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Results for “Absolute quantification”

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PubMed2000

Validity and reproducibility of echocardiographic measurement of left ventricular ejection fraction by acoustic quantification with tissue harmonic imaging technique.

The tissue harmonic imaging technique can enhance detection of the cardiac endocardial border. When combined with an acoustic quantification (AQ) method, an improvement of accuracy and reproducibility of real-time measurement of left ventricular (LV) function might be expected. However, few data exist regarding the measurement of LV function by AQ with the harmonic imaging technique. Therefore, we evaluated the validity and reproducibility of AQ measurement of LV ejection fraction with or without harmonic imaging technique. A total of 50 patients (mean age 58 +/- 10 years) who underwent left ventriculography were included in our study. The LV end-diastolic and end-systolic volumes by ventriculography were 131 +/- 52 mL and 72 +/- 43 mL, respectively, and were underestimated by both conventional (70 +/- 32 mL and 36 +/- 25 mL) and harmonic (67 +/- 30 mL and 34 +/- 22 mL) AQ obtained in the apical 4-chamber view. The calculated ejection fraction by ventriculography was 0.49 +/- 0. 11 and correlated with that by conventional AQ (0.51 +/- 0.11; y = 0. 72x + 0.152; r = 0.73). This was a marked improvement when compared with the ejection fraction by harmonic AQ (0.50 +/- 0.11; y = 0.89x + 0.065; r = 0.91). Interestingly, interobserver and intraobserver variabilities of conventional AQ, which were 15.6% and 8.6%, respectively, were much improved by harmonic AQ (8.9% and 4.5%, respectively). These results indicate the feasibility of real-time measurement of LV ejection fraction by harmonic imaging, although absolute LV volume can be underestimated even by this technique.

Aged↗

PubMed1998

Impact of presence of abnormal wall motion on echocardiographic determination of left ventricular function with automated boundary detection technique: re-evaluation.

It is still unclear whether echocardiography with an automated boundary detection technique (ABD) can accurately determine the left ventricular (LV) volume and function particularly in the presence of LV wall asynergy. We intended to re-evaluate the reliability and application of the ABD, which was based on the acoustic quantification technique (Sonos 2500, Hewlett Packard) for the LV volume measurement in patients without or with LV wall asynergy. A total of 80 patients (mean age 56 years) who underwent left ventriculography (LVG) were divided into two groups. The group A consisted of 29 patients with normal LV wall motion and the group B consisted of 51 patients with generalized or regional LV wall motion abnormality. In group A patients, the LV end-diastolic volume (LVEDV) was 96 +/- 25 ml by ABD and 112 +/- 33 ml by LVG and those of LV end-systolic volume (LVESV) were 44 +/- 14 ml by ABD and 48 +/- 17 ml by LVG, thus resulting in the underestimation of LV volume by 12% in average. Under these conditions, the LV ejection fraction (LVEF) by ABD, 54 +/- 8%, correlated well with that by LVG, 58 +/- 7%. Although underestimation of LV volume by 17% in average also occurred in groups B (N.S.), LVEF was found to correlate well with that by LVG; 27 +/- 8% vs 30 +/- 11% (r = 0.87, SEE = 3.1%) for 21 patients with the generalized LV asynergy; 39 +/- 10% vs 39 +/- 12% (r = 0.86. SEE = 3.3%) for 30 patients with the regional LV asynergy. These results demonstrate the feasibility of the ABD in determining the LVEF, although underestimation can occur in measuring the absolute LV volume in patients with or without LV asynergy.

Case-Control Studies↗

PubMed1996

Noninvasive quantification of iodine-123-iomazenil SPECT.

UNLABELLED: The feasibility of a noninvasive method for quantification of [123I]iomazenil binding using a standardized arterial input function and a single venous blood sample was assessed in normal volunteers. METHODS: Serial SPECT images and blood data from six healthy male volunteers after intravenous injection of [123I]iomazenil were used. The standardized input function was derived by averaging the six subjects' arterial curves. Individual input functions were estimated by calibrating the standardized input function with one-point venous blood radioactivity concentration. Ligand transport (K1) and receptor binding were computed from the estimated input functions and two separate SPECT scans using a table look-up procedure based on a three-compartment, two-parameter model. Reference values for K1 and receptor binding were determined from the serial SPECT data and individual arterial curves using a three-compartment, three-parameter model and curve fitting. RESULTS: Analyses of the error caused by the calibration in relation to the time postinjection revealed that the optimal calibration time was 30 min postinjection. Receptor binding obtained by this simplified method correlated well with the reference values (r = 0.941) and was estimated within an error of 10% in the cerebral cortical regions. Although the estimated K1 showed relatively poor correlation (r = 0.699) with the reference value, it was an excellent relative measure in each subject. CONCLUSION: Our method provided an absolute measure of the benzodiazepine receptor binding and a relative measure of ligand transport from two SPECT scans and a venous blood sample. This method would be useful for quantitative assessment of benzodiazepine receptors in clinical settings.

Adult↗

PubMed1993

Ultrasonic backscatter system for automated on-line endocardial boundary detection: evaluation by ultrafast computed tomography.

OBJECTIVES: The purpose of this study was to evaluate the accuracy of the recently developed echocardiographic on-line endocardial border detection system using ultrafast computed tomography, an independent and proved tomographic imaging modality. BACKGROUND: The automated system for on-line endocardial border detection identifies the blood-tissue interface by acoustic quantification of the ultrasonic backscatter signal. METHODS: Eighteen subjects were screened by conventional echocardiography and acoustic quantification. Ten of these, with high quality echocardiographic images, were also examined by ultrafast computed tomography. Comparable image planes at the midpapillary level were analyzed. Measurements of left ventricular cavity area were compared at end-diastole and end-systole and time course analyses of cavity area during the cardiac cycle were performed. RESULTS: There was good correlation between values for left ventricular end-diastolic area (r = 0.99), end-systolic area (r = 0.93) and fractional area change (r = 0.91) using the two methods. The on-line backscatter system underestimated end-diastolic area (p < 0.001), but the negative bias was small (-1.6 cm2) and the 95% confidence intervals were narrow (-3.6 cm2 to +0.4 cm2). In contrast, the backscatter system overestimated end-systolic area (p < 0.02); the positive bias for this variable was also small (+2.6 cm2) but the confidence intervals were relatively wide (+7.9 to -2.8 cm2). The negative bias of backscatter values for cavity area was fairly constant during diastole and early systole (range -5% to -10%), but during the second half of systole, backscatter values increased progressively relative to computed tomographic values. Real time values for fractional area change measured by the backscatter system were 13% smaller than those determined by ultrafast computed tomography (p < 0.001), with wide confidence intervals (+3% to -30%). Absolute peak rates of area change during systole and diastole were lower by 39% (p < 0.001) and 41% (p < 0.01), respectively, using the on-line ultrasonic backscatter system. Time course analyses revealed the errors to be consistent with cardiac cycle-dependent alterations in gain sensitivity of the ultrasonic backscatter system. CONCLUSIONS: The ultrasonic backscatter system is associated with cyclic cavity area measurement errors that need to be addressed if its early promise for on-line assessment of ventricular function is to be fulfilled. Incorporation of an electrocardiographically triggered time-varying gain control may improve accuracy for on-line analysis of ventricular performance.

Echocardiography↗

PubMed1993

Body composition, health status and urinary magnesium excretion among elderly people (Dutch Nutrition Surveillance System).

Magnesium (Mg) requirements depend on body weight. Recommended dietary allowances (RDA) are mostly given in absolute amounts and are mostly similar for adults, despite differences in body composition in the adult age range. We therefore studied the interrelations of 24 h urinary Mg excretion with body build and body composition indicators, as well as with factors potentially affecting the metabolism of Mg, among 520 elderly men and women seen in a nationwide survey. Mg intake data were also obtained in the Dutch Food Composition Survey amount 5898 subjects. The results showed lower mean Mg intake among elderly people (especially men) and positive associations of body weight and body height with urinary Mg excretion, but not when expressed per mmol of creatinine excreted. Mg excretion per mmol of creatinine was not associated with body weight, body height, body mass index or body fatness (women), suggesting that the amount of fat mass did not affect Mg excretion. Mg excretion was positively correlated with Mg intake, creatinine clearance, excretion of sodium, potassium and calcium, and coffee consumption, indicating interactions at the kidney level that may increase Mg losses. Elderly people using diuretics and/or anticholinergics and male diabetics had a higher mean Mg excretion per mmol of creatinine. The results show that, besides the Mg intake, several factors affect the urinary Mg excretion among elderly people. These factors were body build and body composition, dietary intake, drug use, kidney function, and diabetes. A quantification of the effects cannot be made yet, and controlled studies on those factors potentially affecting the requirement of Mg are needed.

Aged↗

PubMed1990

Minocycline-treatment of diabetic rats normalizes skin collagen production and mass: possible causative mechanisms.

Daily minocycline-treatment of streptozotocin-induced diabetic rats not only prevented a diabetes-caused atrophy of skin collagen mass (10-mos old rats), but also normalized skin collagen mass to match that of growing (ca. 1%/d) non-diabetic controls (4- and 5-mos old rats). The causative mechanism by which minocycline-treatment normalizes skin collagen mass must, in part, be related to a general anabolic effect on growth (body weight) because the effect on skin collagen mass correlates strongly to that on body weight. Consequently, a minocycline-stimulated increase of a systemic factor (such as insulin-like growth factor) is not unlikely. The anabolic effect of minocycline-treatment of diabetic rats is also expressed as a normalized cellular ribosome mass (an index of total protein synthetic capacity) and a normalized absolute rate of collagen production. (Calculation of an absolute rate was justified by an apparent maximum saturation of the prolyl-tRNA pool(s) of skin, maximum saturation obtained by the pool-flooding approach). The normalized skin ribosome amount does not, however, explain a selective effect of minocycline-treatment on collagen production as opposed to that for non-collagen protein, this selective effect measured as relative collagen production. To explain such selectivity, the inhibition of diabetes-induced excess skin collagenase activity seems unlikely. (This inference is based on results from a preliminary study indicating that recently [less than 2 h] synthesized collagen is not degraded by the excess collagenase in skin of diabetic rats). Thus, the principal collagen fraction acted on by pathologically excess collagenase might be collagen at a later stage (greater than 2 h after synthesis) in its life cycle. (Another possibility for the selective effect of minocycline on collagen production, as yet untested, is reduced intracellular procollagen degradation.) Overall, this is the first study aimed at discerning the mechanism(s) by which minocycline-treatment enhances the rate of collagen production in tissues of a diabetic rat. For future studies, the extent to which the positive effect on growth, ribosome mass, and rate of collagen production contributes to the change of collagen mass must, along with the known minocycline-inhibition of collagenase activity, be quantified. Such quantification is a prerequisite for evaluating the chemotherapeutic efficacy of minocycline-treatment on collagen-degradative diseases.

Animals↗

PubMed2005

Theoretical accuracy of model-based shape matching for measuring natural knee kinematics with single-plane fluoroscopy.

Quantification of knee motion under dynamic, in vivo loaded conditions is necessary to understand how knee kinematics influence joint injury, disease, and rehabilitation. Though recent studies have measured three-dimensional knee kinematics by matching geometric bone models to single-plane fluoroscopic images, factors limiting the accuracy of this approach have not been thoroughly investigated. This study used a three-step computational approach to evaluate theoretical accuracy limitations due to the shape matching process alone. First, cortical bone models of the femur tibia/fibula, and patella were created from CT data. Next, synthetic (i.e., computer generated) fluoroscopic images were created by ray tracing the bone models in known poses. Finally, an automated matching algorithm utilizing edge detection methods was developed to align flat-shaded bone models to the synthetic images. Accuracy of the recovered pose parameters was assessed in terms of measurement bias and precision. Under these ideal conditions where other sources of error were eliminated, tibiofemoral poses were within 2 mm for sagittal plane translations and 1.5 deg for all rotations while patellofemoral poses were within 2 mm and 3 deg. However, statistically significant bias was found in most relative pose parameters. Bias disappeared and precision improved by a factor of two when the synthetic images were regenerated using flat shading (i.e., sharp bone edges) instead of ray tracing (i.e., attenuated bone edges). Analysis of absolute pose parameter errors revealed that the automated matching algorithm systematically pushed the flat-shaded bone models too far into the image plane to match the attenuated edges of the synthetic ray-traced images. These results suggest that biased edge detection is the primary factor limiting the theoretical accuracy of this single-plane shape matching procedure.

Algorithms↗

PubMed1993

Contrast-enhanced first pass myocardial perfusion imaging: correlation between myocardial blood flow in dogs at rest and during hyperemia.

The sensitivity of contrast-enhanced MR first pass perfusion imaging in detection and quantification of hypoperfused myocardium was evaluated using an instrumented, closed-chest dog model where graded regional hypoperfusion was induced by applying predetermined levels of stenosis to the left anterior descending artery (LAD). All measurements were performed at rest and under stress induced by dipyridamole (DIP). Myocardial perfusion was assessed both with MR and radiolabeled microspheres injected immediately before the administration of the MR contrast agent. Ultrafast MR imaging was performed using a Turbo FLASH sequence with a 180 degrees inversion prepulse. A Gd-DTPA bolus was injected into the left atrium and T1-weighted images were acquired with every heart beat. Signal intensity measured from the images in regions of the LAD and left circumflex (LCx) perfusion beds was plotted against time to generate signal intensity versus time curves (SI time curve). Various flow indices were derived according to the indicator dilution theory, and compared with and without volume correction due to vasodilation to the myocardial blood flow (MBF) calculated from radiolabeled microspheres. Correlation of the MR and MBF data demonstrated that different transmural and regional myocardial perfusion levels can be easily visualized in the perfusion images and accurately monitored by the SI time curves. Detection of the impairment of myocardial perfusion improved significantly after administration of DIP. The inverse mean transit time calculated from the SI time curve was found to yield a linear correlation to absolute MBF derived from the microsphere data. These results suggest that with intracardiac injections of exogenous contrast agent, myocardial perfusion can be assessed parametrically with first pass contrast enhanced ultrafast MRI.

Animals↗

PubMed2000

Molecular quantification of cell cycle-related gene expression at the protein level.

BACKGROUND: Immunofluorescence cytometry of antigen and DNA content provides relative measurements of the cell cycle phase distribution of a specific epitope. Measurement of correlated expression of epitopes on signaling and regulatory proteins will be useful in the study of the complex pathways involved in cell cycle regulation and carcinogenesis. However, to formulate regulatory pathway models, measurements of molecules per cell would be more useful than relative measurements of intensity. Here, we report on a system in which the relationship between molecules and fluorescence is determined for a reference set of cell lines that are then used to directly calculate the number of molecules for unknowns. To demonstrate the process, we calculated the cell cycle phase distribution of SV40 large T antigen (Tag) in the reference cells. METHODS: A set of cell line clones expressing different levels of Tag were isolated. Quantitative Western blots of these cells and purified, recombinant Tag were performed. Cells from the same sample were stained and analyzed by flow cytometry for Tag and DNA. The relationship between molecules and fluorescence was established and calculations were performed for the phase distributions of Tag. RESULTS: The five cell lines had 0.11, 0.27, 1.06, 2.44, and 2.63 x 10(6) molecules of Tag per cell, determined by Western blot. The average coefficient of variation was 10.6%. The relationship of molecules to fluorescence fit a linear equation (r(2) = 0.96) over the range, 0.11 - 2.63 x 10(6) molecules, however, the same equation did not fit the relationship between 0 molecules, defined by isotype staining controls, and the lowest expressing cell line. To calculate the phase distributions of molecules in the lowest cell line, a second linear equation from 0 to 110,000 molecules was used. CONCLUSIONS: This work describes a system where fixed cells expressing various levels of a target antigen quantified by Western blots can be used to standardize flow cytometric measurements of gene expression in absolute terms.

Animals↗

PubMed1999

Relative efficacy of 32P and 89Sr in palliation in skeletal metastases.

UNLABELLED: 32p and 89Sr have been shown to produce significant pain relief in patients with skeletal metastases from advanced cancer. Clinically significant pancytopenia has not been reported in doses up to 12 mCi (444 MBq) of either radionuclide. To date, no reports comparing the relative efficacy and toxicity of the two radionuclides in comparable patient populations have been available. Although a cure has not been reported, both treatments have achieved substantial pain relief. However, several studies have used semiquantitative measures such as "slight," "fair," "partial" and "dramatic" responses, which lend themselves to subjective bias. This report examines the responses to treatment with 32P or 89Sr by attempting a quantification of pain relief and quality of life using the patients as their own controls and compares toxicity in terms of hematological parameters. METHODS: Thirty-one patients with skeletal metastases were treated for pain relief with either 32P (16 patients) or 89Sr (15 patients). Inclusion criteria were pain from bone scan-positive sites above a subjective score of 5 of 10 despite analgesic therapy with narcotic or non-narcotic medication, limitation of movement related to the performance of routine daily activity and a predicted life expectancy of at least 4 mo. The patients had not had chemotherapy or radiotherapy during the previous 6 wk and had normal serum creatinine, white cell and platelet counts. 32P was given orally as a 12 mCi dose, and 89Sr was given intravenously as a 4 mCi (148 MBq) dose. The patients were monitored for 4 mo. RESULTS: Complete absence of pain was seen in 7 of 16 patients who were given 32P and in 7 of 15 patients who were given 89Sr. Pain scores fell by at least 50% of the pretreatment score in 14 of 16 patients who were given 32P and 14 of 15 patients who were given 89Sr. Mean duration of pain relief was 9.6 wk with 32P and 10 wk with 89Sr. Analgesic scores fell along with the drop in pain scores. A fall in total white cell, absolute granulocyte and platelet counts occurred in all patients. Subnormal values of white cells and platelets were seen in 5 and 7 patients, respectively, with 32P, and in 0 and 4 patients, respectively, after 89Sr therapy. The decrease in platelet count (but not absolute granulocyte count) was statistically significant when 32P patients were compared with 89Sr patients. However, in no instance did the fall in blood counts require treatment. Absolute granulocyte counts did not fall below 1000 in any patient. There was no significant difference between the two treatments in terms of either efficacy or toxicity. CONCLUSION: No justification has been found in this study for the recommendation of 89Sr over the considerably less expensive oral 32P for the palliation of skeletal pain from metastases of advanced cancer.

Administration, Oral↗

PubMed2006

Quantification and visualization of three-dimensional inconsistency of the globus pallidus internus in the Schaltenbrand-Wahren brain atlas.

The major shortcomings of the Schaltenbrand-Wahren (SW) brain atlas include 3-dimensional (3D) inconsistency and spatial sparseness. This work quantifies and visualizes 3D inconsistency of the globus pallidus internus (GPi), a stereotactic target for the treatment of Parkinson's disease, dystonia and Huntington disease. The GPi 3D models 3D-A, 3D-C and 3D-S are reconstructed from the SW axial, coronal and sagittal microseries, respectively, by applying a shape-based (Nonuniform Rational B Splines) method. All three 3D models, placed in the SW coordinate system, are compared quantitatively in terms of location (centroids), size (volumes), shape (normalized eigen values), orientation (eigen vectors) and mutual spatial relationships (overlaps and inclusions). The analysis is done in 3D within each orientation and across them. The reconstructed 3D GPi models substantially differ in location, size and inclusion rate. The centroid of 3D-C is located more medially (15.6 mm) than those of 3D-A (17.5 mm) and 3D-S (18.2 mm), and that of 3D-A more ventrally (-2.3 mm) than those of 3D-C (-0.1 mm) and 3D-S (-0.4 mm). 3D-S has the smallest volume (347.3 mm3); 3D-A is 1.18 and 3D-C 1.85 times larger. The highest inclusion rate is for 3D-S (54.3 and 56.3%) and the lowest for 3D-C (28.8 and 30.6%). A smaller variability is observed in shape, orientation and overlap size (196.8, 196.1 and 185.5 mm3). To get a better correspondence between 3D-C and 3D-S, the coronal microseries were scaled laterally by 1.1667. This results in a substantial improvement of the inclusion rate of 3D-S (87.9%), though raising the volume mismatch to 2.16. The GPi in the SW atlas has a substantial 3D inaccuracy within each orientation and across them. Therefore, absolute and direct reliance on the original atlas is unsafe, and this atlas has to be used with great care and understanding of its limitations. As matching various SW microseries by global scaling is not feasible, we propose the target-dependent scaling based on structure centroid matching.

Brain↗

PubMed1981

Radioimmunoassay and pharmacokinetic profile of bupropion in the dog.

A radioimmunoassay (RIA) procedure for the quantification of bupropion (dl-2-tert-butylamino-3'-chloropropiophenone) in biological fluids is described. Immunization of rabbits with conjugates of bovine serum albumin and p-succinoyl propylbupropion or p-carbomethoxybupropion resulted in the production of antisera which are capable of detecting less than 1 ng ml-1 (100 pg actual mass) of bupropion in the RIA, utilizing [6-3H] bupropion as radioligand. The antisera used in these studies have low cross-reaction (approximately 0.1% or less) with known side chain metabolites of bupropion, but exhibit significant cross-reaction with p-hydroxybupropion (30.3%). Excellent agreement was obtained between RIA and high-pressure liquid chromatography determinations of bupropion concentrations in human plasma samples, but plasma or serum from bupropion-treated dogs, rats and mice required extraction from basic medium to remove some interference before RIA. The assay was applied to a study of bupropion disposition in two beagles of each sex after i.v. and p.o. administrations of bupropion hydrochloride (100 mg). The pharmacokinetic profile in dogs was best described by an open two-compartment model after either route of drug administration. Peak plasma bupropion levels after oral dosing were highly variable, ranging from 12.9 to 63.5 ng ml-1 at 26 to 32 min after drug administration. The mean terminal phase half-life of bupropion was calculated to be 1.73 hr after either route and the absolute oral bioavailability of the drug varied from 2.0 to 6.5%.

Animals↗

PubMedDate not supplied

Quantification of in vivo anaerobic metabolism in the normal cat retina through intraretinal pH measurements.

We examined intraretinal [H+] in the intact retina of anesthetized cats using H+-sensitive microelectrodes to obtain spatial profiles of extracellular [H+]. One H+ is produced when an anaerobically generated ATP is utilized. We theorized that H+ production directly reflects anaerobic glucose consumption. From the choroid (pH approximately 7.40), [H+]o steadily increased to a maximum concentration in the proximal portion of the outer nuclear layer (pH approximately 7.20). The shape of the profile was always concave down, indicating that a net production of H+ occurred across the avascular outer retina. A three-layer diffusion model of the outer retina was developed and fitted to the data to quantify photoreceptor H+ extrusion into the extracellular space (Q(OR-H+)). It was determined that the outer segment (OS) layer had negligible H+ extrusion. The data were then refitted to a special three-layer model in which the OS layer Q(H+) was set equal to zero, but in which the inner segments and outer nuclear layer produced H+. The resulting Q(OR-H+) was several orders of magnitude lower than previous measurements of Q(OR-lactate), which were based on choroidal mass balances of lactate. Stoichiometrically, one H+ is produced for each lactate produced, so we concluded that Q(OR-H+) is a measure of net rather than total H+ production. Because retinal acid production is so high, the retina must contain efficient H+ clearance and/or neutralization mechanisms that prevent severe acidosis. The effect of light on retinal extracellular [H+] and Q(OR-H+) was also examined. As expected, light adaptation caused a retinal alkalinization that resulted from a 52% reduction in Q(OR-H+). This is in agreement with previous studies that have shown that both oxidative (e.g. Haugh et al., 1990) and glycolytic metabolism (Wang et al., 1997a,c) in the photoreceptor are decreased by a factor of 2 during light adaptation. Although we could not obtain absolute values for outer retinal glycolysis, changes in Q(OR-H+) appear to directly reflect changes in glycolytic metabolism.

Adaptation, Ocular↗

PubMed1997

[Leptin in persons with simple obesity].

Obesity is a disease with distinct genetic determination and its phenotype is defined by the still unknown number of genes whose expression can be influenced by environmental factors. Several years ago, "obesity gene" was isolated in animals. This gene, coding protein which consists of 165 amino acids, is called leptin. Leptin is supposed to be a key substance controlling homeostasis of body weight and energy balance; it is produced by adipocytes and its value correlates highly significantly with anthropometric parameters that characterize physical constitution and amount of subcutaneous fatty tissue. The obese individuals often display hyperleptinemia which is frequently caused by a postreceptor disorder; sporadically, a different leptin structure or hypoleptinemia (caused by genetic anomaly) are reported. It is supposed that either absolute or relative leptin deficiency in obese persons are associated with causal obesity (e.g. appetite stimulation). Leptinemia values correlate with percentage of subcutaneous fatty tissue, insulinemia and sometimes with glycemia. In our study we examined 200 probands, patients of the Metabolic and Diabetologic Out-Patient Department, Hospital in Sternberk. A very close correlation between the amount of subcutaneous fatty tissue (measured by a caliper in 10 skinfolds) and the leptine serum concentration was found. The values of leptinemia in men of normal constitution ranged within 1-11 ng/ml, non-obese women had 3-4 times higher values. Leptinemia in some obese individuals reached up to 70 ng/ml. However, the currently calculated and reported parameters of physical constitution (BMI, WHR, Grant index) did not correlate significantly with leptinemia. Similarly, biochemical parameters considered as general markers of insulin resistance (often associated with obesity) did not correlate significantly with leptinemia. This finding indicates that some calculated parameters, quantifying and gualifying physical constitution, may be ambiguous and leptinemia was found to give more detailed information about the amount of subcutaneous fatty tissue than WHR or BMI. An accidental finding was an important positive correlation between myoglobin concentration and creatinemia. At monitoring the effect of hypolipidemic agents we use the myoglobin examination and therefore we consider this correlation to be very important and every physician performing this analysis should be informed about it. The present study thus confirmed that a more accurate quantification of subcutaneous fatty tissue is required. On the other hand, we believe that examination of leptinemia can contribute significantly to stratification of patients into risk groups (with respect to clinical, economic and time differentiation) and subsequently to the treatment of these patients. In future, criteria for quantification of leptinemia and leptine resistance should be defined precisely.

Adipose Tissue↗

PubMed2002

Quantitative assay for measuring the Taura syndrome virus and yellow head virus load in shrimp by real-time RT-PCR using SYBR Green chemistry.

Taura syndrome virus (TSV) and yellow head virus (YHV) are the two RNA viruses infecting penaeid shrimp (Penaeus sp.) that have caused major economic losses to shrimp aquaculture. A rapid and highly sensitive detection and quantification method for TSV and YHV was developed using the GeneAmp 5700 Sequence Detection System and SYBR Green chemistry. The reverse transcriptase polymerase chain reaction (RT-PCR) mixture contained a fluorescent dye, SYBR Green, which exhibits fluorescence enhancement upon binding to double strand cDNA. The enhancement of fluorescence was found to be proportional to the initial concentration of the template cDNA. A linear relationship was observed between input plasmid DNA and cycle threshold (C(T)) values for 10(6) down to a single copy of both viruses. To control for the variation in sample processing and in reverse transcription reaction among samples, shrimp beta-actin and elongation factor-1alpha (EF-1alpha) genes were amplified in parallel with the viral cDNA. The sensitivity and the efficiency of amplification of EF-1alpha was greater than beta-actin when compared to TSV and YHV amplification efficiency suggesting that EF-1alpha is a better internal control for the RT-PCR detection of TSV and YHV. In addition, sample to sample variation in EF-1alpha C(T) value was lower than the variation in beta-actin C(T) value of the corresponding samples. The specificity of TSV, YHV, EF-1alpha and beta-actin amplifications was confirmed by analyzing the dissociation curves of the target amplicon. The C(T) values of TSV and YHV samples were normalized against EF-1alpha C(T) values for determining the absolute copy number from the standard curve of the corresponding virus. The method described here is highly robust and is amenable to high throughput assays making it a useful tool for diagnostic, epidemiological and genetic studies in shrimp aquaculture.

Actins↗

PubMed1989

The chick's marginal zone and primitive streak formation. II. Quantification of the marginal zone's potencies--temporal and spatial aspects.

When a posterior fragment of the chick's marginal zone (PM) was exchanged with equal sized lateral marginal zone fragment (LM), of the same blastoderm, its capacity to initiate an ectopic primitive streak (PS) was found to be both size and stage dependent. Good correlation was demonstrated between the areas of PM fragments and the number of cells they contained. In stage X blastoderms, PM fragments containing less than 1200 cells were incapable of initiating an ectopic PS. Transplanted PMs containing between 1200 and 1500 cells initiated a lateral ectopic PS in 50% of the cases, while in the other 50% a posterior PS developed from the original posterior side. PMs containing 1500 cells or more in all cases initiating an ectopic PS and inhibited the formation of a posterior PS. At stage XI, laterally transplanted PMs containing less than 1800 cells were not effective. Stage XI PMs containing 1800-2300 cells in some cases succeeded in initiating a lateral ectopic PS, in addition to the posterior one. Stage XI PMs containing 2300 cells or more invariably promoted the development of an ectopic PS, but were unable to suppress the formation of a posterior PS, so that two PSs developed in the same blastoderm, one posterior and one ectopic. When a stage XI PM fragment was laterally transplanted into a younger, stage X blastoderm, the minimal effective cell number needed to initiate an ectopic PS increased to at least 3000 cells, again without inhibiting the formation of a posterior PS. The inductive potential of a stage X PM is therefore at least twice that of a stage XI PM. The marginal zone belt of stage X blastoderms was checked for the decrease in its developmental potential from the posterior to the lateral side by evaluating its effect on the developmental expression of two competing stage X PMs, one located posteriorly and the other inserted laterally. The developmental expression of the laterally inserted PM was consistently inferior to that of the posterior PM. The developmental expression of each PM was not related to absolute size, but depended on the size ratio of lateral PM/posterior PM. When the ratio was 1.2 or less, only posterior PSs developed. When the ratio was 1.3-1.4, three different results were encountered: (1) only a posterior PS, (2) posterior plus lateral, and (3) only lateral PS. When the ratio was 1.5 or more, only a lateral PS developed, which suppressed the posterior PS.

Animals↗

PubMed2026

Manual, digital, and AI tumour-infiltrating lymphocyte scoring: a secondary analysis of the APHINITY randomised trial.

BACKGROUND: Stromal tumour-infiltrating lymphocytes (sTILs) are prognostic in early-stage HER2-positive breast cancer, but their role in the context of dual HER2 blockade remains undefined. We evaluated manual, digital, and artificial intelligence (AI)-based sTIL quantification, together with AI-derived spatial metrics, for prognostic and treatment-benefit stratification using tumour samples from the phase 3 APHINITY trial. METHODS: In the APHINITY trial, 4805 patients were randomly assigned to receive chemotherapy plus trastuzumab with pertuzumab or chemotherapy plus trastuzumab with placebo. Median follow-up was 74·1 months (IQR 68·3-75·4). We analysed 4262 haematoxylin and eosin-stained images using manual assessment, an automated digital approach, AI-based lymphocyte quantification (AI percentage lymphocytes), and two AI-derived spatial features (AI-TIL and immune hotspot). Interobserver reproducibility was assessed in 262 randomly chosen tumour samples scored independently by five pathologists. Multivariable Cox models were used to assess associations between TIL levels and invasive disease-free survival (primary outcome in APHINITY), distant recurrence-free interval, and overall survival. The heterogeneity of pertuzumab benefit was evaluated using subgroup analyses, subpopulation treatment effect pattern plot analyses, and nested Cox models with treatment-by-biomarker interaction terms. FINDINGS: Manual scoring showed high interobserver reproducibility (intraclass correlation coefficient 0·84 [95% CI 0·79-0·88]). Concordance between manual and automated methods was modest. AI-based scoring (AI percentage lymphocytes) reclassified 120 (11·6%) of 1035 node-positive tumours from immune-low (by manual scoring) to immune-high; this subgroup of patients showed greater separation of 5-year invasive disease-free survival curves between pertuzumab and placebo groups compared with patients whose tumours were concordantly classified as immune-low by both manual and AI-based approaches. Higher levels of TILs were associated with improved invasive disease-free survival for all sTIL measurement approaches and spatial measurements (hazard ratios [HRs] 0·41-0·93). Pertuzumab was associated with improved invasive disease-free survival at higher sTIL levels across all measurement approaches (HRs 0·36-0·48), but was not associated with higher values of spatial measures. The largest 6-year absolute improvements with pertuzumab were observed in patients with node-positive disease whose tumours scored in the highest level of immune infiltration of manual sTIL scoring (≥70·0%; mean absolute improvement 12·1 percentage points [SD 2·8]). In nested prognostic and predictive models, AI-based immune hotspot scores provided the most consistent additional information when combined with any sTIL measurement (all p<0·010). INTERPRETATION: Standardised manual sTIL scoring was reproducible, and digital and AI-based methods showed consistent prognostic stratification and potential for treatment-benefit stratification despite only modest correlation between platforms. AI spatial metrics provided complementary information beyond sTIL density and could support more scalable immune assessment. Future studies are needed to validate these approaches in independent cohorts and to clarify their clinical utility for stratifying contemporary HER2-directed therapies. FUNDING: None.

Humans↗

PubMed1999

Mass isotopomer distribution analysis at eight years: theoretical, analytic, and experimental considerations.

Mass isotopomer distribution analysis (MIDA) is a technique for measuring the synthesis of biological polymers. First developed approximately eight years ago, MIDA has been used for measuring the synthesis of lipids, carbohydrates, and proteins. The technique involves quantifying by mass spectrometry the relative abundances of molecular species of a polymer differing only in mass (mass isotopomers), after introduction of a stable isotope-labeled precursor. The mass isotopomer pattern, or distribution, is analyzed according to a combinatorial probability model by comparing measured abundances to theoretical distributions predicted from the binomial or multinomial expansion. For combinatorial probabilities to be applicable, a labeled precursor must therefore combine with itself in the form of two or more repeating subunits. MIDA allows dilution in the monomeric (precursor) and polymeric (product) pools to be determined. Kinetic parameters can then be calculated (e.g., replacement rate of the polymer, fractional contribution from the endogenous biosynthetic pathway, absolute rate of biosynthesis). Several issues remain unresolved, however. We consider here the impact of various deviations from the simple combinatorial probability model of biosynthesis and describe the analytic requirements for successful use of MIDA. A formal mathematical algorithm is presented for generating tables and equations (APPENDIX), on the basis of which effects of various confounding factors are simulated. These include variations in natural isotope abundances, isotopic disequilibrium in the precursor pool, more than one biosynthetic precursor pool, incorrect values for number of subunits present, and concurrent measurement of turnover from exogenously labeled polymers. We describe a strategy for testing whether isotopic inhomogeneity (e.g., an isotopic gradient or separate biosynthetic sites) is present in the precursor pool by comparing higher-mass (multiply labeled) to lower-mass (single- and double-labeled) isotopomer patterns. Also, an algebraic correction is presented for calculating fractional synthesis when an incomplete ion spectrum is monitored, and an approach for assessing the sensitivity of biosynthetic parameters to measurement error is described. The different calculation algorithms published for MIDA are compared; all share a common model, use overlapping solutions to computational problems, and generate identical results. Finally, we discuss the major practical issue for using MIDA at present: quantitative inaccuracy of instruments. The nature and causes of analytic inaccuracy, strategies for evaluating instrument performance, and guidelines for optimizing accuracy and reducing impact on biosynthetic parameters are suggested. Adherence to certain analytic guidelines, particularly attention to concentration effects on mass isotopomer ratios and maximizing enrichments in the isotopomers of interest, reduces error. Improving instrument accuracy for quantification of isotopomer ratios is perhaps the highest priority for this field. In conclusion, MIDA remains the "equation for biosynthesis," but attention to potentially confounding factors and analytic performance is required for optimal application.

Isotopes↗