer.

Use of hydroxypropylmethylcellulose 2% for removing adherent silicone oil from silicone intraocular lenses.

BACKGROUND/AIMS: To investigate the effect of hydroxypropylmethylcellulose (HPMC) on the physical interaction (contact angle) between silicone oil and a silicone intraocular lens (IOL). METHODS: In vitro experiments were performed, to determine the effect of HPMC (0.5%, 1% or 2%), with or without an additional simple mechanical manoeuvre, on the contact angle of silicone oil at the surface of both silicone and acrylic (control) IOLs. A balanced salt solution chamber was used. The study group comprised 21 silicone and nine acrylic IOLs. RESULTS: The median contact angle of silicone oil on silicone IOL was 99 degrees. The addition of HPMC 2% alone did not significantly alter the contact angle. HPMC 2% combined with an additional single mechanical manoeuvre increased the contact angle to 180 degrees (greater non-wetting), with complete separation of silicone oil from silicone IOL within 1 min. The manoeuvre alone, or in conjunction with a lower concentration of HPMC (0.5 or 1%), was ineffective in increasing the contact angle. CONCLUSION: We present a novel, non-toxic technique of using hydroxypropylmethylcellulose 2% combined with a simple mechanical manoeuvre, for the removal of adherent silicone oil droplets from silicone intraocular lenses.

Most low-level microsatellite instability in colorectal cancers can be explained without an elevated slippage rate.

Many cancers show a low level of microsatellite slippage and are labelled MSI-L (microsatellite instability--low). However, it is unclear whether this slippage can be attributed to some underlying genetic change that results in a mutator phenotype, analogous to mismatch repair deficiency in MSI-H cancers, or whether the apparent instability is the result of relatively frequent normal somatic slippage. Here, we have used a mathematical model of microsatellite slippage during cancer growth to estimate the degree of microsatellite slippage expected in a cancer due to normal somatic slippage. We compared the model to the slippage observed in 42 non-MSI-H cancers that were macro-dissected into four distinct regions and genotyped at N = 9 microsatellite loci. When the slippage rate was set at mu = 10(-5) per locus per division, ten cancers showed a level of slippage in at least one region that was too severe to be expected from normal somatic slippage alone, suggesting that these cancers had acquired MSI-L. Only one of these ten cancers had putative MSI-L in all four regions. When we considered a slightly higher slippage rate of mu = 5 x 10(-5), none of the cancers showed a degree of slippage that could not be reasonably explained by normal somatic slippage. Counting the number of 'unstable' loci was a poor indicator of putative MSI-L status. We conclude that most low-level microsatellite instability in colorectal cancers can be explained without requiring an elevated slippage rate during neoplastic development, and hence there is little evidence for a discrete MSI-L group of cancers. Putative MSI-L status is indicated by the presence of at least one locus that has multiple alleles that differ by at least five motif repeats from the germline. If an underlying genetic change does cause MSI-L, it appears to be a relatively uncommon event that occurs late in oncogenesis.

Analysis of copy number changes suggests chromosomal instability in a minority of large colorectal adenomas.

We have examined chromosomal-scale mutations in 34 large colorectal adenomas (CRAs). A small number of changes (median = 2, IQR = 0-4) were found by array-comparative genomic hybridization (aCGH) in most tumours. The most common changes were deletions of chromosomes 1p, 9q, 17, 19, and 22, and gains of chromosomes 13 and 21. SNP-LOH analysis and pseudo-digital SNP-PCR analysis detected occasional copy-neutral LOH. Some aCGH changes found frequently in colorectal carcinomas, such as deletions of chromosomes 4q and 18q, were very infrequent in the adenomas. Almost all copy number changes were of small magnitude, far below the predicted levels even for single copy gain/loss; investigation suggested that these changes were either artefactual or occurred in sub-clones within the tumours. In some cases, these sub-clones may have represented progression towards carcinoma, but comparison with aCGH data from carcinomas showed this to be unlikely in most cases. In two adenomas, there was evidence of a large, outlying number of copy number changes, mostly resulting from part-chromosome deletions. Overall, moreover, there was evidence of a tendency towards part-chromosome deletions-consistent with chromosomal instability (CIN)--in about one-sixth of all tumours. However, there was no evidence of CIN in the form of whole-chromosome copy number changes. Our data did not support previous contentions that CRAs tend to show chromosome breakage at fragile sites owing to CIN associated with an elevated DNA damage response. Chromosomal-scale mutations occur in some CRAs; although CIN is not the norm in these lesions, it probably affects a minority of cases.

Mechanism and function of heat shock-dependent IkappaBalpha expression.

OBJECTIVE: Heat shock is known to inhibit activation of the NF-kappa B pathway. One potential mechanism of this effect is de novo expression of the intracellular NF-kappaB inhibitor, Ikappa Balpha. Herein we sought to elucidate the mechanisms by which heat shock induces Ikappa Balpha gene expression and the functional consequences of heat shock-mediated Ikappa Balpha gene expression in A549 cells. METHODS: Nuclear run-on assays demonstrated that heat shock had a small effect on transcription of the Ikappa Balpha gene relative to the level of steady state Ikappa Balpha mRNA that is seen following heat shock. Accordingly, we determined the effect of heat shock on Ikappa Balpha mRNA stability by treating cells with actinomycin D to induce transcriptional arrest. RESULTS: The half-life of IkappaBalpha mRNA was 36 +/- 7.2 min in control cells and 101 +/- 3.7 min in cells subjected to heat shock. These data were consistent with heat shock-mediated increased stability of Ikappa Balpha mRNA. Heat shock induced activation of p38 MAP kinase and inhibition of p38 MAP kinase substantially reduced heat shock-dependent expression of Ikappa Balpha mRNA. After a 4 h recovery period from heat shock, there was inhibition of tumor necrosis factor-alpha-mediated NF-kappaB activation. The introduction of an Ikappa Balpha anti-sense oligonucleotide reversed this inhibitory effect of heat shock. CONCLUSIONS: We conclude that heat shock increases IkappaBalpha gene expression primarily by increasing Ikappa Balpha mRNA stability and this effect is partially dependent on p38 MAP kinase. The functional consequence of heat shock-mediated Ikappa Balpha gene expression is inhibition of NF-kappaB activation.

German cockroach proteases regulate matrix metalloproteinase-9 in human bronchial epithelial cells.

BACKGROUND: Matrix metalloproteinases (MMPs) digest extracellular matrix proteins and may play a role in the pathogenesis of bronchial asthma. MMP-9 levels are increased in the bronchoalveolar lavage fluid and sputum of asthmatics compared with that of controls. As exposure to cockroaches is an environmental risk factor for asthma, we sought to investigate the role of German cockroach fecal remnants (frass) on MMP-9 expression. METHODS: Human bronchial epithelial cells (16HBE14o-) and primary normal human bronchial epithelial cells were treated with cockroach frass in the absence or presence of tumor necrosis factor (TNF)alpha. MMP-9 mRNA, protein levels and pro-MMP-9 activity were determined using real-time polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA) and zymogram assays. Pretreatment of frass with aprotinin abolished protease activity. PD98059, a chemical inhibitor of extracellular signal regulated kinase (ERK), and SLIGKV, an activator of protease-activated receptor (PAR)-2 were also used. AP-1DNA binding was determined by electrophoretic mobility shift assay (EMSA) and ERK phosphorylation by Western blot analysis. RESULTS: Cockroach frass augmented TNFalpha-mediated MMP-9 mRNA and protein expression by a mechanism dependent on active serine proteases within frass and not on endogenous endotoxin. Frass increased ERK phosphorylation, and chemical inhibition of ERK attenuated cockroaches' effects on MMP-9. Serine proteases are known to activate the PAR-2 receptor. We found that selective activation of PAR-2 using the peptide SLIGKV augmented TNFalpha-induced MMP-9 protein levels and increased ERK phosphorylation. Frass and SLIGKV each increased AP-1 translocation and DNA binding. CONCLUSIONS: These data suggest that German cockroach frass contains active serine proteases which augment TNFalpha-induced MMP-9 expression by a mechanism involving PAR-2, ERK and AP-1.

Expression and adaptive regulation of amino acid transport system A in a placental cell line under amino acid restriction.

Trans-placental transport of amino acids is vital for the developing fetus. Using the BeWo cell line as a placental model, we investigated the effect of restricting amino acid availability on amino acid transport system type A. BeWo cells were cultured either in amino acid-depleted (without non-essential amino acids) or control media for 1, 3, 5 or 6 h. System A function was analysed using alpha(methyl-amino)isobutyric acid (MeAIB) transcellular transport studies. Transporter (sodium coupled neutral amino acid transporter (SNAT1/2)) expression was analysed at mRNA and protein level by Northern and Western blotting respectively. Localisation was carried out using immunocytochemistry. MeAIB transcellular transport was significantly (P < 0.05) increased by incubation of the cells in amino acid-depleted medium for 1 h, and longer incubation times caused further increases in the rate of transfer. However, the initial response was not accompanied by an increase in SNAT2 mRNA; this occurred only after 3 h and further increased for the rest of the 6-h incubation. Similarly, it took several hours for a significant increase in SNAT2 protein expression. In contrast, relocalisation of existing SNAT2 transporters occurred within 30 min of amino acid restriction and continued throughout the 6-h incubation. When the cells were incubated in medium with even lower amino acid levels (without non-essential plus 0.5 x essential amino acids), SNAT2 mRNA levels showed further significant (P < 0.0001) up-regulation. However, incubation of cells in depleted medium for 6 h caused a significant (P = 0.014) decrease in the expression of SNAT1 mRNA. System L type amino acid transporter 2 (LAT2) expression was not changed by amino acid restriction, indicating that the responses seen in the system A transporters were not a general cell response. These data have shown that placental cells adapt in vitro to nutritional stress and have identified the physiological, biochemical and genomic mechanisms involved.

Increasing hCG concentrations during storage at (+)4 degrees C with the Bayer Centaur Total hCG method.

BACKGROUND: The interpretation of changes in serum human chorionic gonadotropin (hCG) concentrations in early pregnancy requires a knowledge of the day-to-day imprecision of the method at the concentrations measured. We therefore attempted to determine the analytical imprecision of our total hCG method over a 4-week period at concentrations up to 20,000 IU/L. METHODS: Serum specimens with hCG concentrations between 100 and 20,000 IU/L were collected during early pregnancy and analysed using the Bayer Centaur Total hCG method at weekly intervals after storage at (+)4 degrees C and (-)20 degrees C. RESULTS: The reproducibility of hCG results over a 4-week period in refrigerated specimens with an initial hCG concentration below 1000 IU/L was poor (mean coefficient of variation = 17.4%) and there was an apparent increase in serum hCG concentrations of up to 68%. Similar changes occurred in some specimens stored at (-)20 degrees C. Further experiments confirmed that significant increases can occur during the first week following specimen collection. CONCLUSIONS: Total hCG concentrations in serum specimens collected during early pregnancy increase significantly during storage when measured by the Bayer Centaur Total hCG method, possibly due to a conformational change in hyperglycosylated hCG. When this method is used for monitoring early pregnancy, specimens should ideally be analysed on the day of collection.