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Differentiation of isomers by wavelength-tunable infrared multiple-photon dissociation-mass spectrometry: application to glucose-containing disaccharides.

Variation in the wavelength of irradiation in infrared multiple-photon dissociation (IR-MPD) of lithium-tagged glucose-containing disaccharide ions (1-2-, 1-3-, 1-4-, and 1-6-linked isomers of both anomeric configurations) resulted in marked differences in their mass spectral fragmentation patterns. Two-dimensional plots of the fragment yield versus infrared wavelength for each mass spectral product ion were unique for each isomer and can be considered a spectral fingerprint. Individual product ions or diagnostic ratios of key product ions can be optimized at specific IR wavelengths. The technique permits both linkage position and anomeric configuration to be assigned. The ratio of the fragments derived by cleavage at the glycosidic bond (m/z 169/187) is much enhanced for beta-anomers compared to alpha-anomers. Differences in the diagnostic product ions 169 and 187 were largest in the range of 9.0-9.4 microm, where the maximum dissociation yield was observed. Conversely, at 10.6 microm, the wavelength of nontunable CO2 lasers that accompany commercial Fourier transform ion cyclotron resonance mass spectrometers, the dissociation yield was poor and anomeric differentiation was not possible. In contrast to previous studies by collision-induced dissociation, the trends in dissociation behavior between anomers using IR-MPD are significant and allow simple diagnostic rules to be established. By depositing energy into these isobaric ions via narrow-band IR excitation, the resulting internal energy can be finely controlled, thereby obtaining high reproducibility in dissociation patterns. Given the multidimensionality of variable-wavelength IR-MPD of lithiated disaccharides, it is expected that this approach can overcome some of the current limitations in isomer differentiation.

Carbohydrate Sequence↗

An algorithm for diagnosis and therapy of management and complications of urolithiasis during pregnancy.

From 1 January 1986 to 30 June 1989, 15 pregnant women were diagnosed as having urolithiasis. Patients presented in the last two trimesters with an infection of the lower part of the urinary tract (60 per cent), flank and abdominal pain (27 per cent) and hematuria (13 per cent). Ultrasonographic findings confirmed the diagnosis in 47 per cent of the patients. Other roentgenologic procedures were required in the remaining patients. Initially, therapy was conservative in all, and in 67 per cent of patients, no further intervention was necessary. Only 33 per cent required invasive measures; cystoscopic passage of an internal ureteral stent was the initial procedure of choice at our institution. Three of five patients who underwent invasive procedures had surgical intervention for relief of ureteral obstruction. Intensive care management was necessary for one of these three patients who had acute hemorrhage occur during the procedure. These data emphasize the need for the accurate diagnosis of urolithiasis during pregnancy. Ultrasonography was a valuable diagnostic technique, but a limited excretory urogram is safe and appropriate when there is uncertainty. Conservative management (hydration, analgesia and antibiotics as indicated) of obstructive uropathy was successful in the majority of instances. A specific clinical algorithm facilitated the successful management of patients necessitating operative intervention. Optimal management requires clinical suspicion and a precise diagnostic and therapeutic plan.

Adult↗

Copious production of SARS-CoV nucleocapsid protein employing codon optimized synthetic gene.

The severe acute respiratory syndrome coronavirus (SARS-CoV) nucleocapsid protein (NP) is one of the predominant antigenic protein and the most abundant shed antigen throughout the SARS-CoV infection. This feature makes it a suitable molecular target for diagnostic applications. In this study the full length codon optimized NP gene and its subfragment gene segment was cloned in a bacterial expression vector. The full length NP could be expressed in E. coli at very high level within inclusion bodies. The inclusion bodies were successfully solubilized, purified under denaturing conditions employing IMAC column and refolded. The non-glycosylated NP was used to immunize mice for hybridoma development. The polyclonal antiserum from animals immunized with this recombinant NP protein was found to specifically recognize the NP and its subfragments, thus demonstrating the immunogenic nature of the recombinant protein. The NP antigen or a subfragment could be useful for developing a sensitive serum diagnostic assay to monitor SARS-CoV outbreaks by detecting the early human anti-SARS antibodies. In addition, the availability of the NP fragments could facilitate epitope mapping of anti-NP monoclonals for identifying suitable sandwich pairs.

Animals↗

Analysis of morphologic patterns of fine-needle aspiration of the breast to reduce false-negative results in breast cytology.

BACKGROUND: The identification of specific morphologic diagnostic criteria is of paramount importance to optimize the accuracy of fine-needle aspiration cytology (FNAC) and to reduce the rate of false-negative results. In the current study, the authors reviewed a consecutive series of false-negative findings observed in the study center to define the presence and degree of cytologic abnormalities. False-negative cases were randomly mixed with true-negative cases and were reviewed by a panel of expert readers in a blinded fashion. The main objective of the current study was to identify a morphologic pattern that may permit the reduction of false-negative findings while maintaining the specificity of FNAC. METHODS: A blind review of a set of 41 consecutive false-negative and 49 true-negative breast aspiration samples was performed by a panel of 10 expert cytologists who were asked to give a final report and to classify the samples according to classic morphologic parameters. RESULTS: The majority final report sensitivity was 54% (range, 19-61%) and specificity was 73% (range, 65-92%). The average concordance with the majority report, adjusted for chance agreement (kappa statistic), was moderate at 0.54 (range, 0.40-0.65). Enlarged nuclear size, a hyperchromatic nucleus, the absence of naked nuclei, and the absence of apocrine metaplasia were reported more frequently in carcinoma cases, although not to a significant extent. The only variable found to be associated significantly (P = 0.041) with a diagnosis of carcinoma was the presence of microcalcifications, which nevertheless were found to occur in only a minority of carcinoma cases (7 of 41 cases) or controls (2 of 49 controls). Multivariate analysis demonstrated that the presence of microcalcifications (odds ration [OR] of 3.0; 95% confidence interval [95% CI], 1.2-7.4), the absence of naked nuclei (OR of 2.4; 95% CI, 1.3-4.4), and enlargement of the nucleus (OR of 1.9; 95% CI, 1.1-3.4) were all independently associated with false-negative findings. Diagnostic accuracy using a morphology-based score did not appear to improve the results substantially compared with the final report (sensitivity of 0.46 vs. 0.54 [P = 0.508] and a specificity of 0.80 vs. 0.73 [P = 0.218]). CONCLUSIONS: The results of the current study confirm that breast FNAC false-negative results are at least partially the result of underreporting of abnormalities that may be noted at review. Detailed analysis of a single morphologic characteristic was found to be of limited diagnostic value, suggesting that operators do perceive abnormalities but cannot translate these findings into distinct morphologic categories.

Biopsy, Fine-Needle↗

Performance and Cost evaluation of one commercial and six in-house conventional and real-time reverse transcription-pcr assays for detection of severe acute respiratory syndrome coronavirus.

We evaluated seven reverse transcription-PCR (RT-PCR) assays, including six in-house assays and one commercial assay for the detection of severe acute respiratory syndrome coronavirus (SARS-CoV) RNA in clinical specimens. RT-PCR assays targeted different genomic regions and included three conventional assays (one nested and two non-nested) run on a conventional heat block and four real-time assays performed in a LightCycler (LC; Roche Diagnostics). All in-house assays were optimized for assay parameters, including MgCl2, primer, and probe concentrations. The commercial assay was the RealArt HPA CoV RT-PCR assay (Artus), which was run in the LC. Testing serial dilutions of cultured SARS-CoV showed that the analytical sensitivity of the assays ranged from 10(-8) to 10(-6), corresponding to 1 and 100 copies of viral RNA, respectively. Significant differences in analytical sensitivities were observed between assays (P < 0.01, probit regression analysis for 50% sensitivity levels for the top two assays versus the others). Testing 68 clinical specimens (including 17 respiratory tract specimens, 29 urine samples, and 22 stools or rectal swabs) demonstrated that six of the seven assays detected at least 17 of 18 positives (defined as positive in at least two assays), and two of the assays had a sensitivity of 100%. There were no significant differences in sensitivity between the assays (P = 0.5 [Cochrance Q test, least sensitive 15 of 18 versus 18 of 18]). The specificities of the assays ranged from 94.0 to 100% without significant differences (P = 0.25 to 0.5 [McNemar test]). The reagent and technologist cost of performing the in-house PCR assays ranged from $5.46 to $9.81 Canadian dollars (CDN) per test. The commercial assay cost was considerably higher at $40.37 per test. The results demonstrated good performance for all assays, providing laboratories that need to do SARS RNA testing with a choice of assay formats.

Costs and Cost Analysis↗

Differential tumor markers and hepatitis markers profile in liver tumors.

The differential diagnostic utility of AFP, CEA, CA19.9 and TPA was evaluated in liver tumors. They were determined in the sera of 61 patients with primary hepatocellular carcinoma (HCC), 18 with secondary liver metastasis, 61 of benign liver cirrhosis in comparison to 20 normal healthy control subjects. The association of either HBV or HCV infection and HCC was also studied through the assay of HbSAg, HbSAb, and HCV-Ab. The optimal cut-off values were determined using the diagnostic accuracy measurements and the receiver operating characteristic (ROC) curves. AFP at an optimal cut-off value of 100 ng/ml and TPA at 160 U/L showed the highest sensitivity and specificity in detecting liver metastasis (100% and 87% for AFP; 100% and 54% for TPA respectively). The obtained data indicated that the combined assay of AFP and TPA resulted in a better discrimination of HCC among patients with hepatic focal lesions. HCV-Ab was detected in a higher ratio of HCC patients (83.6%) compared to HbsAg (68.9%), and both were detected in (34%) of HCC patients. This high incidence of HCV-Ab may suggest the implication of HCV in the molecular events leading to hepatic carcinogenesis.

Adult↗

[A resources-activities analysis in cardiology: a report on Piemonte experience. The Cardiology Network of the Piemonte Section of the Associazione Nazionale Medici Cardiologi Ospedalieri].

We are living in an era characterized by cost-containment. Thus, expenditure for the health system represents one of the most important items of the national balance. Nowadays, the optimal management of health organization is essential. In addition, patients expect hospitals to provide more and more effective and efficient cures. With respect to this need, in order to achieve high standard of health cure, the identification of work and the time that the medical departments dedicate to a specific therapeutic and/or diagnostic procedure represent a fundamental tool for the optimal management of patients. However, to provide reliable results, these data should be collected in the largest number of structures possible. For this purpose, 17 departments of cardiology have organized a network with the goal of creating the most important data base on this matter available in Italy. The evaluation of the data obtained (more than 8000 medical procedures) allows us to identify the resources normally used for curing patients admitted to cardiological departments. These provide an extensive view of the work load in the Public Health Services fundamental to implementing a health service in accordance with the criteria of Total Quality Management. The present paper illustrates the scope of this study and the methods used to create the data base. In addition, it discusses some cultural aspects related to the introduction of managerial criteria in the medical arena. The detailed results, regarding the items analyzed, will be presented in a special paper.

Cardiology↗

Nuclear medicine in diagnosis and therapy of bone and joint diseases.

Concerning bone and joint diseases therapy of rheumatic synovitis (= radiosynoviorthesis) was introduced in 1952 before clinically relevant diagnostic procedures were developed. Radionuclides of Sr and later on 99mTc phosphonates then started the wide use of bone scintigraphy since > 30 years. The diagnostic methods have an excellent sensitivity for detection of local abnormalities of bone metabolism, the specificity of such studies, however, is low. Modifications of the technique (3-phase-bone-scintigraphy, pinhole collimators, ROI-technique), increasing knowledge of pathological scan patterns and introduction of other radionuclide studies (67Ga, 201Tl, inflammation scans with 99mTc-leukocytes or 99mTc-HIG) as well as 18FDG-PET have increased the specificity significantly in recent years and improvements of imaging systems (SPECT) also increased the accuracy of diagnostic methods in diseases of bone and joints. Therapy of such diseases has made considerable progress: inflamed, swollen joints can effectively be treated with 90Y-, 186Re, 169Er-colloids or with 165Dy-particles by radiosynoviorthesis. Severe pain due to disseminated bone metastases of cancer or polyarthritis can be controlled by radionuclide therapy with 89Sr, 153Sm-EDTMP, 186Re- or 188Re-HEDP and possibly 117mSn-DTPA with an acceptable risk of myelodepression. Possibilities, technical details and limitations of radionuclide applications for diagnostic and therapeutic purposes must be considered if optimal benefit for individual patients should be achieved. Overall Nuclear Medicine can become an essential element in management of bone and joint diseases. The relationship of Nuclear Medicine to bone and joint pathology is peculiar: In 1952 treatment of rheumatic synovitis by radiosynoviorthesis with 198Au Colloid was started by Fellinger and Schmid before diagnostic approaches to bone pathology existed. Bone scintigraphy was introduced only in 1961 using 85Sr but obviously the unfavourable radiation characteristics of this radionuclide limited it's broad application and 87mSr did not improve this situation. Only when 99mTc phosphonates were developed by Subramanian the importance of bone scintigraphy became apparent: The excellent imaging properties of these radiotracers showed, that abnormal bone metabolism could be visualized even before morphological alterations in the skeleton become visible on radiographies or even CT-scans. Moreover, proposals made earlier to use 32P or 89Sr for palliation of pain in patients with disseminated skeletal metastases were picked up again and led also to other radiopharmaceuticals (186Re-HEDP, 153Sm-EDTMP, 117mSn-DTPA) which are applied today for the same purpose with very good success. Therefore Nuclear Medicine today has a broad program for diagnostic and therapeutic approaches to diseases of bone and joints. In bone scanning the high sensitivity led to inclusion of this method for routine staging and re-staging programs in a variety of cancer forms which have a trend to develop bone metastases (e.g. breast, lung, prostate, melanoma) but the low specificity of abnormal patterns on such scans can impair the diagnostic value of the technique. To increase specificity and to define inflammatory lesions, radiotracers used for "inflammation scanning" were introduced such as labeled granulocytes, 99mTc Human Immunoglobulin and others but also a simple modification of bone scanning--triple phase bone scintigraphy--was used. Recently the excellent properties of 18F for PET of the skeleton were rediscovered again and emission CT scanning--possibly with overlay with transmission CT or MRT pictures--can enhance the diagnostic impact of radionuclide bone studies.

Journal Article↗

[CT studies in children. Methods and indications].

Technical advantages of spiral technology are of great usefulness in diagnostic imaging of thorax and abdomen in children. The rapid imaging time allows the examination of an entire organ while scanning with breath holding or while the child is quietly breathing, resulting in a decreased need for sedation. In addition especially examinations of the chest an to some extend CT-investigations of the abdomen are performed with reduced radiation dose achieved by reducing the tube loading. Radiation dose reduction of 70% is possible without loss of relevant diagnostic information. Spiral low-dose CT in pediatric chest imaging is useful in evaluating parenchymal lung diseases, mediastinal masses and central airway malformations. Whether abdominal helical CT can be replaced by MRI has to be decided in each individual case. When an exact protocol is used, sufficient contrast material given, scan-delay optimized and postprocessing reconstructions are performed, useful additional diagnostic informations are obtained by the method.

Adolescent↗

Rapid diagnostic tests for infectious ocular disease.

There is a current and ongoing need for the rapid, accurate diagnosis of ocular infection and the prompt initiation of appropriate therapy to prevent visual loss, maximize use of health care resources, and reduce physicians' medicolegal liability. For the busy clinician, and ideal rapid diagnostic test to determine ocular pathogens would be one whose result is available before the patient leaves the doctor's office, so that optimal treatment can be promptly instituted. Today, the major rapid diagnostic tests available to the busy clinician are the clinical examination and the stained microscopical smear of conjunctival and corneal specimens. The latter technique is grossly underutilized and would contribute significantly to improved accuracy in early diagnosis of ocular infections. In the near future, improved enzyme immunoassays and simplified PCR kits will provide for highly sensitive and specific diagnostic capabilities for detecting ocular pathogens in the laboratory and, possibly, office settings.

Clinical Laboratory Techniques↗

[Evaluation of the diagnostic efficacy of a radioimmunologic assay: the example of thyroid tests].

Optimization of the utilization of a new radioimmunoassay (RIA) requires the identification of explicit criteria regarding when this new RIA should be used. The first step consists in the definition of the normal range on a well defined learning sample of patients. The authors describe the methods which can be used; mean and standard deviation, ROC analysis, Bayes theorem. These limits must be therefore validated on a control sample. The second step of this diagnostic evaluation must estimate the impact of the new RIA on the medical decision procedure. The historical account of thyroid function testing illustrates these different points.

Bayes Theorem↗

New dimensions in cancer biology and therapy.

The most successful and productive approach to defeat cancer relies on highly integrated and interchanging cooperation between basic research, diagnosis, and therapeutic innovation. Nevertheless, much remains to be done to achieve a consistent and continuous flow of theoretical and practical information among scientists actively involved in these equally relevant fields. The major objective of the International Conferences, "New Dimensions in Cancer Biology and Therapy," has been identified in gathering together basic scientists and clinicians who represent scientific leaders in their field, whose working efforts are focused on specific human malignancies. Thus, in pursuit of this well-defined goal, the third edition of the Conference has focused on human immunodeficiency virus (HIV) and cancer, cutaneous melanoma, and colorectal carcinoma, which are ideal clinical examples for innovative diagnostic and therapeutic intervention, as well as optimal models to unveil new mechanisms of tumor pathogenesis.

Colorectal Neoplasms↗

Slide-based cytometry for predicting malignancy in solid salivary gland tumors by fine needle aspirate biopsies.

BACKGROUND: To minimize hospitalization and morbidity for a patient with a solid tumor of a salivary gland, malignancy must be confirmed or excluded as soon as possible. This information cannot be obtained preoperatively by existing standard procedures. Minimal-invasive approaches with adequate diagnostic analysis represent a promising precondition for optimized therapy. METHODS: For fine needle aspirate biopsies (FNABs), laser scanning cytometry (LSC) offers a semi-automated slide-based technology for objective and quantitative analysis. We have established an assay for FNABs from salivary gland tumors. FNAB cells were stained for cytokeratin and DNA followed by LSC analysis. The cells were subsequently HE-stained and were relocalized on the slide. The LSC analysis quantitatively determines the DNA index (DI) of the tumor cells taking leukocytes as internal DNA diploid standard. Histograms with 0.95 < DI < 1.05 and 1.9 2.5 (i.e., 5c exceeding rate, 5cER) was calculated. Samples with DNA aneuploid peaks or with 5cER > 5% were classified as malignant. Routine histopathology was performed as a control. RESULTS: FNABs from 51 solid salivary gland tumors (41 parotid gland, six submandibular, four parapharyngeal) were analyzed with this assay. Eleven of 14 malignant tumors were DNA aneuploid by LSC analysis. All benign tumors showed diploid DNA content. The positive predictive value for malignancy was 1.0, the negative predictive value was 0.93, the correlation with routine histopathology was highly significant (p = 7.6 x 10(-9), Fisher's exact test). The calculated specificity of LSC analysis was 1.0 and the sensitivity was 0.79. CONCLUSIONS: This pilot study demonstrates the validity of slide-based cytometry for the preoperative prediction of malignancy in solid tumors being inaccessible for incision biopsy but suitable for FNABs such as those of the parotid gland.

Adenoma, Pleomorphic↗

Ultrafast T2-weighted imaging of the abdomen and pelvis: use of single shot fast spin-echo imaging.

Single shot (SS) rapid acquisition with relaxation enhancement (RARE) and half Fourier SS-RARE (HFSS-RARE, HASTE, or SS-FSE) sequences allow ultrafast imaging acquisition and generate high imaging quality. Images can be acquired within a very short time, without artifacts from physiologic motion. They are widely applied in the abdominal MRI. Clinical application of the ultrafast SS-RARE imaging techniques provide not only improved temporal resolution but better spatial resolution, higher SNR, and higher tissue contrast. Imaging parameters must be optimized for different MR scanners to obtain diagnostic images.

Abdomen↗

Automatic analysis of heart rate variation: I. Method and reference values in healthy controls.

Many patients referred to an electrophysiological laboratory may have autonomic dysfunction. Some parasympathetic tests are based on the assessment of heart rate variation induced by breathing, Valsalva maneuver, and standing. We have developed fast and practical computer-based methods to analyze heart rate variation using standard EMG equipment and a personal computer. For quantitative description we have evaluated different algorithms, both earlier described and new ones. Findings in patients with diabetes have been compared with those obtained from healthy subjects in order to determine the diagnostic utility of the various algorithms. The optimal algorithm has been chosen by this and other criteria, and a reference database from healthy subjects has been developed.

Adolescent↗

A comparison of Australian and UK obstetricians' and midwives' preferences for screening tests for Down syndrome.

OBJECTIVE: To establish and compare obstetricians' and midwives' preferences for hypothetical prenatal screening tests for Down syndrome. METHODS: A cross-sectional questionnaire survey was completed by 296 obstetricians and midwives at two teaching hospitals: one in Melbourne, Australia (n = 175), and one in London, UK (n = 94). Conjoint analysis was undertaken using random effects probit regression. RESULTS: No significant differences were seen in any measurements when comparing obstetricians in Australia and the UK or midwives in Australia and the UK. Obstetricians and midwives shared similar relative values regarding the importance of the detection rate of the screening tests. However, obstetricians placed higher relative values on both timing of prenatal tests and risk associated with the subsequent diagnostic test than did midwives when considering optimal tests to offer women. Marginal rates of substitution suggest that, compared with midwives, obstetricians would wait longer and accept a greater decrease in detection rate for a test if it was safer. Younger midwives placed higher value on both detection rate and safety of prenatal tests than older midwives. Female obstetricians placed higher value on the timing of a test than male obstetricians. CONCLUSION: Obstetricians in Australia and UK placed almost identical importance on test attributes, as did the midwives in the two countries. However, different attitudes towards tests were seen between obstetricians and midwives.

Adult↗

Idiopathic focal foveolar hyperplasia in infants.

Idiopathic focal foveolar hyperplasia (FFH) is a rare cause of gastric outlet obstruction in infants. We present two cases, including the first reported association with infantile hypertrophic pyloric stenosis. The diagnostic ultrasound appearance of FFH is described. Optimal treatment is surgical excision of the involved mucosa with pyloroplasty or pyloromyotomy.

Diagnosis, Differential↗