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[Comparison of positivity between density gradient centrifugal method and whole blood method in lymphocyte surface marker analysis with flow-cytometry].

The lymphocyte surface marker analysis with flow-cytometry is usually performed using a density gradient centrifugal method or a whole blood method. We have compared these two methods using several monoclonal antibodies and have found the significant difference in positivity all monoclonal antibodies except TCR-1 (anti alpha/beta receptor on T-lymphocyte). We found the significantly higher positivity of Leu12 and HLA-DR as well as lower of the other monoclonal antibodies by using whole blood method than by density gradient centrifugal method. We conclude that it is necessary to make a distinct description of the method used in the surface marker analysis.

Adult↗

Determination of sulfur dioxide in grapes: comparison of the Monier-Williams method and two ion exclusion chromatographic methods.

Results for determination of sulfur dioxide in grapes were compared by 3 methods: the modified Monier-Williams method, acid distillation/ion exclusion chromatography with electrochemical detection (AD/IEC-EC), and alkali extraction/ion exclusion chromatography with electrochemical detection (AE/IEC-EC). An unusual positive response was observed during the later stage of the Monier-Williams distillation of both control grapes and sulfited grapes. Development of volatile acidic compounds in parallel with this Monier-Williams response and darkening of sample was also observed by collection in an alkali trap and analysis using anion exclusion chromatography and photodiode array detection. No parallel increase in sulfite was observed by the more selective AD/IEC-EC method, which clearly demonstrated that the response observed during the later stage of the Monier-Williams method is a false positive, probably due to caramelization reaction products. Monier-Williams results for grapes containing ca 10 ppm sulfite were in reasonably good agreement with those by either the AD/IEC-EC or AE/IEC-EC methods, presumably because the false positive response in the Monier-Williams analysis compensated for the somewhat incomplete recovery of sulfite. The AE/IEC-EC method is recommended because it is rapid, sensitive, straightforward, and free from interference. Accurate results by Monier-Williams analysis could be obtained by limiting distillation to 60 min and correcting for recovery.

Chromatography, Liquid↗

Total and direct-reacting bilirubin values by automated methods compared with liquid chromatography and with manual methods for determining delta bilirubin.

This study compares total and direct-reacting bilirubin values in 40 serum samples from patients with various diagnoses, as measured by automated methods (Beckman Synchron CX-5, Beckman Astra 8, Kodak Ektachem 700) and HPLC and by a manual method for delta bilirubin. For total bilirubin, within-run CVs were less than 6%. The Ektachem 700 method underestimated bilirubin with serum samples from patients with Crigler-Najjar syndrome and from newborns in whom unconjugated bilirubin concentrations were increased but conjugated bilirubins were not present or were present only in small amounts. The Astra 8 and Synchron CX-5 methods were inaccurate with cholestatic serum samples, in which conjugated bilirubin concentrations were increased and other compounds such as bile acids could be expected to interfere. We conclude that each automated method examined provides reasonable estimates for total and direct-reacting bilirubin values for routine clinical use. The need for each laboratory to select the appropriate bilirubin method for its particular situation is obvious.

Autoanalysis↗

Methods of analysis for toxic elements in food products. 3. Limit of determination of methods for assuring safety.

To evaluate the suitability of the analytical methods used in determining food safety, a new metrological characteristic "MQS" is suggested. MQS is defined as the absolute minimum quantity in micrograms of a substance that can be determined in a test solution (solubilized test portion). MQS accounts for 2 factors: (a) the necessity for a reliable determination of ML (maximum permitted level, i.e., regulatory tolerance), and (b) the optimum quantity of test portion of a food product to be analyzed, and thus assists in evaluating the suitability of a method to assure food safety. The MQS of 8 toxic elements in any food are As, 3; Cd, 0.5; Cu, 20; Fe, 50; Hg, 0.2; Pb, 4; Sn, 200; Zn, 100 micrograms. To characterize the applicability of any given method, the specific minimum limit of determination, MQSM, must be established for each method. The method in question may be used to control food safety only if MQSM is less than MQS. MQSM values are given for the common polarographic and colorimetric methods for determining these elements.

Food Analysis↗

[Hemodynamic change in the liver following hepatic resection in the dog. A comparative study by the 198Au colloid method, KICG and the H2 clearance method].

On the basis of our hypothesis that blood flow in the residual hepatic parenchyma is a determinant factor of posthepatectomy liver function, measurements were made of blood flow in the residual liver after 40% hepatic resection in dog. Adult mongrel dogs were divided into two groups, i.e., normal control and 40% hepatectomy, and evaluated for hepatic hemodynamics by 198Au colloid method, KICG and H2 clearance method. Blood biochemistry and, histology of the liver were also examined. The results are as follows: In the control group, hepatic blood flow by 198Au colloid method, KICG and H2 clearance method was found to correlate significantly with each other. In the 40% hepatectomy group, the hepatic blood flow index by both 198Au colloid method and KICG decreased in proportion to the amount of resected hepatic tissue initially and then increased as the liver regenerated. H2 clearance method showed that hepatic blood flow per unit weight of tissue decreased in the face of reduced hepatic volume in early stage of posthepatectomy period and returned to the preoperative level as the liver regenerated.

Alanine Transaminase↗

[Assessment of bacterial counts in the meat-processing industry. II. The plate-loop method and the droplet method (author's transl)].

Two methods used in making bacterial counts of meat and surfaces are discussed. When the plate-loop method is used, dilutions are prepared with calibrated loops in spotting tiles. These loops are also used to swab droplets of these dilutions on quadrants of previously dried agar plates. A formula is presented, which may be used in calculating bacterial counts. When the droplet method is adopted, an apparatus is used to prepare dilutions in tubes containing liquid agar. Drops of agar then are transferred into empty Petri dishes by this apparatus. After incubation, the colonies which have grown in these droplets are counted by a viewer included in the apparatus. Both the plate-loop method and the droplet method (if slightly modified) were found to satisfy the criteria simplicity, reliability and saving time and material. This only holds good when accurate counts are made and when bacterial counts are more than 2,500 bacteria per gram or per sq. cm. When these conditions are complied with, the two methods are suited for routine studies of large numbers of samples.

Animals↗

Precision of glucose measurements in control sera by isotope dilution/mass spectrometry: proposed definitive method compared with a reference method.

This improved isotope-dilution gas chromatographic/mass spectrometric (GC/MS) method, in which [13C]glucose is the internal standard, meets the requirements of a Definitive Method. In a first study with five reconstituted lyophilized sera, a nested analysis of variance of GC/MS values indicated considerable among-vial variation. The CV for 32 measurements per serum ranged from 0.5 to 0.9%. However, concentration and uncertainty values (mmol/L per gram of serum) assigned to one serum by the NBS Definitive Method (7.56 +/- 0.28) were practically identical to those obtained with the proposed method (7.57 +/- 0.20). In the second study, we used twice more [13C]glucose diluent to assay four serum pools and two lyophilized sera. The CV ranged from 0.26 to 0.5% for the serum pools and from 0.28 to 0.59% for the lyophilized sera. In comparison, results by the hexokinase/glucose-6-phosphate dehydrogenase reference method agreed within acceptable limits with those by the Definitive Method but tended to be slightly higher (up to 3%) for lyophilized serum samples or slightly lower (up to 2.5%) for serum pools.

Blood Chemical Analysis↗

Comparison of modified automatic Dumas method and the traditional Kjeldahl method for nitrogen determination in infant food.

This study compares 2 methods for determining nitrogen and protein in various types of infant food: the Kjeldahl method, developed in 1883, which is time consuming and labor intensive, and a newer, automatic method, based on the Dumas method. In each category of infant food considered, the results obtained from both methods are shown to be comparable; however, the modified Dumas method is quicker, easier, and does not pollute the laboratory environment.

Autoanalysis↗

The Ames Clinitek 200/Multistix 9 urinalysis method compared with manual and microscopic methods.

We assayed 315 urine specimens by the Ames Clinitek 200/Multistix 9 semi-automated method and by corresponding standard methods (listed in parentheses) for the following analytes: protein (sulfosalicylic acid precipitation), glucose (Lilly Tes-Tape), ketones (Boehringer Mannheim Chemstrip K), leukocyte esterase (microscopic examination for leukocytes), blood (microscopic examination for erythrocytes), nitrite (microscopic examination for bacteria, Gram stain, or culture), and pH (pH meter). The Clinitek and standard methods agreed well at all concentrations for protein, ketones, and glucose. The Clinitek leukocyte esterase, nitrite, and blood methods were less sensitive than microscopic methods for detection of pyuria, bacteriuria, and hematuria, respectively. The Clinitek pH method produced falsely low pH values for urines with true pH less than 6.5, and falsely high values for urines with true pH greater than 6.5.

Autoanalysis↗

[A quantitative study of brain atrophy on computed tomography--multivariate analysis for comparison between the linear measurement method and the pixel count method].

We previously reported the newly developed quantitative measurement of the cerebral atrophy. The data indicated that the volume of the cerebrospinal fluid not always gradually increases during the life course but remains relatively constant until age 50 and thereafter increases with a wide variation. Though the technique, which is called the pixel count method, is highly quantitative, it is quite troublesome as it needs the computer to count out each pixels. On the other hand, the linear measurement method is easier than the pixel count method, but is far less quantitative. We examined seventy four subjects using both the linear measurement method and the pixel count method, and compared them by means of the multivariant analysis. Three different linear measurement methods were selected by the stepwise multiple regression analysis, those are B (distance between the caudate nuclei), E (greatest distance between the lateral ventricles at the level of the cella media) and G (number of visible sulci whose width are more than 3.1 mm at the level of 3 cm above the corpus callosum). B and E were calibrated by the maximum internal width of the skull (H). The highest correlation was achieved with a formula employing these parameters as follows; y = 42.66 X B/H + 12.52 X E/H + 0.232 X G - 2.92 (y means the estimated value of the CCR (CSF-cranial ratio), which is obtained by dividing the CSF volume by the cranial cavity). Multiple correlation coefficient was 0.76, and was statistically significant (p less than 0.001). The authors emphasized that using this formula we can easily predict CCR as the index of the brain atrophy without any computer.

Adult↗

Performance characteristics of methods of analysis used for regulatory purposes. I. Drug dosage forms. E. miscellaneous methods.

Precision parameters of miscellaneous methods for the analysis of drug dosage forms approved by AOAC since 1972, and not previously reviewed in this series, were recalculated on a consistent statistical basis by using the computer program FDACHEMIST. Seventeen published collaborative studies were reviewed; the studies encompassed 19 analytes in 80 different materials (dosage forms), 102 collaborative assays, approximately 10 laboratories per study, and principally direct spectrophotometric, polarographic, and spectroscopic methods, for a total of 1451 determinations. The average repeatability relative standard deviation (within-laboratories, RSDo) for the instrumental methods was 1.5%; the reproducibility relative standard deviation (among-laboratories, including within-, RSDx) was 2.6%; the ratio RSDo/RSDx of the averages was 0.57, with an average outlier rate of 2.7% of the reported determinations. The line of best fit of RSDx for the instrumental methods plotted against the negative logarithm of the concentration increases slightly with decreasing concentration, extending from an RSDx of approximately 2.0% at 100% concentration to an RSDx of 3.4% at 0.001% (10 ppm) concentration; this represents an RSDx change of approximately 0.3% (absolute) for each 10-fold decrease in concentration, independent of analyte, matrix, and method. A method for determining precipitated allergenic protein by the micro-Kjeldahl technique appeared to be outside this general relation, showing an RSDx of about 13% at a concentration of 0.015% (150 ppm) nitrogen.

Allergens↗

Mobility of the pubic symphysis. In vivo measurements with an electromechanic method and a roentgen stereophotogrammetric method.

Mobility of the pubic symphysis can be measured in vivo by two new methods. One is electromechanic (EM), with steel pins inserted into the bone. The other is roentgen stereophotogrammetric analysis (RSA), with tantalum bone markers implanted into the hip bones. With the first method, movements are directly registered with resistive transducers mounted on the pins. With the second method, movements are deduced with kinematic analysis from roentgen stereophotogrammetric measurements of the positions of the bone markers. Both methods have high precision, about 0.1 mm for translations and 0.1 degrees for rotations. The methods were used in two young volunteers, a man and a woman. In provocative tests with identical loading (alternating, active straight-leg raising in the supine position and standing on alternating left or right leg) there was a good correlation between the two methods. The measured values, i.e., translations of up to 2 mm and rotations of up to 3 degrees, form a basis for more comprehensive studies of normal and pathologic mobility of the pubic symphysis.

Adult↗

Lipoprotein quantification: an electrophoretic method compared with the Lipid Research Clinics method.

We compared a turbidimetric electrophoretic method (Lipidophor) for lipoprotein quantification with the standardized Lipid Research Clinics (LRC) method. In the Lipidophor procedure, major lipoproteins are separated by electrophoresis on agarose gels, precipitated on the gels with phosphotungstate-Mg2+ reagent, and the resulting turbidity is measured densitometrically. Measurements of relative turbidity were converted into lipoprotein cholesterol values by the use of numeric constants provided by the manufacturer. Among-day CVs (n = 46) for the Lipidophor method were 6.0%, 3.6%, and 9.9% for cholesterol in the alpha-, beta-, and pre-beta lipoproteins, respectively. The Lipidophor alpha-cholesterol was significantly lower (n = 171 specimens) than the LRC high-density lipoprotein (HDL) cholesterol (514 vs 586 mg/L), and beta-cholesterol was significantly higher than the corresponding LRC low-density lipoprotein (LDL) cholesterol values (1505 vs 1409 mg/L). The linear relation between the two methods for lipoprotein cholesterol quantification is as follows: Lipidophor alpha = 0.77 LRC HDL + 63 mg/L with correlation coefficient (r) of 0.87; Lipidophor beta = 0.95 LRC LDL + 166 mg/L (r = 0.96); Lipidophor pre-beta = 0.57 LRC very-low-density lipoprotein + 39 mg/L (r = 0.82). We derived a revised algorithm for estimating lipoprotein cholesterol from turbidity measurements. Lipoprotein cholesterol values by the Lipidophor method agree well with those obtained by the LRC method when these constants are used.

Adult↗

Comparison of automated method and improved AOAC Kjeldahl method for determination of protein in meat and meat products.

The Kjel-Foss automated method for protein determination meat and meat products was compared with the improved AOAC Kjeldahl method. Meat samples were separated into 3 categories based on fat content and analyzed in duplicate by both methods. No significant difference was found in a paired comparison of the 2 methods in each of the 3 meat categories, using Student's t-test at the 99% confidence level. A number of additional meat samples analyzed 6-9 times by the automated method showed an overall average range of 0.55% protein and average standard deviation of 0.20. The Kjel-Foss automated method was applicable for total protein determination in a wide variety of meat and meat products.

Animals↗

Comparative and multilaboratory studies of two immunodiffusion method enrichment protocols and the AOAC/Bacteriological Analytical Manual culture method for detection of Salmonella in all foods.

The single enrichment immunodiffusion (1-step), the preenrichment and selective enrichment immunodiffusion (2-step), and the AOAC/Bacteriological Analytical Manual culture methods for Salmonella were evaluated for equivalence in 2 separate studies, a comparative evaluation and a multilaboratory dilution study. In the comparative study, all 3 methods were performed on 10 food types. For 550 samples, analyses resulted in 99.3 and 99.6% agreement between the culture method and the 1-step and 2-step methods, respectively. False negative rates were 0.9 and 0.3% for 1-step and culture, and 0.0% and 0.6% for the 2-step and culture, respectively. Subsequently, 6 food types were included in a multilaboratory dilution-to-extinction study. A sequential dilution series of Salmonella in foods was analyzed by the 3 methods to determine their lower limits of detection for Salmonella. A total of 1185 samples analyzed resulted in 98.9% agreement between 1-step and culture, and 99.7% agreement between 2-step and culture. False negative rates were 1.8 and 0.1% for 1-step and culture, and 0.4 and 0.1% for 2-step and culture, respectively. During these evaluations, 1735 samples and controls representing 10 different naturally contaminated and inoculated foods were tested. The data indicate statistical equivalence of all 3 methods when analyzing all food types.

Bacteriological Techniques↗

Egg viability in urinary schistosomiasis. I. New methods compared with available methods.

A new method of quantifying viability and hatchability of Schistosoma haematobium eggs in urine has been developed. It consists of filtering urine samples through Nytrel filters followed by mercurochrome staining to assist in the detection of viable (motile) miracidia in egg shells, and iodine staining to show hatched miracidia. The new method is compared with two existing techniques for viability/hatchability determination, centrifugation hatching and Nuclepore filtration trypan blue staining. Seventy-three egg positive urine samples are compared; the overall proportions of viable eggs are very close for centrifugation hatching and the new method, but 33% of eggs are lost in centrifugation and 19% of eggs are unclassifiable by Nuclepore filtration trypan blue staining. The main advantages and disadvantages of the new and existing methods are described and discussed; the cheapness, simplicity and ease of performance of the new method make it suitable for large-scale field use. Suggestions for improvements and standardization of the new method are made.

Animals↗

Thoracoscopic surgery for lung cancer using the two small skin incisional method. Two windows method.

Pulmonary lobectomy and mediastinal lymph node dissection was performed in 25 patients with Stage I lung cancer under thoracoscopic guidance using the two-windows method. A posterior skin incision (3 cm) and lateral skin incision (2 cm) were made in the 4th intercostal space centering on the inferior angle of the scapula. The site closest to the operating surgeon was used for direct vision, while the distant site was used for insertion of the thoracoscope. The mean operative time was 2 hours and 15 minutes, and the mean blood loss was 82.6 ml. The mean number of dissected mediastinal lymph nodes was 32. The length of hospitalization ranged from 5 to 17 days. Recovery was uneventful, and analgesics were not required by postoperative day 6. The two-windows method overcomes the three-dimentional inaccuracy due to the one-directional observation of the operative field employed during conventional thoracoscopy. In addition, since we developed this method for mediastinal lymph node dissection, the tracheal bifurcation can be confirmed under direct vision, increasing the accuracy of the procedure. The advantages of the two-window thoracoscopic method of pulmonary lobectomy are cosmesis, preservation of respiratory function, and reduced postoperative pain. In addition, there is reduced intraoperative bleeding and shortened operative time, while achieving mediastinal dissection similar to that of standard thoracotomy. The two-windows method of thoracoscopic pulmonary lobectomy is equal or superior to standard thoracotomy in every respect. This method should become the standard surgical technique for Stage I lung cancer.

Adenocarcinoma↗

Problems in evaluating radiographic findings in rheumatoid arthritis using different methods of radiographic scoring: examples of difficult cases and a study design to develop an improved scoring method.

Radiographic scoring is generally accepted as the "gold standard" to follow the course of rheumatoid arthritis (RA). Several methods have been developed to quantify radiographic changes due to RA. The most widely used methods have been proposed by Larsen and Sharp. Problems of the interpretation of the radiograph and the design of the scoring methods are not resolved. The design of established scoring methods lacks correct correlation to the amount of change on radiographs, leading to confounding of the results of the radiographic analysis. Concerns arise about the correct interpretation of findings on plain radiographs that are caused by having a 2-dimensional picture of a 3 dimensional object. There is no agreement which of the different changes caused by RA on the radiograph have to be considered and which can be neglected. Examples for these problems are given. We compared the established scoring methods of Larsen and Sharp on the same set of radiographs with longterm followup using the correct data analysis to provide the information needed to design an improved scoring method. Details of the study design are reported.

Arthritis, Rheumatoid↗