Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “protocol optimization”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 775 records · Page 43Linked to original sources
PubMed1999

Identification and optimization of regeneration conditions for affinity-based biosensor assays. A multivariate cocktail approach.

A general regeneration, identification, and optimization (RO) protocol for Biacore systems was developed. The RO protocol uses six multi-ingredient stock solutions that represent the six most common chemical properties employed as regeneration agents. The regeneration effect of different regeneration cocktails of these six stock solutions were tested iteratively until a satisfactory result was obtained. The RO protocol was designed with an ease-of-use and multivariate approach. The RO protocol was tested on 13 different antibody-antigen systems. For 10 of these, only the first screening session was tested. For 9 of the 13 systems, the RO-protocol screening session identified cocktails that removed more than 90% of the bound analyte in a 30 s pulse. For 5 systems, the RO protocol identified cocktails that regenerated the surface completely and that were more gentle than previously used regeneration conditions. Furthermore, the regeneration optimization results can be interpreted as a characterization of the interacting molecules. The relevance of testing cocktails was justified by the fact that at least one cocktail was significantly better than all diluted stock solutions for all tested model systems. By using the multivariate approach, the risk of missing relevant combinations of stock solutions was minimized. This resulted in an unexpected discovery of excellent properties of EDTA as an additive in regeneration cocktails containing chaotropic agents and ions in high concentration.

Biosensing Techniques↗
PubMed2004

[Evaluation of different 16-row CT colonography protocols using a porcine model].

PURPOSE: To develop and to test an easily produced biological colon model with simulated polypoid lesions. Application of this phantom for the selection of an optimized scan protocol of 16-row CT colonography (CTC) for clinical use. METHODS AND MATERIAL: Six polypoid lesions (1 - 6 mm) were simulated with sutures on the inner face of a porcine colon segment (20 cm). After distending the colon segment with air, the phantom was placed in a water quench and CT scans were performed on a MDCT-scanner (Somatom Sensation 16, Siemens, Forchheim). At constant values for collimation (16x0.75 mm) and voltage (120 kV), 54 different combinations of mAs values (50, 75 and 100 mAs), pitch factors (1, 1.25 and 1.5) and slice thicknesses (0.75, 1.0, 1.5, 2.0, 3.0 and 5.0 mm) were tested systematically. The phantom was scanned in the longitudinal and transverse axis to simulate the different orientation of the colon in the abdomen. Axial slice images and virtual endoscopic views of all data sets were presented separately to 2 radiologists who independently determined number and size of detectable polyps. Dose exposure was measured with an Alderson phantom. RESULTS: The colon model offered a realistic imitation of a polyp-covered, human colon. The experimental set-up allowed a systematic evaluation of polyp detection related to lesion size, orientation of the colon and CTC parameters, with other influencing factors mostly excluded. Polyps were significantly better detected in the longitudinal than in the transverse orientation of the colon. For the detection of lesions of at least 3 mm, a low dose (50 mAs) 16-row CTC should be combined with a pitch of 1.5 and a maximum slice thickness of 3 mm. For the depiction of polyps smaller than 3 mm, slice thickness and pitch should amount to 1 mm and 1.0, respectively. Effective dose of this low dose protocol is 4.08 mSv. CONCLUSION: The porcine colon phantom represents a realistic and easily produced alternative to other colonography models. It allows a preselection of a CTC-protocol for subsequent clinical testings. If the high in vitro performance of the low-dose 16-row CTC-protocol is confirmed on a human collective, the use of 16-row technique would represent a big step for CTC toward a screening method.

Animals↗
PubMed1991

A qualitative approach to cell growth modeling and simulation for cancer chemotherapy.

A qualitative model of cell growth, based on qualitative process theory, is presented. The model can be used to analyze the effects of the interaction of antiproliferative drugs on cells when the effects of each specific drug are known, mainly when designing multi-drug protocols for optimal cancer treatment.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed2005

Imaging in breast cancer: magnetic resonance imaging.

Over the past 5 years there has been a marked increase in the use of magnetic resonance imaging (MRI) of the breast. Multiple research studies have confirmed improved cancer detection, diagnosis, and evaluation of response to therapy with breast MRI compared with mammography and ultrasound. As this exciting new technology advances, focused work in optimal scan protocols, appropriate clinical applications, and image interpretation are needed. Both the potential benefits and harms need to be evaluated to guide optimal use of this imaging modality in select patient populations.

Biopsy↗
PubMed2026

Establishment of an efficient Agrobacterium-mediated genetic transformation protocol for Saccharum officinarum using Black Cheribon as a model genotype.

Efficient Agrobacterium-mediated transformation (AMT) is vital for the biotechnological improvement of sugarcane (Saccharum spp.). Saccharum officinarum is the main ancestor of all modern cultivars, yet little research has been conducted on its AMT system. In this work, an efficient AMT protocol for S. officinarum was developed, with Black Cheribon as the model genotype owing to its superior tissue culture performance and regeneration capacity. The optimized agro-infection protocol comprised the following main parameters: concentration of acetosyringone (AS) in Agrobacterium culture, concentration of AS for infection, Agrobacterium concentration at OD600 = 0.4, infection time of 30 minutes, vacuum infiltration time of 10 minutes and co-cultivation time of 3 days. To further improve transformation efficiency, 0.5 mg/L thidiazuron and 200 mg/L citric acid were added to the regeneration medium, which enhanced the regeneration of shoots. A modified stage-dependent selection strategy (FlexII) was established by using glufosinate-ammonium at concentrations of 2.0, 1.0, and 0.75 mg/L in the callus proliferation, shoot regeneration, and rooting stages, respectively. This strategy was more successful than the minimum inhibitory concentration-based strategy in S. officinarum transformation. The optimized protocol further boosted the transformation efficiency of Black Cheribon from 1.12% to 7.17%. The resulting transgenic lines were confirmed by PCR amplification of T-DNA regions and immunochromatographic detection of Bar protein expression in primary transformants, respectively. These results provide a sound technical foundation for the functional genomics and biotechnological optimization of S. officinarum germplasm, and may serve as a reference for future transformation studies in other sugarcane germplasm.

Agrobacterium↗
PubMed2002

Conformal radiotherapy optimization with micromultileaf collimators: comparison with radiosurgery techniques.

PURPOSE: Conformal radiotherapy (CRT) consists of irradiating the target volume while avoiding the healthy peripheral tissues and organs at risk as far as possible. One technique used to treat intracranial tumors consists of using micromultileaf collimators (MMLCs). Given the dose constraints involved, it is of interest to optimize MMLC irradiation parameters and compare the results of this technique with those of conventional radiosurgery (RT) techniques (Gamma Knife and linear accelerator stereotactic RT). METHODS AND MATERIALS: MMLC protocols are optimized in two stages. The orientation of the fields, delimited by a beam's eye view technique, is determined using a genetic algorithm method. The weighting of the fields and subfields when using intensity modulation and the position of the leaves are optimized using a simulated annealing method. We compared the results obtained for 8 clinical cases using 5 intensity-modulated fields with those obtained using the two radiosurgery techniques. The comparison indexes are those defined by the Radiation Therapy Oncology Group (RTOG). RESULTS: The results of this study demonstrated the advantages of using intensity modulation and the improvement obtained for the RTOG indexes in the case of CRT with MMLC, although the healthy peripheral tissues were less exposed to radiation with the radiosurgery techniques. The results also highlight the difficulty encountered with radiosurgery techniques in obtaining satisfactory dose homogeneity when the protocol is defined with numerous iosocenters. CONCLUSION: In CRT with MMLC, intensity modulation makes it possible to reduce the number of fields used. It is especially useful to optimize the orientations in the case of target volumes of complex shape or when volumes at risk are in the vicinity of the target. If used correctly, MMLC can be a valuable alternative to conventional radiosurgery techniques.

Algorithms↗
PubMed2001

Timing of leukocyte filtration during cardiopulmonary bypass.

The effects of leukocyte filtration on the outcome of cardiac surgery with cardiopulmonary bypass (CPB) have been shown by numerous investigators. In the majority of cases a leukocyte filter is placed in the arterial line instead of a standard arterial line filter and used throughout CPB. However, protocols to optimize onset and duration of leukocyte filtration have not been sufficiently evaluated to date. In this paper, current efforts to improve such protocols are demonstrated and discussed. These efforts are based on studies of leukocyte pathogenicity during cardiac surgery. A first study (double-blind randomized) was performed in routine coronary artery bypass graft (CABG) patients to evaluate whether short-term leukocyte filtration during reperfusion by release of the aortic crossclamp would reduce reperfusion-associated myocardial damage. Further data compare the efficacy of three different filtration concepts to reduce CPB- and/or reperfusion-associated leukocyte pathogenicity. Clinical endpoints, standard laboratory variables and functional in vitro assays are provided and discussed.

Cardiopulmonary Bypass↗
PubMed2004

Microarray results improve significantly as hybridization approaches equilibrium.

Dual-channel long oligonucleotide microarrays are in widespread use. Although much attention has been given to proper experimental design and analysis regarding long oligonucleotide microarrays, relatively little information is available concerning the optimization of protocols. We carried out a series of microarray experiments designed to investigate the effects of different levels of target concentration and hybridization times using a long oligonucleotide library. Based on principles developed from nucleic acid renaturation kinetics studies, we show that increasing the time of hybridization from 18 h to 42 h and 66 h, especially when lower than optimal concentrations of target were used, significantly improved the quality of the microarray results. Longer hybridization times significantly increased the number of spots detected, signal-to-noise ratios, and the number of differentially expressed genes and correlations among replicate arrays. We conclude that at 18 h of incubation, target-to-probe hybridization has not reached equilibrium and that a relatively high proportion of nonspecific hybridization occurs. This result is striking, given that most, if not all, published microarray protocols stipulate 8-24 h for hybridization. Using shorter than optimal hybridization times (i.e., not allowing hybridization to reach equilibrium) has the consequence of underestimating the fold change of differentially expressed genes and of missing less represented sequences.

Algorithms↗
PubMed2002

DNA topoisomerases in cancer chemotherapy: using enzymes to generate selective DNA damage.

DNA topoisomerase II targeting agents such as etoposide and doxorubicin are well-established cancer chemotherapeutic agents. Topotecan (Hycamtin) and irinotecan (Camptosar) are launched drugs that target topoisomerase I and have significant activity against many solid malignancies. These agents have important mechanistic similarities, converting their target enzyme(s) to generate DNA damage. Recent structural and biochemical studies on targeting of topoisomerases by antitumor agents are providing a framework for understanding drug action at the enzyme level, and at the level of cellular pathways important for responses to this unique type of DNA damage. These investigations into the mechanisms of action of topoisomerase-targeting agents should aid in the design of dinical protocols that optimize the activity of these agents.

Animals↗
PubMedDate not supplied

Factors influencing vascular and hepatic enhancement at CT: experimental study on injection protocol using a canine model.

PURPOSE: The purpose of this work was to evaluate the effects of contrast medium injection parameters on aortic, portal vein, and hepatic enhancement at spiral CT and to assess optimal injection protocol for hepatic CT. METHOD: Ten 15 kg dogs underwent single level dynamic CT through the hepatic hilum at 5 s intervals just after the injection of contrast medium for 3 min. With use of different volumes (1, 2, and 3 ml/kg), injection rates (0.5, 1, and 2 ml/s), and concentrations (150, 200, and 300 mg/ml), a total of 270 spiral CT scans were performed. In each scan, time-attenuation curves of aorta, portal vein, and liver were obtained. The degree of maximum contrast enhancement (Imax), time to maximum enhancement (Tmax), and time to equilibrium phase (Teq) for to each injection protocol were analyzed. RESULTS: Alterations in contrast material volume, injection rate, and concentration had significant impact on contrast enhancement of the liver. With increasing volume of contrast medium, Imax, Tmax, and Teq of aorta, portal vein, and liver increased (p < 0.005). With increasing rate of injection, on the other hand, Imax of aorta and liver increased (p < 0.05), but Tmax and Teq decreased (p < 0.005). Change of concentration of contrast medium had a significant effect on Imax of vessels (p < 0.05). CONCLUSION: Maximum contrast enhancement of liver and vessels was influenced mainly by injection volume of contrast medium and the time to peak enhancement by injection rate of contrast medium. Under given amounts of contrast medium, therefore, the strategy of increasing volume by dilution and faster injection might give better Imax values without penalty for the duration of an optimal temporal window (Tmax and Teq).

Animals↗
PubMed2000

Adult rat and human bone marrow stromal cells differentiate into neurons.

Bone marrow stromal cells exhibit multiple traits of a stem cell population. They can be greatly expanded in vitro and induced to differentiate into multiple mesenchymal cell types. However, differentiation to non-mesenchymal fates has not been demonstrated. Here, adult rat stromal cells were expanded as undifferentiated cells in culture for more than 20 passages, indicating their proliferative capacity. A simple treatment protocol induced the stromal cells to exhibit a neuronal phenotype, expressing neuron-specific enolase, NeuN, neurofilament-M, and tau. With an optimal differentiation protocol, almost 80% of the cells expressed NSE and NF-M. The refractile cell bodies extended long processes terminating in typical growth cones and filopodia. The differentiating cells expressed nestin, characteristic of neuronal precursor stem cells, at 5 hr, but the trait was undetectable at 6 days. In contrast, expression of trkA, the nerve growth factor receptor, persisted from 5 hr through 6 days. Clonal cell lines, established from single cells, proliferated, yielding both undifferentiated and neuronal cells. Human marrow stromal cells subjected to this protocol also differentiated into neurons. Consequently, adult marrow stromal cells can be induced to overcome their mesenchymal commitment and may constitute an abundant and accessible cellular reservoir for the treatment of a variety of neurologic diseases.

Animals↗
PubMed1991

An improved method of loading pH-sensitive liposomes with soluble proteins for class I restricted antigen presentation.

We have recently shown that ovalbumin (OVA) entrapped in pH-sensitive liposomes could sensitize mouse thymoma cells for lysis by MHC class I-restricted cytotoxic T lymphocytes (CTL) (Reddy et al. (1991) J. Immunol. Methods, 141, 157-163). The present studies were designed to optimize the antigen delivery system. A simple freeze-thaw method was developed to load OVA into pH-sensitive liposomes, and the protocol was optimized in terms of the choice of buffer, pH and ionic strength of the medium, lipid composition, lipid and OVA concentrations and the number of freeze-thaw cycles. Under optimized conditions, approximately 25% of OVA could be entrapped in pH-sensitive liposomes at 172 micrograms protein/mg lipid. This compares to only about 5% entrapment (70 micrograms protein/mg lipid) using the previous method. OVA loaded to pH-sensitive liposomes using the improved method led to a sensitive measure of CTL activity. The approach promises to be suitable to measure CTL against less available soluble antigens such as viral proteins.

Animals↗
PubMed1996

Simultaneous enumeration of chromosomes 13, 18, 21, X, and Y in interphase cells for preimplantation genetic diagnosis of aneuploidy.

A fluorescence in situ hybridization (FISH) protocol to simultaneously enumerate chromosomes 13, 18, 21, X, and Y in interphase cell nuclei for application in preimplantation genetic diagnosis (PGD) of aneuploidy was tested. Strict scoring criteria were developed to minimize recording errors. The protocol used optimized probes for chromosome-specific DNA repeat and single-copy loci and showed a significantly higher efficiency (95%) than previously published protocols. The other purpose of this study was to differentiate between two signals originating from a single split target or from two targets in close proximity. These criteria were based on the FISH results obtained from the analysis of all or most of the cells from 50 chromosomally normal or mosaic human embryos, 20 aneuploid embryos, and five polyploid embryos donated for research. Subsequently, 183 human embryos underwent PGD of aneuploidy in one of their cells. In 64 embryos that were not transferred back to the uterus, the rest of cells were also analyzed, and the previous results were confirmed in 91% of these embryos, with 0% (0/21) of the embryos classified as normal embryos being abnormal and 14% (6/43) of the embryos classified as abnormal being normal. Compared to previous protocols, these criteria minimize the risk of transferring abnormal embryos after PGD analysis.

Aneuploidy↗
PubMed2006

Poplar (Populus spp.).

Although species within the genus Populus are, in general, easier to transform and regenerate in vitro than most other trees, many poplar species are very recalcitrant. Many protocols that previously have been reported were developed for a specific genotype or species. Thus, it has often been necessary to re-optimize a protocol each time research is initiated with a new genotype. The method presented in this chapter has been effective for a wide variety of poplar genotypes.

Agrobacterium tumefaciens↗
PubMed1995

High-throughput method for determination of apolipoprotein E genotypes with use of restriction digestion analysis by microplate array diagonal gel electrophoresis.

Molecular epidemiological research has identified the association of a common apolipoprotein E (apo E) isoform (E4 as opposed to E3), with risk both of coronary artery disease and of Alzheimer dementia. In addition, the role of apo E genotype (usually E2/E2) in Type III hyperlipidemia is well known. However, both for diagnostic and research purposes, apo E genotyping is cumbersome. The preferred approach is electrophoretic sizing of restriction digestion fragments, enabling simultaneous analysis of the two codons (112 and 158) that represent the six common genotypes (E2/E2; E2/E3; E2/E4; E3/E3; E3/E4; E4/E4). However, the consequent demands of high-yield PCR, high-resolution, high-throughput electrophoresis, and sufficient detection sensitivity have left shortfalls in published protocols. In conjunction with a high-throughput electrophoresis system we described recently, microplate array diagonal gel electrophoresis (MADGE), we have constructed extensively optimized, simplified protocols for DNA isolation from mouthwash samples for PCR setup and high-yield PCR, for restriction digestion, and for subsequent MADGE gel image analysis. The integral system enables one worker to readily undertake apo E genotyping of as many as hundreds of DNA samples per day, without special equipment.

Apolipoproteins E↗
PubMed1997

13C-urea breath test in Helicobacter pylori diagnosis and eradication. Correlation to histology, origin of 'false' results, and influence of food intake.

BACKGROUND: Which protocol is optimal for the 13C-urea breath test (UBT) for Helicobacter pylori detection is controversial. This study aimed to characterize a very simple UBT protocol for the clinical routine (two-point-analysis performed with 75 mg 13C-urea and citric acid) with special consideration of 'false' UBT results. RESULTS; UBT was evaluated in reference to histology (Warthin-Starry). In mismatching results re-gastroscopy was performed. By UBT, 74 of 77 patients with H. pylori-positive histology were detected (sensitivity, 96%). The false-negative UBTs were due to low colonization densities during spontaneous H. pylori elimination or pyloric obstruction. Seven of 49 patients with negative histology had a positive UBT, but re-gastroscopy showed that all of them had a positive histology when multiple antral biopsy specimens were taken (UBT specificity, 100%). UBT correlated only weakly with H. pylori colonization density. No correlation was found between UBT and gastric neutrophil and lymphocyte infiltration. UBT reproducibility was excellent (93 of 94 in a 6-month period). Non-fasting conditions induced a shift to lower UBT results in H. pylori-positive and to higher UBT results in negative patients, resulting in 2 of 10 false-positive and 1 of 10 false-negative UBTs. CONCLUSION: This simple version of the urea breath test combines the highest sensitivity with excellent reproducibility. It is superior to histologic detection of H. pylori in the clinical routine and an optimal tool for monitoring H. pylori eradication. Fasting conditions are required for the test.

Breath Tests↗
PubMed2004

Echo-planar BOLD fMRI of mice on a narrow-bore 9.4 T magnet.

The feasibility of BOLD fMRI in association with electrical somatosensory stimulation on spontaneously breathing, isoflurane-anesthetized mice was investigated using spin-echo, echo-planar imaging (EPI) on a vertical narrow-bore 9.4 T magnet. Three experiments were performed to derive an optimal fMRI protocol. In Experiment 1 (n = 9), spin-echo BOLD responses to 10% CO2 challenge under graded isoflurane (0.25-1.25%) ranged from 10 +/- 2% to 3.5 +/- 0.9%; the optimal BOLD contrast-to-noise ratio peaked at 0.75% isoflurane. In Experiment 2 (n = 6), hindpaw somatosensory stimulations using 1-7 mA under 0.75% isoflurane revealed the optimal BOLD response was at 6 mA. In Experiment 3 (n = 5), BOLD responses to 4 and 6 mA stimulation under 0.75% and 1% isoflurane were evaluated in detail, confirming the optimal conditions in Experiment 2. These results demonstrated that BOLD fMRI using single-shot, spin-echo EPI in a mouse somatosensory stimulation model could be routinely performed on high-field, vertical, narrow-bore magnets. This protocol might prove useful for fMRI studies of transgenic mice.

Anesthetics, Inhalation↗
PubMed1995

Radiation doses to paediatric patients undergoing micturating cystourethrography examinations and potential reduction by radiation protection optimization.

Dose-area product values were measured in paediatric micturating cystourethrography examinations in two dedicated facilities. The results are compared, taking into account features of the X-ray rooms and differences in examination protocol. Several optimization procedures were simulated or implemented, in order to estimate the dose savings which could be attained. Potential savings identified in the dose-area values reach about 85% for both centres.

Adolescent↗