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PubMed2005

Biochemical markers of neurotoxicity in wildlife and human populations: considerations for method development.

Disruption of neurochemical parameters in blood and brain tissues can be used as early biomarkers of neurotoxicity in human and wildlife epidemiological studies. To investigate the feasibility of biomarker measurements in field samples obtained from remote locations, tissue storage limits were determined with human blood and mink cortex tissue using efficient and cost-effective microplate assays. Results show that isolated blood platelets and plasma can be stored at 4 degrees C for 4 wk before measurement of monoamine oxidase (MAO) and cholinesterase (ChE) activities, while human lymphocytes can be stored at 4 degrees C for up to 2 d before muscarinic acetylcholine (mACh) receptor binding analysis. Blood cells stored frozen resulted in decreased MAO activity and mACh receptor function. These data suggest that mink brain tissue obtained from field samples can be stored at various temperatures without affecting dopamine (D2) and mACh receptor densities; however, MAO and ChE activities were most stable in samples stored in a -20 degrees C domestic freezer or at 4 degrees C. Multiple freeze/thaw cycles alter mACh and D2 receptors and MAO activity in mink cortex samples and should therefore be minimized. In conclusion, these neurochemical biomarkers can efficiently be measured in large human and wildlife neurotoxicity studies, provided proper storage conditions are maintained.

Adult↗
PubMed2005

Laboratory safety during assisted reproduction in patients with blood-borne viruses.

For couples where one or both partners are infected with human immunodeficiency virus or hepatitis C, the doors to receiving fertility care are opening as a result of better antiviral medication, better long-term prognosis and consequent changes in attitude. In line with this, fertility centres electing to treat couples with blood-borne viral (BBV) infection need to re-examine their policies and procedures to ensure the safety of their staff and both non-infected and infected patients during assisted reproduction treatments. At a time when the European Tissue Directive aims to introduce quality standards for assisted reproduction throughout Europe, we highlight the risks involved when treating patients with known BBV infections and argue that safety cannot be met with any certainty unless samples from such patients are handled within a separate high security laboratory or laboratory area, technically adapted to ensure minimal cross-contamination risk to uninfected gametes and embryos.

Blood-Borne Pathogens↗
PubMed1995

Clinical evaluation and reproducibility of the Pastorex Aspergillus antigen latex agglutination test for diagnosing invasive aspergillosis.

AIMS: The performance of the Pastorex Aspergillus antigen latex agglutination test for the detection of galactomannan in sera of patients at risk for invasive aspergillosis was evaluated, and the impact of storage on the reproducibility of the antigen titre was tested. METHODS: During a one year period, 392 serum samples were obtained from 46 patients at risk for invasive aspergillosis and tested for the presence of galactomannan using an Aspergillus latex agglutination test (Pastorex). Twenty three positive serum samples which had been stored at -20 degrees C for 2-16 months were retrospectively retested. Furthermore, two positive serum samples were stored at -20 degrees C and -70 degrees C and prospectively tested at three month intervals for a period of 15 months. RESULTS: The Pastorex Aspergillus test was positive in eight patients with microbiological, radiological, or histological evidence for invasive aspergillosis, but was negative in the initial serum sample from five of these patients. In two patients with histological evidence for invasive aspergillosis no positive reaction was found in six samples. Six of 13 (45%) serum samples which had been stored at -20 degrees C for longer than six months had lost reactivity, while one of 10 (10%) samples had lost reactivity when stored up to six months. Two serum samples which had been stored at -20 degrees C and -70 degrees C and prospectively retested at three month intervals for 15 months, maintained stable antigen titres. CONCLUSIONS: The Pastorex Aspergillus test is too insensitive to diagnose invasive aspergillosis in an early stage, but may contribute to the diagnosis when cultures remain negative and serial samples are obtained. To maintain a good reproducibility, serum samples should be stored at -70 degrees C when the period of storage exceeds six months.

Adolescent↗
PubMed2003

Measurements of serum S-100B protein: effects of storage time and temperature on pre-analytical stability.

Measurement of S-100B protein in serum or cerebrospinal fluid is increasingly used as a molecular marker of brain damage. However, there is no information about the pre-analytical in vitro stability of S-100B. We have investigated whether storing blood samples at different time periods and conditions affected the measured levels of S-100B protein. Blood samples were taken from 29 neurosurgical patients with normal serum S-100B values and from seven patients with increased serum S-100B values. Blood samples were immediately divided into nine aliquots for measuring S-100B immediately and after 4, 8, 12, and 24 hours, stored at room temperature or at 4 degrees C. Measurement of S-100B was performed using the LIAISON assay (Byk-Sangtec Diagnostica, Dietzenbach, Germany). Moreover, in 10 additional patients the effect of freezing the serum and thawing the sample after 24 hours was investigated. There were no differences between the results of S-100B measurements after storing the sample at different temperatures and time periods. There was no trend towards higher or lower values in all three groups. Therefore, blood samples may be collected as part of the daily clinical routine without time constraint and even stored overnight without affecting S-100B serum levels when measured with the LIAISON Sangtec 100 assay.

Blood Preservation↗
PubMed1996

A comparison of time, temperature, and refreezing variables on frozen sperm motility recovery.

OBJECTIVE: To compare motility recovery among methods of thawing semen in serial freeze-thaw processes. DESIGN: A comparison of treatments. SETTING: Semen cryobank laboratory. PARTICIPANTS: Semen samples from an existing cryobank. INTERVENTIONS: Portions of each of 35 ejaculates were thawed serially in either a 37 degrees C water bath for 10 minutes or at room temperature for 30 minutes. Motility was estimated, and the samples were immediately refrozen. Thaws were performed initially (T1), at 24 hours (T2), and at 1 week (T3). MAIN OUTCOME MEASURE: Percent motility. RESULTS: The initial motility of all samples averaged 73.2%. Serial thaws at 37 degrees C yielded mean motility recoveries of 34.0%, 21.8%, and 8.3% for T1, T2, and T3, respectively. Those at room temperature were 22.4%, 7.9%, and 1.3%. Motilities progressively decreased from initial to final thaw regardless of temperature. Semen thawed at 37 degrees C had a higher motility recovery at all thaw times compared with samples thawed at room temperature. CONCLUSIONS: For sperm frozen rapidly in liquid nitrogen and for sperm subjected to multiple freeze-thaw processes, motility recovery is improved if thaw is carried out at 37 degrees C versus room temperature.

Cryopreservation↗
PubMed2004

Efficient production of sex-identified and cryosurvived bovine in-vitro produced blastocysts.

To establish a protocol for production of bovine in-vitro produced (IVP) blastocysts that were sex-identified and cryopreserved, we examined the sexing efficiency and accuracy of Day-3 and Day-4 embryos by polymerase chain reaction (PCR), the development of the biopsied embryos into Day-7 blastocysts and the freezability of these blastocysts by vitrification in gel-loading tips. One or two blastomeres were isolated from IVP embryos at either the 8-cell or 16-cell stage (Days 3 and 4, respectively) by a pressing-out method, and were then subjected to primer extension preamplification (PEP)-PCR. The successful sex-identification rate of biopsied samples amplified, purified and analyzed for sex by a second PCR (88.9%) was higher than that of those amplified and analyzed without purification (32.0%). Developmental rates into Day-7 blastocysts of biopsied embryos (Day-3, 65.5%; Day-4, 70.8%) were similar to those of non-biopsied control embryos (Day-3, 74.5%; Day-4, 65.1%). Total cell numbers and the inner cell mass (ICM) ratio of blastocysts derived from biopsied embryos were also comparable with those of control embryos. Blastocysts were vitrified-warmed in the presence of 20% DMSO, 20% ethylene glycol and 0.6M sucrose using gel-loading tips as containers. The proportions of biopsied blastocysts that were hatched or hatching rates after warming were high, regardless of the biopsy time (Day-3, 94.1%; Day-4, 91.9%), similar to the rates for control blastocysts (Day-3, 97.5%; Day-4, 96.9%). In conclusion, a protocol that allows sexing of Day-3 and Day-4 bovine embryos without compromising either the developmental ability to the blastocyst stage or freezability of Day-7 blastocysts was developed.

Animals↗
PubMed2003

Storage of red deer epididymides for four days at 5 degrees C: effects on sperm motility, viability, and morphological integrity.

The purpose of this study was to assess the sperm motility, the plasma membrane integrity and the morphology of red deer spermatozoa when maintained within epididymides stored for 4 days at 5 degrees C, and to evaluate whether such stored spermatozoa are able to withstand a refrigeration process. Thirty pairs of testes, with attached epididymides, were collected from 30 hunter-killed mature stags (Cervus elaphus hispanicus), and spermatozoa from each one of the pairs were immediately collected in Triladyl medium, evaluated and refrigerated (Control Group). The remaining testes and epididymides were gradually cooled to 5 degrees C and stored for 1, 2, 3, and 4 days (Experimental Groups), after which spermatozoa were processed as described previously for the control group. The effects on spermatozoa that had been stored within epididymides for various times were determined by assaying sperm motility index (SMI), plasma membrane integrity and sperm morphology (SM). In the same way, SMI and SM were assessed after spermatozoa refrigeration at 5 degrees C for 3 hours in different groups (SMI-R, SM-R). There was no significant decrease in plasma membrane integrity of spermatozoa recovered from epididymides stored at 5 degrees C for 4 days. Similarly, the percentage of morphologically normal spermatozoa remained unaffected during the first 3 days of storage. In contrast, during storage sperm motility evaluation revealed significantly (P<0.05) lower SMI values for samples from epididymides stored 2, 3, and 4 days (47.7+/-3.6, 45.5+/-4.4, 44.1+/-5.2) than that of the control group (57.6+/-1.6). Similar results were obtained after refrigeration of spermatozoa in Triladyl at 5 degrees C. These data suggest that it might be possible to recover functional spermatozoa from red deer epididymides stored at 5 degrees C during several days when epididymal spermatozoa cannot be collected and cryopreserved immediately.

Animals↗
PubMed1993

Antifreeze glycoproteins increase solution viscosity.

A new microsensor that can analyze microliter volume samples was used to measure the viscosity of aqueous solutions of antifreeze glycoproteins as a function of temperature and concentration. The results show that at physiological concentrations which naturally occur in the fish, the antifreeze proteins significantly increase aqueous solution viscosity. The probability for ice nucleation is inversely proportional to viscosity. Therefore, the increased viscosity could explain, in part, reports on the beneficial effects of antifreeze glycoproteins during cryopreservation by vitrification. Reducing the probability for ice nucleation could be also beneficial for the survival of cold-water fish in their natural habitat. Millimolar concentrations of antifreeze proteins increase aqueous solution viscosity to values comparable with those of conventional cryoprotectants in molar concentrations.

Animals↗
PubMed1986

Identification of isolate-specific proteins on sorbitol-enriched Plasmodium falciparum infected erythrocytes from Gambian patients.

We have compared the surface radio-iodinated proteins of uninfected and Plasmodium falciparum-infected erythrocytes from natural infections of human patients. Cryopreserved infected blood from Gambian children with falciparum malaria was thawed, cultured to the middle trophozoite stage, and surface radio-iodinated. Trophozoite-infected cells were enriched about 10-fold on a Percoll gradient newly designed to separate cells based on their differential permeability to sorbitol. Infected blood was radio-iodinated and erythrocytes from the fraction enriched in parasitized cells and uninfected erythrocytes from the same sample obtained from the gradient and compared by SDS-PAGE and autoradiography. In each sample, parasitized erythrocytes contained one or more polypeptides of very high molecular weight (Mr 250 000-300 000) that were not found on uninfected erythrocytes from the same patient. These proteins were isolate-specific in size and number, suggesting that natural isolates contain a variable number of different P. falciparum phenotypes for this surface protein. In addition, these radio-iodinated surface proteins could not be extracted from the host cell membrane by the non-ionic detergent Triton X-100, but were extracted by SDS. The properties of these proteins suggest they are the equivalent for natural infections of the strain-dependent antigen previously described (Leech, Barnwell, Miller & Howard, 1984) on the surface of P. falciparum-infected Aotus erythrocytes. In addition, we observed a second parasite-dependent modification of labelled proteins on infected erythrocytes with the appearance of a new band of Mr 30 000. There were also variations in the pattern of radio-isotope labelled proteins on uninfected erythrocytes from different patients.

Animals↗
PubMed2026

A Clinically Integrated Pediatric Patient-Derived Xenograft Program Enables Evaluation of Cohort and Patient-Specific Biology and Therapeutic Strategies.

UNLABELLED: Preclinical translational research has increasingly utilized patient-derived xenograft (PDX) models for mechanistic and experimental therapeutic studies, yet most existing models have been developed from adult cancer types. We describe the establishment of a PDX program to expand the availability of pediatric-specific PDXs for preclinical research and enable studies of pediatric cancer histologies, including ultrarare diseases. Processes for PDX generation were integrated into established clinical workflows to facilitate universal model generation. Methodologies for tissue procurement, processing, and cryopreservation were optimized to enable intra- and interinstitutional PDX model generation. Over a 6-year span, 388 PDX tumor models representing more than 40 diagnoses were generated, including ultrarare tumors and longitudinal models established from pretherapy, posttherapy, and relapse tumors from the same patient. Genomic characterization of these PDXs demonstrates excellent concordance and recapitulation of molecular alterations of the source tumor. Successful PDX generation was enhanced from relapsed samples, was higher in sarcomas compared with other solid tumor types, and was a negative prognosticator for clinical outcome. With a broad portfolio of molecularly annotated models, we demonstrate utility for validating cross-histology biomarker-driven therapeutic strategies by demonstrating antitumor activity of an MAT2A inhibitor in MTAP-deficient PDXs. Universal model creation also allows for experimental validation of therapeutic hypotheses on a patient-specific basis, as we describe the characterization of a novel RAF1 fusion (EPB41L2::RAF1) in an osteosarcoma PDX. Development of a diverse collection of pediatric PDX models enables hypothesis-driven and cross-histology studies that expand our understanding of cancer biology and aid ongoing drug prioritization efforts in rare tumors. SIGNIFICANCE: A clinically integrated, genomically annotated pediatric PDX portfolio supported by systematic benchmarking of model generation facilitates exploratory biomarker-driven and patient-specific translational studies.

Humans↗
PubMed2005

Application of sexed semen technology to in vitro embryo production in cattle.

Use of sexed semen in conjunction with in vitro embryo production is a potentially efficient means of obtaining offspring of predetermined sex. For thousands of years, livestock owners have desired a methodology to predetermine the sex of offspring for their herds. The ability to sort individual sperm cells into viable X- and Y-chromosome-bearing fractions made producers' sex selection dreams reality in the 1990s and now semen can be sexed with greater than 90% accuracy with use of a flow cytometric cell sorter. Several concerns regarding the implementation of sexed semen technology include the apparent lower fertility of sorted sperm, the lower survival of sorted sperm after cryopreservation and the reduced number of sperm that could be separated in a specified time period. These issues are discussed in this review. There are also a number of issues that appear to influence the success rates of using sexed semen to produce bovine embryos in vitro. These issues include reductions in fertilization rates, lower cleavage rates, blastocyst rates and pregnancy rates, partial capacitation of the sperm, dilute sperm samples and sire variation. These subjects are also addressed in this paper. Finally, we will describe a recent field trial in which female Holstein embryos produced using the combined technologies of sex-selected semen and microfluidics were transferred either as single or bilateral twin embryos into beef cattle recipients, demonstrating these technologies' contributions to viable embryo production. The results indicate that large-scale transfer of in vitro produced, Holstein heifer embryos to beef recipients is a feasible production scheme.

Animals↗
PubMedDate not supplied

Persistence of Plasmodium falciparum HRP-2 in successfully treated acute falciparum malaria.

The potential for Plasmodium falciparum histidine-rich protein-2 (PfHRP-2) dipstick tests to predict antimalarial treatment failure was investigated in a prospective study in Thailand of 38 patients admitted with severe malaria and 54 hospitalized with uncomplicated P. falciparum infections. Of these, 40 had subsequent recrudescence of their infections. Overall, 89% of patients with severe malaria and 61% of patients with uncomplicated malaria had positive PfHRP-2 dipstick tests for > 2 weeks following the start of treatment. Persistence was correlated positively with admission parasite counts, PfHRP-2 intensity scores and disease severity. PfHRP-2 tests which remained positive for > 2 weeks and PfHRP-2 reactive intensity scores on admission, at day 7 and day 14 did not predict treatment failure independent of admission parasitaemia. Freezing and thawing the blood samples did not significantly affect PfHRP-2 results tested by the dipstick technique. The PfHRP-2 dipstick test provides a useful indicator of recent severe malaria, but does not predict the therapeutic response.

Adult↗
PubMed1999

Quantitative determination of a potent lipopolysaccharide antagonist, E5564, in rat and dog plasma by high-performance liquid chromatography with fluorescence detection.

The assay method was established for the quantification of a potent lipopolysaccharide (LPS) antagonist, E5564, in rat and dog plasma using HPLC. E5564 and the I.S. (an analogue of E5564) were extracted and derivatized with 9-Anthryldiazomethane (ADAM reagent) to be given fluorescence. LC-MS analysis indicated that single molecule of E5564 was coupled with two molecules of ADAM reagent at one on each of the phosphorus groups. After solid-phase extraction, ADAM derivatives of E5564 and the I.S. were separated on an ODS column using methanol/ethanol containing sodium acetate as a mobile phase at 1.2 ml/min (gradient elution), and detected by a fluorescence detector (excitation: 254 nm, emission: 415 nm). The intra-day and inter-day precision were less than 14.4%, and accuracy were within +/-13.0% in the concentration range of 30 to 20,000 ng/ml plasma in both species. E5564 was stable for at least 13 days in rat and dog plasma at -20 degrees C, and the processed sample was stable for up to 14 days at 4 degrees C. This validated method was successfully applied to the evaluation of the pharmacokinetics of E5564 in rats and dogs after single bolus intravenous doses.

Animals↗
PubMed1994

Co-culture with Vero cell monolayer maintains the motility of asthenozoospermic semen samples.

The clinical effectiveness of co-culture with Vero (Green monkey kidney) cell monolayer in maintaining the motility and viability of fresh asthenozoospermic semen (18 samples) and frozen-thawed semen with poor motility (motility fraction < 50%) (15 samples) in a 24-h period was evaluated. Co-culture with Vero cell monolayer in human tubal fluid (HTF) medium for 24 h resulted in a statistically better maintenance of motility percentage (P < 0.005), mean amplitude of lateral head displacement (ALH) (P < 0.005), and mean track speed (VCL) (P < 0.05) than culture in HTF medium alone. However, these motility parameters (motility percentage, ALH, VCL) declined soon after removal of spermatozoa from the monolayer. Co-culture with Vero cell monolayer also maintained the viability percentage of these sperm samples (52% of the original value) after the 24-h period compared with culture in HTF medium alone (22% of the original) (52% versus 22%, P < 0.05). It is concluded that Vero cell monolayer is effective in the maintenance of motility and viability of asthenozoospermic semen or frozen-thawed semen with poor motility. This co-culture system may be beneficial in enhancing the in-vitro performance of asthenozoospermic semen samples in the practice of assisted reproductive technology. However, its safety needs further evaluation.

Animals↗
PubMed1995

Double-layer coating for high-resolution low-temperature scanning electron microscopy.

Specimen damage caused by mass loss due to electron beam irradiation is a major limitation in low-temperature scanning electron microscopy of bulk specimens. At high primary magnifications (e.g., 100,000 x) a hydrated sample is usually severely damaged after one slow scan (about 3000 e-nm-2). The consequences of this beam damage are significantly reduced by coating the frozen-hydrated sample with a 5-10-nm-thick carbon layer. Since this layer covers up surface details, the sample is first unidirectionally shadowed with a thin heavy metal layer (e.g., 2 nm of platinum) that is in close contact with the biological surface (double layer coating). This heavy metal layer can be visualized in field-emission scanning electron microscopy with the material-dependent backscattered electron signal. The method allows for routine observation of large frozen-hydrated samples. By use of an in-lens field-emission SEM and a sensitive backscattered electron detector, structural information comparable to that obtained with the transmission electron microscopy freeze-fracture replica technique can be achieved.

Carbon↗
PubMed1997

Induction of acrosome reaction in dog sperm by calcium ionophore.

The sensitivity of the plasma membrane to calcium ionophore (A23187) challenge was studied in dog sperm using fluorescein lectin staining for the assessment of acrosomal status and viability. Second fraction ejaculates from 5 dogs were washed, resuspended in Ca(2+)-free (EDTA-treated), 50, 100, 500, 1000 and 2000 microM/l Ca(2+)-containing Sp-TALP medium and induced with 50, 250, 500, 1000, 2500 and 5000 nM/l calcium ionophore. Samples were collected from each aliquot after 30 and 60 min of induction to assess the percentage of acrosome reacted sperm cells (AR rate), viability and motility by fluorescein isothiocyanate conjugated peanut agglutinin (FITC-PNA) and ethidium-homodimer combined staining. On each slide, 200 sperm cells were assessed under epifluorescence microscope (x 1250) in a blind manner. The response to ionophore challenge (AR rate, viability, motility) varied with Ca2+ and ionophore concentration in the suspension. A significantly higher AR rate was detected in samples containing 100, 500, 1000 and 2000 microM/L Ca2+ (> 40%) than in that containing 50 microM/L. Acrosome reaction could not be successfully induced in the EDTA-treated sample and in any of the aliquots in which 50, 250 and 500 nM/L ionophore concentrations were used for induction. Motility decreased drastically in all of the treated samples and stopped in that sample where as significant AR rate could be detected. Viability remained high (> 75%) during the incubation and did not differ significantly in the treated and the control groups.

Acrosome↗
PubMed2001

Dextran sedimentation in a semi-closed system for the clinical banking of umbilical cord blood.

BACKGROUND: The results of current processing procedures for reducing volume and recovering HPCs from umbilical cord blood (UCB) before cryopreservation vary. STUDY DESIGN AND METHODS: Dextran was added to bags containing UCB, followed by sedimentation for 30 minutes. The processed UCB was then frozen. RBCs, nucleated cells, MNCs, CD34+ cells, CFUs and long-term culture-initiating cells (LTC-ICs), viability, and sterility were evaluated. Fractionations in ficoll-hypaque and hydroxyethyl starch (HES) were also run in parallel for comparison. RESULTS: The nucleated cell (NC) recovery and RBC depletion were 86.1 percent and 94.3 percent, respectively (n = 50). Sedimentation with dextran also enabled the recovery of 80.7 percent MNCs and 82.6 percent CD34+ cells (n = 30). Postsedimentation samples displayed no impairment of CFU growth (n = 42, 108.7% CFU-C, 104.6% CFU-GEMM, 107% CFU-GM, and 95.7% BFU-E). Long-term cultures on five paired samples before and after sedimentation generated similar numbers of CFU-C each week (p = 0.88). Limiting dilution analysis of 12 paired pre/postsedimentation samples showed comparable median proportions of LTC-ICs (1/6494 vs. 1/5236; p = 0.18). The cell viability of 24 samples of thawed UCB after sedimentation was 90.3 percent (77.5-96%) and the recovery of CFU-C, CFU-GEMM, CFU-GM, and BFU-E of 11 postsedimentation samples was 93.4 percent, 84.9 percent, 92.3 percent, and 83.4 percent, respectively. NC recovery was significantly higher after treatment with dextran than with ficoll-hypaque (n = 30; 88.5% vs. 29.1%; p<0.005) and HES treatment (n = 21; 88.5% vs. 76.4%; p = 0.004). However, MNCs, CD34+ cells, CFUs, LTC-ICs, and RBCs were comparable. Two cycles of dextran sedimentation recovered 93.9 percent of NCs with cell viability of 98.6 percent (96.5-100%), whereas 11.7 percent of RBCs were retained (n = 20). The final yield volume was 33.5 (28-41) mL. CONCLUSION: In a semi-closed system, dextran sedimentation enabled volume reduction of UCB without significant quantitative and qualitative losses of HPCs.

Blood Banks↗
PubMed2002

Efficacy of enzyme immunoassay for the detection of Helicobacter pylori antigens in frozen stool specimens: local validation.

AIM: To investigate the efficacy of measurement of Helicobacter pylori stool antigen (HpSA) using stored frozen stool specimens, and to assess whether there were factors affecting efficacy in Hong Kong. METHODS: Patients undergoing upper endoscopy at Tuen Mun Hospital were recruited. Stool samples were saved for HpSA testing and questionnaires were completed. Stool samples were frozen immediately upon receipt and stored at -70 degrees C until tested. HpSA results were compared with rapid urease test and histology. RESULTS: One hundred and eighty-one patients were recruited. One hundred and seventy-eight patients were suitable for analysis. Eighty-three were H. pylori positive and 95 were H. pylori negative. The mean duration of storage of the stool samples was 120 days (range, 40-225 days). The sensitivity, specificity and positive and negative predictive values were 84.3%, 97.9%, 97.2% and 88.6%, respectively. In patients with a false negative HpSA test, there was a significant delay in collecting the stool specimen after endoscopy when compared with those with a true positive HpSA test (4.2 vs. 2.3 days; P < 0.05). However, the duration of storage of the specimens was not longer, and consumption of coffee or tea and smoking habits were similar. CONCLUSIONS: HpSA testing showed good sensitivity and specificity, even with frozen stool samples stored for up to 225 days. The efficacy was not affected by coffee, tea or smoking.

Adult↗