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PubMed1982

Specimen handling and high-performance liquid chromatographic determination of furosemide.

A simple high-performance liquid chromatographic method to measure furosemide in plasma and urine is described. Furosemide fluoresces best, but is unstable, at acidic pH and is subject to photochemical degradation. These factors were analysed and the results prompted changes in previously described methods. All specimens were very carefully protected from light; extraction and acidification were done with acetic acid instead of hydrochloric acid. With these precautions no 4-chloro-5-sulphamoylanthranilic acid was found in biological specimens. The main metabolite was furosemide glucuronide (20% of furosemide excretion). Sensitivity was 0.1 and 0.5 microgram/ml for plasma and urine, respectively. The applicability of our method for furosemide studies is demonstrated.

Adult↗
PubMed1996

[Traumatic and iatrogenic ocular infections: specimen handling and role of the laboratory].

Ocular inflammations may be due to a variety of diseases, and microorganisms play a major role in both acute and chronic eye diseases. Treatment of serious infections needs a good microbiological diagnostic. The indications and techniques for investigations are determined by the site of infection, severity of the process, and knowledge of the likely responsible organisms. Immediate inoculation of specimens on culture media in the examining room is often recommended. This article describes organisms associated with ocular infections, and is designed to assists ophthalmologists in the collection, processing and transport of specimens.

Bacteriological Techniques↗
PubMed1981

Application of scanning electron microscopy to x-ray analysis of frozen-hydrated sections. I. Specimen handling techniques.

X-ray microanalysis of frozen-hydrated tissue sections permits direct quantitative analysis of diffusible elements in defined cellular compartments. Because the sections are hydrated, elemental concentrations can be defined as wet-weight mass fractions. Use of these techniques should also permit determination of water fraction in cellular compartments. Reliable preparative techniques provide flat, smooth, 0.5 micrometers-thick sections with little elemental and morphological disruption. The specimen support and transfer system described permits hydrated sections to be transferred to the scanning electron microscope cold stage for examination and analysis without contamination or water loss and without introduction of extraneous x-ray radiation.

Animals↗
PubMed1992

Impact of specimen handling and storage on detection of hepatitis C virus RNA.

Direct detection of hepatitis C virus (HCV) RNA in serum or plasma is useful for validating the performance of anti-HCV assays and for the discrimination of persons with persistent HCV infections from those with resolved infections. Quantitation of HCV RNA may also be useful for disease prognosis and therapeutic monitoring. Previous studies have reported detection of HCV RNA in 50 to 70 percent of blood donors who were positive on anti-HCV supplemental tests. There is concern that specimen processing and storage conditions might influence the stability, and hence the detectability, of HCV RNA. To address this concern, the rate of detection of HCV RNA by the polymerase chain reaction (PCR) using donor pilot tube sera (PTS) previously subjected to routine donor screening and supplemental testing was compared with HCV PCR results obtained with fresh-frozen plasma (FFP) derived from the same donations. All 16 anti-HCV supplemental test-positive donations evaluated were HCV RNA positive with FFP, whereas only 10 (62.5%) were positive with PTS (p = 0.024). None of 11 FFP or PTS samples from HCV enzyme immunoassay-reactive donations not confirmed by supplemental anti-HCV assays tested positive for HCV RNA. Direct comparison of sample type (serum vs. plasma) and various storage conditions using specimens from two seropositive donors showed that room-temperature storage results in marked reduction in HCV RNA signal, while replicate freezing and thawing caused a moderate reduction. These data indicate that well-controlled sample processing and storage conditions are critical to the sensitive and potentially quantitative analysis of HCV RNA.

Blood Preservation↗
PubMed2003

[Dientamoeba fragilis: is it really fragile? Approach to specimen handling and rapid microscopic diagnosis].

Dientamoeba fragilis is a pathogenic protozoan parasite with a world-wide distribution. Interestingly, a resistant cyst stage has not been demonstrated and it is still an unsolved problem how this parasite can survive successfully outside the human host. D. fragilis was found in 2% of approximately 2500 individuals unselected who submitted stools for parasitological examination during 2001 in Padua (Italy). The goal of this study was to detect the protozoan stages and the duration of persistence of this protozoa in faeces stored in different environmental conditions. The trophozoites of D. fragilis were detected up to 60 days after the collection of the faeces stored at 4 degrees C and Giemsa stained. The laboratory detection rate of the organism is greatly enhanced by use of preservative to fix stool specimens immediately after passage. Alternatively, a microscopic observation of the collected stool has to be performed immediately after passage followed by examination of permanently-stained smears. Demonstration of the charateristic "golf-club" and "acanthopodia-like" structures in unstained fixed faecal material by direct microscopy (400x) are suitable for a rapid identification of D. fragilis.

Animals↗
PubMed1981

Specimen handling for the assay of adenylates and glycerate 2, 3-bisphosphate in erythrocytes.

ATP, ADP, AMP and glycerate 2, 3-bisphosphate in blood are frequently analyzed in order to evaluate erythrocytes for blood donation and to diagnose hemolytic anemias. In order to find an uncomplicated and safe procedure for the preanalytical treatment of blood a freezing-storing procedure using dry ice was worked out and combined with a later PCA precipitation. The pre-freezing stability of the components was also investigated and found to be best at room temperature.

2,3-Diphosphoglycerate↗
PubMed1986

Specimen handling in an HPLC determination of phenylbutazone and its major metabolites in plasma, avoiding degradation of the compounds.

A problem usually not taken into account when a quantitative HPLC method for phenylbutazone is developed is the degradation of this drug and its metabolites not only upon storage, but also on extraction under acidic conditions, especially when the temperature is raised. Moreover, the degradation products in the chromatograms may interfere with the determination of gammahydroxyphenylbutazone. In our newly developed HPLC method, using feprazone as an internal standard, extreme care is taken to avoid degradation of the compounds during the extraction procedure. In view of the present results it is concluded that previously published data on phenylbutazone, oxyphenbutazone and gammahydroxyphenylbutazone levels should be considered with reserve.

Chromatography, High Pressure Liquid↗
PubMed2000

Different stability of free and complexed prostate-specific antigen in serum in relation to specimen handling and storage conditions.

The effect of sample collection, storage conditions (time and temperature), and freeze-thaw cycles on the stability of free prostate-specific antigen (fPSA), PSA complexed with alpha1-antichymotrypsin (ACT-PSA), and total PSA (tPSA) in serum was studied. The analytes were quantified using immunoassays for tPSA and fPSA on the Elecsys system 2010 and a research assay for ACT-PSA on the ES system (Roche Diagnostics). The stability of the analytes was calculated as percentages of the values measured in samples 1 h after blood collection. When the samples were stored at 37 degrees C, at room temperature or at 4 degrees C, the stability of ACT-PSA was less impaired than that of fPSA. To avoid erroneous results in the determination of PSA isoforms and their corresponding ratios, serum samples should be preserved at 4 degrees C when the analysis is performed within 8 h after blood collection, or they should be stored at -80 degrees C if the analysis is not feasible during that period.

Blood Chemical Analysis↗
PubMed1994

[Effect of the time of sample collection after exertion and the specimen handling on the lactate content in the blood of horses].

The mean maximal lactate concentration of horses after exercise (Lamax) amounted to 4.5 +/- 2.2 mmol/l for gallop workouts of less than 11 m/s and more than 180 s (n = 35; category A), at more than 11 m/s and less than 180 s duration (n = 17; category B) to 16.3 +/- 4.8 mmol/l and at less than 11 m/s and less than 180 s duration (n = 10; category C) to 4.6 +/- 1.6 mmol/l. Lamax was always measured immediately after exercise following exercise of categories A and C. But for exercise bouts of category B Lamax was measured between the first and the tenth minute after exercise. Following exercise bouts in this category, mean lactate concentrations measured in two-minute intervals before and after Lamax always differed significantly (p < 0.01 to p < 0.001). The lactate concentration did not vary within seven days in whole blood samples deproteinized immediately after collection and in plasma samples. In the same sample material, when kept frozen, lactate contents remained stable for 60 days. In whole blood samples treated with Na-heparin or Na-heparin/Na-fluoride stored at 20-22 degrees C and 4 degrees C, the lactate concentration had already increased significantly within the first 24 hours.

Animals↗
PubMed1998

Stability of hepatitis C virus RNA during specimen handling and storage prior to NASBA amplification.

The influence of different anticoagulants and pre-amplification storage conditions on the stability of hepatitis C virus (HCV)-RNA, as detected by the quantitative HCV NASBA assay (NASBA-QT), was studied. The HCV-RNA load remained stable for at least 15 months when serum or plasma samples (EDTA and heparin) were directly frozen at -70 degrees C in lysis buffer. At 4 degrees C, the HCV-RNA load in serum or plasma stored with lysis buffer did not decline for at least 14 days. At 30 degrees C, however, the load declined significantly after 7 days. When clotted, whole blood was stored at 4 degrees C, the HCV-RNA load was stable for 72 h. However, when EDTA-anticoagulated whole blood was stored at 4 degrees C, the HCV-RNA load declined significantly after 48 h. In paired plasma and serum samples at baseline the HCV-RNA levels were similar. Heparin did not influence the efficiency of the HCV NASBA-QT assay. Clotted blood as well as EDTA or heparin anticoagulated blood can be used for quantifying HCV-RNA using the NASBA-QT assay. Blood samples should be stored at 4 degrees C after collection and serum or plasma separated within 24 h. Preferably, after separation, samples should be frozen in lysis buffer at -70 degrees C until NASBA-QT analysis.

Buffers↗
PubMed1998

Optimization of specimen-handling procedures for accurate quantitation of levels of human immunodeficiency virus RNA in plasma by reverse transcriptase PCR.

Human immunodeficiency virus type 1 (HIV-1) RNA levels in plasma are currently widely used clinically for prognostication and in monitoring antiretroviral therapy. Accurate and reproducible results are critical for patient management. To determine the effects of specimen collection and handling procedures on quantitative measurement of HIV-1 RNA, we compared anticoagulants and sample processing times. Whole blood was collected from 20 HIV-1-infected patients in EDTA, acid citrate dextrose (ACD), and heparin tubes, aliquoted, and stored at room temperature. Plasma was separated from whole-blood aliquots prepared at < or =1, 3, 6, 24, and 48 h postcollection and then stored at -70 degrees C until use. HIV-1 RNA levels were determined by the AMPLICOR HIV-1 MONITOR assay. Heparinized plasma samples, which were pretreated with heparinase prior to analysis, had the lowest baseline HIV-1 RNA levels. In the first 6 h, HIV-1 RNA levels decreased by 10, 20, and 31% in EDTA, ACD, and heparin tubes, respectively. From 6 to 48 h postcollection, HIV-1 RNA levels decreased in all anticoagulants, albeit at a slower, more consistent rate. Our results indicate that EDTA should be the anticoagulant of choice for plasma HIV-1 RNA measurement by reverse transcriptase PCR, but ACD tubes are acceptable if the plasma is separated within 6 h of blood collection. Caution must be applied in the interpretation of absolute HIV-1 RNA copy number values obtained with suboptimal specimen collection and processing procedures.

Adolescent↗
PubMed1976

Error introduced by specimen handling before determination of inorganic phosphate concentrations in plasma and serum.

Values for serum inorganic phosphate (Pi) concentrations in groups of healthy adults vary widely, the coefficient of variation ranging from 10 to 15%. We undertook to determine in 23 healthy adults whether part of this variation could be accounted for by (a) drawing blood in syringes vs. evacuated tubes (b) the time between blood sampling and separation of serum or plasma, and (c) the prevention of clotting. Values were unaffected by a, decreased significantly with time at room temperature between blood sampling and separation of cells in both serum and plasma, and were significantly lower in plasma than in serum. The group coefficient of variation for Pi averaged 13% and was uninfluenced by the blood-processing technique.

Blood Specimen Collection↗