Effect of Prior Formal Education on Successful Thoracic Epidural Placement By Anesthesia Residents.

OBJECTIVE: Catheter placement for thoracic epidural analgesia (TEA) is technically challenging; however, methods for teaching this technique to anesthesia residents have not been well-studied. The present study aimed to determine optimal teaching methods for proficient TEA catheter placement by comparing video-based formal resident education with traditional bedside training by attending physicians. DESIGN: Prospective, randomized study. SETTING: Large academic hospital, single institution. PARTICIPANTS: The study comprised 76 postgraduate year 3 and 4 anesthesiology residents (38 intervention, 38 control). INTERVENTIONS: Formal education included an instructional video on proper TEA technique. MEASUREMENTS AND MAIN RESULTS: Measures of proficiency in TEA catheter placement included the time needed to complete the procedure successfully and the success of placement as indicated by patient confirmation. Residents who received formal video instruction had similar success in catheter placement and similar procedure times compared with the traditionally trained residents. The overall success rate was 99.2%, with faculty intervention required in only 17% of cases. More experienced residents (ie, having placed more epidural catheters) were faster at TEA catheter placement. CONCLUSIONS: Formal video education for TEA catheter placement provided no additional improvement of resident proficiency compared with traditional training at a high-volume academic center. The success rate was very high in this group of residents; however, experiences at other institutions may vary. Future studies are needed to determine optimum teaching strategies for TEA.

Novel test method (sickle confirm) to differentiate sickle cell anemia from sickle cell trait for potential use in developing countries.

The objective of this study was to develop a diagnostic testing method to detect HbS, distinguish sickle cell homozygotes from heterozygotes, and overcome testing barriers encountered in laboratories in underdeveloped countries. Blood samples positive and negative for sickle cell were subjected to the standard hemoglobin solubility test followed by a variety of centrifugation and filtration procedures. Each procedure was evaluated for the ability to remove insoluble HbS from the sample. The hemoglobin types that remain (HbA, HbA2 and HbF) were measured spectrophotometrically or estimated visually allowing samples to be categorized into three genotypes (AA, AS and SS) as confirmed by hemoglobin electrophoresis. De-identified EDTA blood samples were obtained from Saint Louis University and Cardinal Glennon Children's hospitals and tested in the Department of Clinical Laboratory Science at Saint Louis University. The main outcome measures were turbidity of the solubility solution; color of the supernatant and the material on the surface of the solution following centrifugation; precipitate trapped on the filter paper; absorbance of the filtrate; and hemoglobin electrophoresis patterns. Centrifugation and filtration successfully separated HbS from HbA/A2/F allowing for the differentiation of seven sickle cell homozygotes from sixteen heterozygotes with a sensitivity and specificity of 100%. This method has the potential to reliably distinguish homozygous from heterozygous sickle cell patients and it is fast, inexpensive, and simple. These characteristics make Sickle Confirm a desirable method in developing countries like Haiti and Africa where sickle cell anemia is prevalent and modern diagnostic methods like electrophoresis, HPLC and nucleic acid testing are impractical.

A method of HbF determination for potential use in underdeveloped countries.

The objective of this study was to develop a simple, cost-effective method of HbF determination potentially useable in underdeveloped countries to determine sickle cell patient response to hydroxyurea treatment. Normal adult blood (HbA), cord blood (HbF), and a 50:50 mixture (HbA+F) were the three sample types used in procedure development. Normal blood samples were collected from the research team, and de-identified cord blood samples were provided by Cardinal Glennon Pediatric Research Institute, St. Louis, MO. The hematocrit of all blood samples was standardized to 35%. The method, based on the Kleihauer-Betke (K-B) test principle, used a citrate solution to selectively elute HbA from RBCs while HbF remained intracellular, and spectrophotometric absorbance of the eluate was the primary outcome measure. A procedure was developed and optimized utilizing a 395 nm wavelength, 30 sec centrifugation time, 6 min incubation time, 20 microL blood volume, and 0.07 M sodium citrate in a 0.06 M sodium phosphate buffer solution. Reproducibility was demonstrated (N = 39) with a mean HbA absorbance of 1.285 (SD 0.069), mean HbA+F absorbance of 0.690 (SD 0.050), and mean HbF absorbance of 0.035 (SD 0.005), also exhibiting linearity (r2 = 0.99). This simple, cost-effective method of HbF determination shows potential as a basis for determining sickle cell patient response to hydroxyurea treatment in underdeveloped countries.

Research in clinical laboratory science: professionals' involvement.

OBJECTIVES: To describe current qualitative and quantitative aspects of research engagement and other scholarly activities conducted by clinical laboratory science (CLS) professionals across a range of employment settings. DESIGN: A link to a 3-part online survey was sent by electronic mail to 7,572 members of the American Society for Clinical Laboratory Science and 500 program directors. SETTING: email message, on-line survey PARTICIPANTS: all ASCLS members and all directors of accredited clinical laboratory educational programs MAIN OUTCOME MEASURES: Quantitative and qualitative measures of professionals' engagement in research and other scholarly activities RESULTS: 556 of 7572 (7.3%) persons completed the survey. Thirty-two percent of survey respondents reported spending between 1 to > 40 work hours per week conducting research with 68% of respondents not participating in research activities. Conducting research is an employment requirement for 18% of survey participants. Twenty-nine percent of respondents have published at least one research article, and 47% of respondents who conduct research have published studies in the journal Clinical Laboratory Science. More than 57% of respondents participate in non-research scholarly activities as part of their employment. CLS professionals who conduct research are more likely to do applied, clinical, or educational research than other types of research. Fifty-seven percent of respondents who conduct research lack external funding for their work. Ninety-three percent of total research dollars is obtained by respondents who hold the Ph.D. degree. The perception of the importance of conducting research varies by employment position. Barriers to participation in research include lack of inclusion of research in the job description, time constraints, inadequate research funding, limited opportunity, and lack of space and equipment. CONCLUSIONS: CLS professionals participate in research in limited numbers, and are more likely to engage in non-research types of scholarly activities. Numerous barriers are identified which impose limits to conducting research. Over half of CLS's research efforts lack external funding. Although there was broad representation among participants across educational levels, employment settings, and job positions, the number of survey respondents was limited. Possible directions for future research include conducting this survey using members of additional professional organizations.

Research in clinical laboratory science: professionals' educational preparation.

OBJECTIVE: To describe the educational preparation of CLS professionals for conducting research. DESIGN: A link to 3-part online survey was sent by electronic mail to 7,572 members of the American Society for Clinical Laboratory Science and 500 program directors research project. Barriers to participation in research by undergraduates include time limitations within the curriculum, insufficient faculty time, and lack of funds, space, and equipment. Increased emphasis on developing research skills is found in educational programs at the master's degree level. CONCLUSIONS: The formal educational background of many CLS professionals may leave them unprepared or underprepared for conducting research. Although there was broad representation among participants across educational levels, employment settings, and job positions, the number of survey respondents was limited. Possible directions for future research include conducting this survey using members of additional professional organizations.

National Institutes of Health Consensus Development Project on Criteria for Clinical Trials in Chronic Graft-versus-Host Disease: IIb. The 2020 Preemptive Therapy Working Group Report.

Chronic graft-versus-host disease (GVHD) commonly occurs after allogeneic hematopoietic cell transplantation (HCT) despite standard prophylactic immune suppression. Intensified universal prophylaxis approaches are effective but risk possible overtreatment and may interfere with the graft-versus-malignancy immune response. Here we summarize conceptual and practical considerations regarding preemptive therapy of chronic GVHD, namely interventions applied after HCT based on evidence that the risk of developing chronic GVHD is higher than previously appreciated. This risk may be anticipated by clinical factors or risk assignment biomarkers or may be indicated by early signs and symptoms of chronic GVHD that do not fully meet National Institutes of Health diagnostic criteria. However, truly preemptive, individualized, and targeted chronic GVHD therapies currently do not exist. In this report, we (1) review current knowledge regarding clinical risk factors for chronic GVHD, (2) review what is known about chronic GVHD risk assignment biomarkers, (3) examine how chronic GVHD pathogenesis intersects with available targeted therapeutic agents, and (4) summarize considerations for preemptive therapy for chronic GVHD, emphasizing trial development, including trial design and statistical considerations. We conclude that robust risk assignment models that accurately predict chronic GVHD after HCT and early-phase preemptive therapy trials represent the most urgent priorities for advancing this novel area of research.

Estimated prevalence of sickle cell in northern Haiti.

OBJECTIVE: The purpose of this study is to estimate the prevalence of sickle hemoglobin in northern Haiti. DESIGN: Sickle cell testing occurred from 2002-2009. Blood samples from 1035 subjects were collected for diagnostic purposes, de-identified, and made available for the study. SETTING: Bethesda Medical Center and Eben-Ezer Clinic in northern Haiti. SUBJECTS: Study subjects included prenatal patients, their companions, clinic staff and volunteers. All subjects were Haitian and selected to most closely represent healthy patients present at the clinic. Deidentification of the blood samples precluded the need for informed consent. MAIN OUTCOME MEASURES: Each subject was tested for sickle hemoglobin using a standard hemoglobin solubility test and results were recorded as either positive or negative. RESULTS: The estimated prevalence of sickle hemoglobin was 15.1% with a 95% confidence interval of 12.2-18%. CONCLUSIONS: These prevalence rates validate the clinical significance of sickle cell disorder, help guide clinical decisions, and suggest the need to develop intervention programs among the people of northern Haiti.

A suite of algorithms for the comprehensive analysis of complex protein mixtures using high-resolution LC-MS.

MOTIVATION: Comparing two or more complex protein mixtures using liquid chromatography mass spectrometry (LC-MS) requires multiple analysis steps to locate and quantitate natural peptides within a single experiment and to align and normalize findings across multiple experiments. RESULTS: We describe msInspect, an open-source application comprising algorithms and visualization tools for the analysis of multiple LC-MS experimental measurements. The platform integrates novel algorithms for detecting signatures of natural peptides within a single LC-MS measurement and combines multiple experimental measurements into a peptide array, which may then be mined using analysis tools traditionally applied to genomic array analysis. The platform supports quantitation by both label-free and isotopic labeling approaches. The software implementation has been designed so that many key components may be easily replaced, making it useful as a workbench for integrating other novel algorithms developed by a growing research community. AVAILABILITY: The msInspect software is distributed freely under an Apache 2.0 license. The software as well as a Zip file with all peptide feature files and scripts needed to generate the tables and figures in this article are available at http://proteomics.fhcrc.org/.

Development of a micro-ESR system with potential for in-home use.

OBJECTIVE: To develop a micro-erythrocyte sedimentation rate (ESR) system with potential to be self-administered by patients at home using capillary blood. DESIGN: For each subject, three tubes of blood were collected in ethylenediaminetetraacetic acid (EDTA), centrifuged, and the cells separated from the plasma. Plasma was pooled and divided into three aliquots, two of which were spiked with defined amounts of fibrinogen creating a normal ESR and two distinct degrees of ESR acceleration. Fifteen hundred microliters of pooled autologous cells were resuspended in 1500 microL of the three prepared autologous plasmas to standardize hematocrit values. ESRs were performed using the Westergren method and four potential micro-ESR systems, utilizing a micro-hematocrit tube, S/P capillary blood gas tube, Natelson blood collecting tube, and Caraway micro blood collecting tube. SETTING: Saint Louis University, St. Louis MO. PATIENTS/SAMPLES: Twenty-eight volunteers between the ages of 18 and 60 years with no underlying conditions participated in the study. INTERVENTIONS: Hematocrit was standardized to approximately 40% for all samples and fibrinogen concentrations of approximately 200 mg/dL, 382 mg/dL, and 563 mg/dL were achieved for each subject. MAIN OUTCOME MEASURES: ESRs were measured in mm/hour and in percentage. Micro-ESR values were plotted versus the paired Westergren ESR value and the data were analyzed using Pearson's r correlation. RESULTS: When compared to the Westergren ESR method, the following correlation coefficients were achieved: S/P tube (r = 0.808), Caraway tube (r = 0.797), Natelson tube (r = 0.719), and micro-hematocrit tube (r = 0.655). CONCLUSION: Three of the four micro-ESR methods achieved correlation coefficients acceptable to the authors, with the potential of being converted into a capillary ESR system for in-home use.

Chronic myelocytic leukemia--Part I: History, clinical presentation, and molecular biology.

DATA SYNTHESIS: Chronic myelocytic leukemia (CML) was initially described in 1845 and is considered one of the first leukemias to be discovered. Diagnosis of CML was dramatically improved with the discovery of the Philadelphia chromosome by Nowell and Hungerford in 1960. However, the rudiments of our understanding of the molecular cause of CML began in 1973 when Janet Rowley discovered that the Philadelphia chromosome is a reciprocal translocation between chromosomes 9 and 22. The leukemogenic mechanisms of CML were hypothesized 20 years later when it was discovered that the t(9;22) translocation produced a fusion gene involving the BCR gene from chromosome 22 and the ABL protooncogene from chromosome 9 [corrected] Multiple breakpoints in BCR produce fusion genes that are translated into chimeric protein products of different lengths that are associated with different leukemic subtypes. CONCLUSION: Although CML has a rich history of interest to hematologists, it also represents a leukemogenic paradigm to the molecular biologist. Nearly all malignancies result from a series of mutagenic events, which culminate in full malignant transformation. However, it appears that CML results from a single mutagenic event involving the t(9;22) translocation leading to the development of the BCR/ABL fusion gene and the corresponding fusion protein. The successful transcription and translation of the BCR/ABL fusion protein led researchers to carefully study its involvement in leukemogenesis. The BCR/ABL fusion protein exhibits increased and constitutive tyrosine kinase activity that differs depending on which BCR breakpoint is expressed, resulting in varying clinical presentations.

Chronic myelocytic leukemia--Part II: Approaches to and molecular monitoring of therapy.

DATA SYNTHESIS: Chronic myelocytic leukemia (CML) was initially described in 1845 and is considered one of the first leukemias discovered. Effective approaches to therapy were not instituted until arsenic was first administered in 1865. Since then, four major therapeutic milestones have been achieved; the development of alkylating agents like busulphan and 6-thioguanine in 1953, alpha interferon in 1983, bone marrow transplantation in 1986, and tyrosine kinase inhibitors in 1998. The discovery that the protein product of this fusion gene expresses constitutive tyrosine kinase activity prompted the synthesis of a designer drug, imatinib mesylate, which binds the fusion protein and neutralizes the tyrosine kinase activity. Molecular methods of detecting BCR-ABL transcripts are showing promise in confirming drug resistance and predicting patient outcomes in response to imatinib mesylate therapy. Evidence of drug resistance can guide physicians in selecting alternative therapeutic approaches early in the course of the disease to potentially rescue non responders. Although the success of clinical trials has been dramatic, drug resistance and disease relapse are issues to be considered. CONCLUSION: The discovery that the BCR/ABL fusion protein exhibits increased and constitutive tyrosine kinase activity led investigators to develop an inhibitor to this activity. The synthesis of imatinib mesylate, currently marketed as Gleevec(TM) or Glivec(R), is in stage III clinical trials and has proven to be the most successful antileukemic drug to date. As in CML, an understanding of the leukemogenic mechanisms involved in other leukemias will provide the groundwork for the development of therapeutic interventions tailored to the specific molecular defects identified, eventually rendering obsolete the shotgun approaches to massive cell killing produced by chemotherapy.

Advances in acute lymphoblastic leukemia.

DATA SOURCES: Current literature. DATA SYNTHESIS: Acute lymphoblastic leukemia (ALL) is a stem cell disorder characterized by an overproduction of lymphoblasts in the bone marrow that eventually spill into circulation, producing lymphocytosis. As with the other acute leukemias, the most common symptoms experienced by patients include fatigue, bleeding, and recurrent infections resulting from the suppression of normal hematopoiesis in the bone marrow by the accumulating blasts. ALL primarily affects children and exhibits the best response to standard chemotherapy as compared to acute myeloblastic leukemias (AML). Further, remission rates are highest among ALL patients, many of whom are experiencing sustained remissions suggesting cure. In light of early treatment successes, researchers began to investigate modifications of standard treatment regimens to accommodate variability in weight, age, and response to therapy among children with ALL. Individualized treatment plans were implemented where some patients received a reduced intensity course of therapy to minimize drug toxicity while others received drug intensification to maximize response. More recently, research efforts have been directed at the elucidation of leukemogenic mechanisms implicated in ALL to identify specific protein mutants that can be used to design drugs tailored to interfere with the activity of these mutant protein targets. Identification of chimeric proteins produced from chromosomal translocations and gene expression profiles from microarray analyses are the primary techniques used to identify the potential therapeutic targets. CONCLUSION: Several reliable prognostic indicators have been identified and are being used to improve therapeutic planning and outcome prediction in ALL patients. Individualized treatment regimens have been developed based on the specific characteristics of each patient to minimize treatment related adverse events and maximize response. Through the use of cytogenetic, molecular, and microarray testing, ALL classification schemes have improved and potential therapeutic targets have been identified. It is anticipated that the next major advance in the treatment of ALL will involve the use of designer therapies developed to specifically interfere with particular molecular abnormalities producing the leukemogenic aberration to the normal signal transduction pathways.

The minimum concentration of fibrinogen needed for platelet aggregation using ADP.

OBJECTIVE: Determine the minimum concentration of plasma fibrinogen needed to stimulate the aggregation of platelets, collected from normal subjects, using ADP. DESIGN: Platelet rich plasmas (300 x 10(9) platelets/L) were made and adjusted to final fibrinogen concentrations of 75, 19, 5, and 0 mg/dL using fibrinogen free serum. Each fibrinogen concentration in all twelve subjects was aggregated with ADP SETTING: Research laboratory in the Department of Clinical Laboratory Science at Saint Louis University. PARTICIPANTS: Twelve healthy volunteers of both genders, between the ages of 18 and 60 years who were not pregnant and weighed at least 110 pounds were included in the study. Subjects were excluded from the study if they had ingested aspirin within one week prior to blood collection. In addition, subjects with a history of bleeding disorders such as afibrinogenemia, hypofibrinogenemia, von Willebrand disease, and Bernand-Soulier disease were rejected from the study. MAIN OUTCOME MEASURES: Platelet aggregation tracings were analyzed for amplitude and compared across plasma fibrinogen concentrations. In addition, the type of curve (monophasic vs. biphasic), smoothness and aggregation stability were also noted. RESULTS: The results show that aggregation occurred with every dilution of fibrinogen tested and that the amplitude of the aggregation curves appears not to be dependent on plasma fibrinogen. CONCLUSIONS: The results indicate that platelets from healthy individuals previously exposed to normal fibrinogen levels will aggregate equally well in decreasing plasma fibrinogen concentrations and even in the absence of plasma fibrinogen using ADP as the aggregator.

Ultrasensitive detection and characterization of posttranslational modifications using surface-enhanced Raman spectroscopy.

Posttranslational modification (PTM) of proteins is likely to be the most common mechanism of altering the expression of genetic information. It is essential to characterize PTMs to establish a complete understanding of the activities of proteins. Here, we present a sensitive detection method using surface-enhanced Raman spectroscopy (SERS) that can detect PTMs from as little as zeptomoles of peptide. We demonstrate, using model peptides, the ability of SERS to detect a variety of protein modifications, such as acetylation, trimethylation, phosphorylation, and ubiquitination. In addition, we show the capability to obtain positional information for modifications such as trimethylation and phosphorylation using SERS and wavelet decomposition data analysis techniques. We further show that it is possible to apply SERS to detect PTMs from biological samples such as histones. We envision that this detection method might be a valuable technique that is complementary to mass spectrometry in obtaining orthogonal chemical and modification-specific information from biological samples at sensitive levels.

Denoising array-based comparative genomic hybridization data using wavelets.

Array-based comparative genomic hybridization (array-CGH) provides a high-throughput, high-resolution method to measure relative changes in DNA copy number simultaneously at thousands of genomic loci. Typically, these measurements are reported and displayed linearly on chromosome maps, and gains and losses are detected as deviations from normal diploid cells. We propose that one may consider denoising the data to uncover the true copy number changes before drawing inferences on the patterns of aberrations in the samples. Nonparametric techniques are particularly suitable for data denoising as they do not impose a parametric model in finding structures in the data. In this paper, we employ wavelets to denoise the data as wavelets have sound theoretical properties and a fast computational algorithm, and are particularly well suited for handling the abrupt changes seen in array-CGH data. A simulation study shows that denoising data prior to testing can achieve greater power in detecting the aberrant spot than using the raw data without denoising. Finally, we illustrate the method on two array-CGH data sets.