Rapid Ascertainment Research Articles

Lung biopsy in the diagnosis and management of chILD.

Children's interstitial and diffuse lung disease (chILD) comprises a large number of diverse entities ranging from disorders of lung development, maturation and function unique in infancy to immune-mediated, environmental, vascular and other conditions overlapping with adult disease. Pathologic evaluation of the lung has played a central role in characterizing many of these disorders, resulting in revised nomenclature and classifications to help guide clinical management(1-4). Technological advancements are rapidly uncovering genetic and molecular underpinnings of these conditions, as well as widening the phenotypes which bridge adult disease, often reducing the perceived need for diagnostic lung biopsy. As such the decision to get a lung biopsy in chILD is frequently for rapid ascertainment of disease in a critically ill child or when clinical presentation, imaging and laboratory studies fail to provide a cohesive diagnosis needed for treatment. While there have been modifications in surgical procedures for lung biopsy that minimize postoperative morbidity, it remains a high-risk invasive procedure, especially in a medically complex patient(5). Thus, it is essential that the lung biopsy be handled properly to maximize diagnostic yield, including close communication between the clinician, radiologist, surgeon, and pathologist before biopsy to determine best sampling site(s) and prioritization of tissue utilization. This review provides an overview of optimal handling and evaluation of a surgical lung biopsy for suspected chILD, with emphasis on specific conditions in which pathologic features play a critical role in providing an integrated diagnosis and guiding management.

Testicular cancer management: Population-wide, rapid case ascertainment to drive early expert engagement and reduced practice variation.

39 Background: Research priorities in germ cell tumor (GCT) management have moved sharply from therapeutic improvements to cancer care delivery research and biomarker-based decision making. Early intervention with centralized decision support and oversight by expert teams result in best therapeutic outcomes and survivorship with decreased resource utilization. We describe Kaiser Permanente Northern California’s (KPNC) re-organization of care delivery through rapid case ascertainment and early expert input, as well as early results of reduction in practice variation and system-wide practice change. Methods: In 2016, KPNC reorganized oncology from a distributed generalist model to a model led by a centralized multidisciplinary expert team to share in initial and ongoing care delivery for all GCT patients in the system. Central to the re-organization was rapid ascertainment of the entire population of patients with GCT within the system and early expert engagement in treatment decision-making. Results: Between May 2016 and June 2018, 274 GCT patients were recorded in the tumor registry, of whom 69% were < 40 years of age, 16% were non-white, 56% had seminoma and 63% had stage 1 disease. Rapid case ascertainment identified 89% (95% CI, 86-93%) of the cases, increasing from 79% in 2016 to 97% in 2018 as false negatives were identified and used to improve the case finding algorithm. The overall positive predictive value was 57% (52-62%) and number needed to detect was 1.75 (1.62-1.91). Of the 274 cases, 92% (89-95%) were engaged by the expert team. In addition, the team reviewed 61 testicular cancer patients who had recurrences or metastatic cancers. Among 177 patients with stage I seminoma, the preferred use of active surveillance over adjuvant chemotherapy or radiation therapy rose from 48% (95% CI, 35-62%) in 2015 to 87% (75-99%) in 2018 (p = 0.0005). For patients with nonseminoma, the rate of the preferred option of retroperitoneal lymphadenectomies being performed by a high volume urologic surgeon increased markedly from 62% in 2015 to 95% in 2018. Conclusions: To our knowledge, the KPNC re-organization of GCT care delivery with comprehensive rapid case ascertainment is unique for integrated health care delivery systems in the USA. While early, KPNC has a working platform for early, expert multidisciplinary review and bidirectional communication with local care teams for population-based care. Early evidence points to system-wide reductions in practice variation and improvements in practice.

Systematic identification of facility-based stillbirths and neonatal deaths through the piloted use of an adapted RAPID tool in Liberia and Nepal.

Maternal, fetal, and neonatal health outcomes are interdependent. Designing public health strategies that link fetal and neonatal outcomes with maternal outcomes is necessary in order to successfully reduce perinatal and neonatal mortality, particularly in low- and middle- income countries. However, to date, there has been no standardized method for documenting, reporting, and reviewing facility-based stillbirths and neonatal deaths that links to maternal health outcomes would enable a more comprehensive understanding of the burden and determinants of poor fetal and neonatal outcomes. We developed and pilot-tested an adapted RAPID tool, Perinatal-Neonatal Rapid Ascertainment Process for Institutional Deaths (PN RAPID), to systematically identify and quantify facility-based stillbirths and neonatal deaths and link them to maternal health factors in two countries: Liberia and Nepal. This study found an absence of stillbirth timing documented in records, a high proportion of neonatal deaths occurring within the first 24 hours, and an absence of documentation of pregnancy-related and maternal factors that might be associated with fetal and neonatal outcomes. The use of an adapted RAPID methodology and tools was limited by these data gaps, highlighting the need for concurrent strengthening of death documentation through training and standardized record templates.

Use of Rapid Ascertainment Process for Institutional Deaths (RAPID) to identify pregnancy-related deaths in tertiary-care obstetric hospitals in three departments in Haiti

BackgroundAccurate assessment of maternal deaths is difficult in countries lacking standardized data sources for their review. As a first step to investigate suspected maternal deaths, WHO suggests surveillance of “pregnancy-related deaths”, defined as deaths of women while pregnant or within 42 days of termination of pregnancy, irrespective of cause. Rapid Ascertainment Process for Institutional Deaths (RAPID), a surveillance tool, retrospectively identifies pregnancy-related deaths occurring in health facilities that may be missed by routine surveillance to assess gaps in reporting these deaths.MethodsWe used RAPID to review pregnancy-related deaths in six tertiary obstetric care facilities in three departments in Haiti. We reviewed registers and medical dossiers of deaths among women of reproductive age occurring in 2014 and 2015 from all wards, along with any additional available dossiers of deaths not appearing in registers, to capture pregnancy status, suspected cause of death, and timing of death in relation to the pregnancy. We used capture-recapture analyses to estimate the true number of in-hospital pregnancy-related deaths in these facilities.ResultsAmong 373 deaths of women of reproductive age, we found 111 pregnancy-related deaths, 25.2% more than were reported through routine surveillance, and 22.5% of which were misclassified as non-pregnancy-related. Hemorrhage (27.0%) and hypertensive disorders (18.0%) were the most common categories of suspected causes of death, and deaths after termination of pregnancy were statistically significantly more common than deaths during pregnancy or delivery. Data were missing at multiple levels: 210 deaths had an undetermined pregnancy status, 48.7% of pregnancy-related deaths lacked specific information about timing of death in relation to the pregnancy, and capture-recapture analyses in three hospitals suggested that approximately one-quarter of pregnancy-related deaths were not captured by RAPID or routine surveillance.ConclusionsAcross six tertiary obstetric care facilities in Haiti, RAPID identified unreported pregnancy-related deaths, and showed that missing data was a widespread problem. RAPID is a useful tool to more completely identify facility-based pregnancy-related deaths, but its repeated use would require a concomitant effort to systematically improve documentation of clinical findings in medical records. Limitations of RAPID demonstrate the need to use it alongside other tools to more accurately measure and address maternal mortality.

Barth Syndrome: An Under-Recognized Cause of Chronic Neutropenia

▪Barth syndrome is an X-linked disease characterized by defective remodeling of phospholipid side chains in mitochondrial membranes. Major features include neutropenia, fetal or childhood onset dilated cardiomyopathy, motor delay and proximal myopathy, feeding problems and constitutional growth delay. Improved diagnostic testing, recognition of the wide disease phenotype and a worldwide patient/family support network have recently facilitated much more rapid ascertainment, now with at least 186 living males diagnosed worldwide. We provide a comprehensive survey of neutropenia seen in Barth syndrome and its management based on the study of 90 patients. Neutropenia (i.e. at least one count less than 1.5 x 109/L) was detected in 74 of 90 (82%) patients; 28 of 90 (31%) patients had a mean neutrophil count less than 1.5 x 109/L; 13 of 90 (14%) had a mean count less than 0.5 x 109/L. Neutropenia may be the sole presenting feature in Barth syndrome. Importantly, it can take on any form: intermittent and unpredictable, chronically severe or cyclical, thus mimicking other diseases. Eighty-seven patients had reported monocyte data. Monocytosis was seen in 65 of 87 (75%) patients with at least one monocyte count greater than 1.0 x 109/L and 23 of 87 (26%) had at least one monocyte count greater than 3.0 x 109/L.Some patients who were neutropenic and had a bone marrow evaluation showed myelocyte arrest. Neutropenic Barth patients are highly responsive to granulocyte colony-stimulating factor, although dosing can be challenging because of innate variations in their neutrophil counts. Clinical improvement with reduced signs and symptoms of infections is the usual response to this treatment. Barth syndrome should be considered in any male with neutropenia accompanied by any of the characteristic features of Barth syndrome and in those with idiopathic neutropenia. DisclosuresDale:Amgen: Consultancy, Honoraria, Research Funding.

Establishing a clinic-based pancreatic cancer and periampullary tumour research registry in Quebec.

Enrolling patients in studies of pancreatic ductal adenocarcinoma (pdac) is challenging because of the high fatality of the disease. We hypothesized that a prospective clinic-based study with rapid ascertainment would result in high participation rates. Using that strategy, we established the Quebec Pancreas Cancer Study (qpcs) to investigate the genetics and causes of pdac and other periampullary tumours (pats) that are also rare and underrepresented in research studies. Patients diagnosed with pdac or pat were introduced to the study at their initial clinical encounter, with a strategy to enrol participants within 2 weeks of diagnosis. Patient self-referrals and referrals of unaffected individuals with an increased risk of pdac were also accepted. Family histories, epidemiologic and clinical data, and biospecimens were collected. Additional relatives were enrolled in families at increased genetic risk. The first 346 completed referrals led to 306 probands being enrolled, including 190 probands affected with pdac, who represent the population focus of the qpcs. Participation rates were 88.4% for all referrals and 89.2% for pdac referrals. Family history, epidemiologic and clinical data, and biospecimens were ascertained from 91.9%, 54.6%, and 97.5% respectively of patients with pdac. Although demographics and trends in risk factors in our patients were consistent with published statistics for patients with pdac, the qpcs is enriched for families with French-Canadian ancestry (37.4%), a population with recurrent germ-line mutations in hereditary diseases. Using rapid ascertainment, a pdac and pat research registry with high participation rates can be established. The qpcs is a valuable research resource and its enrichment with patients of French-Canadian ancestry provides a unique opportunity for studies of heredity in these diseases.

Trends in HCV RNA testing among HCV antibody-positive persons in care, 2003-2010.

A test for hepatitis C virus (HCV) RNA is essential to identify persons with active, or current, HCV infection. We assessed trends in HCV RNA testing following a positive HCV antibody result among persons in 4 large healthcare organizations. Data collected from adults with ≥2 clinical encounters during 2003-2010 were analyzed to determine the frequency of, interval between, and factors associated with having an RNA test after a first positive HCV antibody test. From 2003-2010, 5860 persons had a positive antibody test, of whom 3570 (60.9%) had a follow-up RNA test. During this period, the annual frequency of persons with an eventual RNA test did not change significantly; however, the fraction of persons who had the follow-up RNA test within 6 months improved significantly, from 45% in 2003 to 57% in 2010 (P < .001, for trend). Persons born during 1945-1965, men, and those with annual income <$30 000 (by census geocode) were less likely to have had a follow-up RNA test done within 6 months of a positive antibody test. Less than two-thirds of persons with a positive HCV antibody test had a follow-up RNA test. Rapid ascertainment of HCV infection status with reflex testing to RNA is critical to identify persons eligible for treatment.

ONLINE REPORT IN REAL TIME FROM THE NATIONAL PROSTATE CANCER REGISTER (NPCR) OF SWEDEN, A NEW AND EFFICIENT TOOL FOR QUALITY ASSURANCE

IntroductionTraditionally data in The Swedish Cancer register, The Patient Register and other health care registers have been characterized by a very high capture rate but a very long time between...

Measurement of Phospholipids May Improve Diagnostic Accuracy in Ovarian Cancer

BackgroundMore than two-thirds of women who undergo surgery for suspected ovarian neoplasm do not have cancer. Our previous results suggest phospholipids as potential biomarkers of ovarian cancer. In this study, we measured the serum levels of multiple phospholipids among women undergoing surgery for suspected ovarian cancer to identify biomarkers that better predict whether an ovarian mass is malignant.Methodology/Principal FindingsWe obtained serum samples preoperatively from women with suspected ovarian cancer enrolled through a prospective, population-based rapid ascertainment system. Samples were analyzed from all women in whom a diagnosis of epithelial ovarian cancer (EOC) was confirmed and from benign disease cases randomly selected from the remaining (non-EOC) samples. We measured biologically relevant phospholipids using liquid chromatography/electrospray ionization mass spectrometry. We applied a powerful statistical and machine learning approach, Hybrid huberized support vector machine (HH-SVM) to prioritize phospholipids to enter the biomarker models, and used cross-validation to obtain conservative estimates of classification error rates.ResultsThe HH-SVM model using the measurements of specific combinations of phospholipids supplements clinical CA125 measurement and improves diagnostic accuracy. Specifically, the measurement of phospholipids improved sensitivity (identification of cases with preoperative CA125 levels below 35) among two types of cases in which CA125 performance is historically poor - early stage cases and those of mucinous histology. Measurement of phospholipids improved the identification of early stage cases from 65% (based on CA125) to 82%, and mucinous cases from 44% to 88%.Conclusions/SignificanceLevels of specific serum phospholipids differ between women with ovarian cancer and those with benign conditions. If validated by independent studies in the future, these biomarkers may serve as an adjunct at the time of clinical presentation, to distinguish between women with ovarian cancer and those with benign conditions with shared symptoms and features.

Feasibility of Institutional Registry-Based Recruitment for Enrolling Newly Diagnosed Breast Cancer Patients in an Exercise Trial

The purpose of this study was to determine the feasibility of using a tumor registry to recruit newly diagnosed survivors into a randomized controlled exercise trial and to discuss issues related to this recruitment strategy. A tumor registry-based rapid ascertainment system was used to recruit breast cancer survivors into a 6-month home-based, telephone-administered intervention of moderate-to-vigorous intensity exercise or a usual care group. 468 newly diagnosed cases were identified. Of these, 50 women (15.4% of those for which screening calls were made) were enrolled in the study. Women were randomized, on average, 11 weeks after diagnosis (SD = 4.8). Sixty-four percent were randomized before beginning treatment or within the first week of treatment. Time required to obtain physician consent was the primary determinant of diagnosis-to-randomization latency. Enrolled women were more likely than nonenrolled women to be non-Hispanic White and to have a college degree (P < .05). Tumor registries present a feasible means of recruiting breast cancer survivors before or early in adjuvant treatment. The success of recruiting survivors promptly after diagnosis is largely dependent on ability to rapidly obtain physician consent. Specific effort is needed to counteract self-selection effects that may lead to under-representation of minorities.

Rapid screening for the detection of HLA‐B57 and HLA‐B58 in prevention of drug hypersensitivity

HLA-B57 and HLA-B58 are major histocompatibility class (MHC)-I allotypes that are potentially predictive of important clinical immune phenotypes. HLA-B*5701 is strongly associated with hypersensitivity to the HIV drug abacavir, liver toxicity from the antibiotic flucloxacillin and is a marker for slow progression of HIV AIDS. HLA-B*5801 is associated with hypersensitivity to allopurinol used to treat hyperuricaemia and recurrent gout. Here we describe a monoclonal antibody (mAb) specific for HLA-B57 and HLA-B58 that provides an inexpensive and sensitive screen for these MHC-I allotypes. The usefulness of HLA-B57 screening for prediction of abacavir hypersensitivity was shown in three independent laboratories, including confirmation of the mAb sensitivity and specificity in a cohort of patients enrolled in the PREDICT-1 trial. Our data show that patients who test negative by mAb screening comprise 90%-95% of all individuals in most human populations and require no further human leukocyte antigen (HLA) typing. Patients who test positive by mAb screening should proceed to high-resolution typing to ascertain the presence of HLA-B*5701 or HLA-B*5801. Hence, mAb screening provides a low-cost alternative to high-resolution typing of all patients and lends itself to point-of-care diagnostics and rapid ascertainment of low-risk patients who can begin immediate therapy with abacavir, flucloxacillin or allopurinol.

PS2-01: A System for Computer-Assisted Rapid Case Ascertainment of Breast, Lung and Colorectal Cancers

result in recruitable subjects. The proposed presentation will describe our use of text classifiers as part of an efficient computer-assisted recruitment pipeline. Methods: We gathered corpora of pathology reports for breast, lung and colorectal tissue, each with a ‘gold standard’ assessment of whether they discussed a malignancy. We then randomly divided each into 75% training and 25% evaluation subsamples, trained the classifier on the training subsample and compared its categorizations to the gold standard in the evaluation subsample. We performed this training/evaluation cycle repeatedly, generating a distribution of predictive value statistics, thereby getting insight into the classifiers’ sensitivity to the particular random division of corpus reports. More importantly, it allowed us to experiment with tweaks to the classifiers and to the basic text processing that preceded them, and quickly evaluate whether they contributed to the predictive power of the classifier. Results: Several enhancements contributed to the accuracy of the system. In particular, taking multi-word phrases in addition to individual words as features increased the classifier’s predictive power. To our surprise, ‘stemming’ individual words (that is, reducing them to their root forms) did not have a perceptible effect. Once the classifiers were optimized, we were able to cover approximately 1600 path reports/week in approximately 30 minutes of a Research Specialist’s time. As an added benefit, when the RS found that a report was flagged in error, the classifier was trained on that report, thereby improving its future performance. Conclusions: Text classifiers are an effective tool for optimizing the use of staff time in rapidly ascertaining cancer cases for recruitment. Furthermore, because the classifiers’ training (including the corrections made during the study) is easily reduced to a file on disk, it becomes an independent asset, useful for future studies needing to do rapid ascertainment of cancer.

The role of age and race in receiving breast cancer treatment: A cross-sectional survey of an urban population

6047 Background: Breast cancer treatment differences in older patients and in certain groups. These differences may be due to a variety of factors, in addition to age and race, which include: comorbid illnesses, performance status, socioeconomic status, and education. Methods: The sample consisted of a cross-sectional survey of 257 Latina, African American and White women living in Los Angeles County, age 55 or older from the Los Angeles County Cancer Surveillance Program’s (CSP) rapid ascertainment program (the Tumor Registry) who were between three and nine months of diagnosis and at least one month post treatment. These patients were interviewed, their medical records abstracted, and the doctor’s surveyed. The primary outcomes for this analysis were surgical, radiation, hormone, and chemotherapy treatment options. Logistic regression analysis was used controlling for age, race/ethnicity, socio-demographic characteristics, Katz comorbidity, functional status, and stage of disease. Results: Univariate analysis demonstrated significant trends with age and race across most treatment choices. In multivariate analysis, the effect of age and race was less pronounced. Factors such as insurance status, comorbidity, and education were very significant for treatment selection. Medi-Cal patients were less likely to receive lumpectomy (Odds Ratio, OR=0.19) and none received reconstruction post mastectomy. Educated patients were more likely to have an axillary lymph node dissection, LND, (OR=7.5) and post-lumpectomy radiation (OR=3.94). Those with higher Katz comorbidity were less likely to receive a LND (OR=0.70) or chemotherapy (OR=0.76). Nevertheless, older women were less likely to have LND (OR=0.49) and chemotherapy (OR=0.24) compared to younger women. African-American women were less likely to have adjuvant hormone therapy (OR=0.50) compared to Caucasians and Latinas. Conclusion: Although age and race are factors in certain breast cancer treatment decisions, other factors such as education and insurance status and may serve as important targets in order to improve care. No significant financial relationships to disclose.

Serum levels of sex hormones and breast cancer risk in premenopausal women: a case-control study (USA).

High levels of serum estrogens and androgens have been convincingly linked with an increased risk of breast cancer among postmenopausal women. By contrast, the role of blood levels of these hormones in the etiology of premenopausal breast cancer is not well understood. In a case-control study, we sought to examine associations between levels of serum estradiol, sex-hormone binding globulin (SHBG), dehydroepiandrosterone (DHEA), testosterone, androstenedione and progesterone and risk of premenopausal breast cancer. Cases of breast cancer under age 45 were identified using rapid ascertainment systems in Seattle/Puget Sound, Washington and control subjects were identified from the same area through random digit dialing methods. A total of 169 eligible breast cancer cases and 195 control subjects donated blood (either before or six or more weeks after surgery) and were interviewed using a standardized questionnaire. The fully adjusted risk ratios and 95% confidence intervals for the highest versus lowest tertiles of estradiol, according to menstrual cycle phase, were 3.10 (0.8-12.7) for early follicular, 0.54 (0.2-1.7) for late follicular and 0.60 (0.3-1.4) for luteal. Risks for highest versus lowest quartiles of SHBG and androgens were 0.81 (0.4-1.6) for SHBG, 2.42 (1.1-5.2) for DHEA, 1.12 (0.6-2.5) for testosterone, and 1.33 (0.6-2.8) for androstenedione. For luteal progesterone, the RR for the highest versus lowest tertile was 0.55 (0.2-1.4). In summary, we did not find a convincing association between serum SHBG, estradiol, testosterone or androstenedione and premenopausal breast cancer risk. Observed differences between cases and controls subjects in serum levels of DHEA and luteal phase progesterone should be investigated further in large prospective studies.

Study Design for a Case Control Investigation of Cellular Telephones and Other Risk Factors for Brain Tumors in Adults

The aetiology of brain tumours is poorly understood. Due, in part, to public concern about a postulated relationship between the use of cellular telephones or other increasingly prevalent environmental exposures and the incidence of brain cancer in adults, the National Cancer Institute is collaborating with three US hospitals in a comprehensive case-control study of malignant and benign brain tumours. Factors under consideration include use of cellular phones and other wireless communication devices, workplace exposures to chemical agents and electromagnetic fields, dietary factors, family history of tumours, genetic determinants of susceptibility, home appliance use, reproductive history and hormonal exposures, viruses, medical and dental exposure to ionising radiation, and other aspects of medical history. Approximately 800 newly diagnosed brain tumour cases and 800 controls were enrolled at hospitals in Boston, Phoenix and Pittsburgh from 1994 to 1998. Cases include all adults (age ≥ 18 y) newly diagnosed with a histologically confirmed intracranial glioma, histologically confirmed intracranial meningioma or acoustic neuroma. Controls are patients admitted to the same hospitals as the eases, and treated for any of a variety of non-malignant conditions. Key features of the study include its large size, the emphasis on rapid ascertainment of incident cases and interview of study subjects rather than surrogate respondents, the use of detailed, job-specific questions developed by industrial hygienists to ascertain occupational exposures, and the storage of blood samples for future evaluation of inherited susceptibility, biomarkers of exposure and gene-environment interactions.

変異蛋白，翻訳後修飾蛋白のＭＳによる構造解析

Progress in medicine and molecular research has been increased by the need for simple and reliable methods of detection and characterization of aberrant proteins caused either by mutation or by post-translational modification. We applied electrospray ionization mass spectrometry (ESIMS) to detect and characterize abnormal structure proteins, and to determine the ratio of mutant and wild type proteins. Direct examination of hemolysate might well lead to rapid ascertainment of variant hemoglobins (Hbs) provided that the mass difference between the normal and the abnormal chain is larger than the resolution power of standard instruments. Various procedures are required in the presentation of most plasma and cell proteins other than Hb prior to MS. Two simple methods were deviced to prepare proteins. One was immunoprecipitation with antisera against target proteins, followed by reversed-phase HPLC/ESIMS, and the other was 2-dimensional LC (a strong anion exchange region and a reversed-phase resin) connected to ESIMS. With both methods, the ions of intact normal and variant proteins were clearly observed in samples from patients with neurodegenerative diseases such as familial amyloidotic polyneuropathy (FAP) and amyotrophic lateral screlosis (FALS). Using these procedures, we detected more than 40 cases of transthyretins (TTRs) mutants including to 10 different types. Two of these, [101 Gly->Ser] and [38 Asp->Ala], were new. We also successfully detected 4 different mutants of Cu/Zn-binding superoxide dismutase inerythrocytes, and a variant in spinal cord from patients with FALS by immunoprecipitation mehods. Two of these, [4Ala->Ser] and [111Cys->Tyr], were new. Finally we also measured glycated β-globin N-ternimus hexapeptide (HbA1c) by Poroszyme V8 protease digestion and nanoESIMS technique.

電気泳動と質量分析の併用 変異蛋白質同定法のレベルアップ

Direct examination of hemolysate by electrospray ionization mass spectrometry may lead to the rapid ascertainment of variant hemoglobins (Hb) and to the quantification of glycated Hb. Plasma and cell proteins other than Hb can be prepared by immunoprecipitation prior to MS. The detected variants were sequenced by MS using enzyme cleaved peptides. We detected various variants of Hb, transthyretin, and CuZn-superoxide dismutase. New variants included; 1) Hb Sagami [β139 (H17) Asn→Thr], and 2) Hb Hokusetsu [β52 (D3) Asp→Gly]. These were studied to clarify the cause of unusual profiles of HPLC for HbAlc measurement. Hb Sagami was an electrophoretically silent and isopropanol test-negative variant, detected solely by ESIMS. Other new variants included; 3) a variant transthyretin, amyloidogenic, [38 Asp→Ala], and 4) another variant transthyretin, non-amyloidogenic, [101 Gly→Ser]. Variant transthyretins are usually difficult to detect by electrophoresis. Samples containing variant Hb or high carbamylated Hb gave erroneous results of glycated Hb by routine HPLC and immunoassay, but did not by MS.

Simultaneous assessment of loss of heterozygosity at multiple microsatellite loci using semi-automated fluorescence-based detection: subregional mapping of chromosome 4 in cervical carcinoma.

Detection of loss of heterozygosity (LOH) by comparison of normal and tumor genotypes using PCR-based microsatellite loci provides considerable advantages over traditional Southern blotting-based approaches. However, current methodologies are limited by several factors, including the numbers of loci that can be evaluated for LOH in a single experiment, the discrimination of true alleles versus "stutter bands," and the use of radionucleotides in detecting PCR products. Here we describe methods for high throughput simultaneous assessment of LOH at multiple loci in human tumors; these methods rely on the detection of amplified microsatellite loci by fluorescence-based DNA sequencing technology. Data generated by this approach are processed by several computer software programs that enable the automated linear quantitation and calculation of allelic ratios, allowing rapid ascertainment of LOH. As a test of this approach, genotypes at a series of loci on chromosome 4 were determined for 58 carcinomas of the uterine cervix. The results underscore the efficacy, sensitivity, and remarkable reproducibility of this approach to LOH detection and provide subchromosomal localization of two regions of chromosome 4 commonly altered in cervical tumors.