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Biomedical subjects

L O Henderson

Publications and source records attributed to L O Henderson.

At least 19 recordsLinked to original sources
PubMed2000

Serum immunoglobulins and lymphocyte subset distributions in children and adults living in communities assessed for lead and cadmium exposure.

This study assessed the impact of environmental cadmium and lead exposure on the immune system of more than 2000 children and adults. Serum immunoglobulins [immunoglobulins (Ig) A, G, and M] and peripheral blood lymphocyte phenotypes (T cells, B cells, NK cells, and CD4/CD8 subsets) were measured in a total of 2041 children and adults who lived either in sites with elevated soil levels of cadmium and lead (n = 1561) or in comparison communities (n = 480). The blood lead and urine cadmium levels of participants were somewhat higher than national averages. Mean blood lead levels were 7 microg/dl for participants aged 6-35 mo; 6 microg/dl for participants aged 36-71 mo, 4 microg/dl for participants aged 6-15 yr; and 4.3 microg/dl for participants aged 16-75 yr. Multivariate analysis indicated no marked differences in any of the immune marker distributions attributed to lead for adults or children over 3 yr of age. However, in children under age 3, increased blood lead levels, principally those over 15 microg/dl, were associated with increases in IgA, IgG, IgM, and circulating B lymphocytes. Among adults, urine cadmium levels over 1.5 microg/g were associated with higher levels of IgA and circulating B lymphocytes. No evidence of immunosuppression was noted. The findings of potential immunologic effects at lead levels > 15 microg/dl in young children and at urine cadmium levels > 1.5 microg/g in adults are interesting, but too few participants had these high levels to delineate a threshold. Therefore, we find these results intriguing, but requiring confirmation in populations with higher exposure levels.

Adolescent↗
PubMed2000

Fluorescence intensity calibration for immunophenotyping by flow cytometry.

Fluorescence intensity (FI) is the basis for classifying phenotypes by fluorescence-label flow cytometry. FI is customarily recorded as an arbitrary relative value, but with proper calibration it can be expressed in stoichiometric units called molecules of equivalent soluble fluorochrome (MESF) that reflect the concentrations of the fluorescent conjugates and the receptors they stain. Forthcoming availability of authoritative standards and consensus methods will alleviate many of the difficulties encountered in making valid MESF measurements. FI calibration establishes the true values for the critical parameters of the fluorescence measurement, a useful feature for quality control. It further allows the establishment of a comparable window of analysis across different times and laboratories, and it permits numeric assessment of antibody-binding capacity (ABC) values in selected cell populations. The relation between ABC values and receptor expression is complicated by several factors, but careful assessment of the binding chemistry can establish the actual number of receptors on cells stained by fluorescent conjugates.

Antibodies↗
PubMed2000

Glutamic acid decarboxylase and IA-2 autoantibodies in type 1 diabetes: comparing sample substrates for autoantibody determinations.

Large-scale programs designed to assess risk for type 1 diabetes through serologic assessment of autoantibodies to recombinant beta-cell autoantigens are hampered by several limitations, including the methods for sample collection and assay performance, as well as the volume required for autoantibody determinations. The present study was designed to develop a low sample-volume, primary screening method for autoantibody detection of high specificity and sensitivity, and to determine the feasibility of dried blood spots collected on filter paper in serving as vehicles for such determinations. Autoantibodies to glutamic acid decarboxylase (GAD) and ICA512bdc (IA-2), both individually and in combination, were determined in persons with type 1 diabetes, healthy controls, or individuals with other autoimmune disorders. Autoantibody results for serum, plasma, and dried blood spots were compared. GAD, IA-2, and combined GAD/IA-2 autoantibodies were concordant in their measurement from minimal volumes of serum, plasma, and whole blood extracted from dried filter paper. The autoantibody levels from the dried blood spots were, however, lower than corresponding serum samples, and, as currently designed, failed to detect low-titer autoantibodies. Despite this limitation, screening for diabetes risk can be performed using small volumes of whole blood, serum, or plasma collected onto filter paper. These methodological improvements should simplify matters, reduce costs, and increase the efficacy of screening programs for type 1 diabetes. Further development of better substrates/methods for blood-specimen collection seems necessary to exploit the full potential of this and other autoantibody measurement strategies for screening large populations.

Journal Article↗
PubMed1999

Current activities at the Centers for Disease Control and Prevention's National Diabetes Laboratory.

In 1997, the Centers for Disease Control and Prevention established the National Diabetes Laboratory in order to help prevent and treat type 1 diabetes. This state-of-the-art laboratory collaborates with research scientists and key national and international organizations throughout the world to identify and study risk factors for type 1 diabetes by developing measurements for glycosylated proteins, developing and evaluating technology for measuring genetic risk factors for the disease, and working to standardize autoantibody measurements. Developing improved technologies for diagnosing and managing diabetes and developing reference materials for properly calibrating and standardizing blood glucose meters are also critical aspects of the laboratory's work. In addition, the laboratory provides quality storage for valuable collections of biologics and other materials and facilitates sharing of specimens, associated epidemiologic data, and test results. Working with our partners in diabetes research, we are improving the diagnosis, treatment, and prevention of type 1 diabetes.

Autoantibodies↗
PubMed1998

Titration of a CD45-FITC conjugate to determine the linearity and dynamic range of fluorescence intensity measurements on lymphocytes.

To produce biologic calibrators for relative fluorescence intensity (RFI) measurements, we stained leukocytes with serial dilutions of CD45-FITC conjugate and processed them using our regular whole blood lysis procedure. Cells were stained with conjugate concentrations ranging from twice recommended to a million-fold lower. At the highest concentrations of conjugate, the RFI reached a plateau near the top of the third decade, indicating saturation of CD45 binding sites. As the concentration decreased, the RFI declined in a highly linear relationship between the dilution factor and the histogram channel number. For channel numbers corresponding to the lowest percentiles of the RFI distribution, linearity persisted down to the first half decade. The slope of this relationship revealed a true dynamic range of 4.5 decades, which was comparable to the value obtained with microbead standards calibrated in molecules of equivalent soluble fluorochrome (MESF). Our results suggest that the lower limit of linearity for fluorescence intensity from fluorescein isothiocyanate (FITC)-stained lymphocytes is below 500 MESF and that cellular autofluorescence is the major limiting factor in detecting and quantifying FITC-specific staining. This procedure provides an adroit way of characterizing the linearity and dynamic range of measurements for quantitative fluorescence cytometry using exactly the same matrix, stains, and preparation methods as those used for cellular analytes.

Antibodies, Monoclonal↗
PubMed1998

Terminology and nomenclature for standardization in quantitative fluorescence cytometry.

Terminology in any field is a complex mix of established conventions, accepted usages, disputed terms, and occasional misnomers. The terminology that has evolved for quantitative fluorescence cytometry (QFCM) is especially multifarious, in part because QFCM encompasses a range from subjective visual assessments to objective photon counts. Thus, while descriptive terms such as "dim" and "bright" are still quite useful, quantitative terms such as "binding capacity" should be used with collective understanding of their exact meanings. This article reviews current usage and proposes definitions that, with refinement from suppliers and users of QFCM technology, can provide the required clarity.

Calibration↗
PubMed1997

An evaluation of the use of dried blood spots from newborn screening for monitoring the prevalence of cocaine use among childbearing women.

A collaborative March of Dimes study was designed to examine the utility of dried blood spot (DBS) materials routinely collected from newborns as a source for monitoring cocaine exposure and to assess the prevalence of cocaine use among childbearing women in Georgia. We used a modified urinary radioimmunoassay (RIA) to anonymously detect the cocaine metabolite benzoylecgonine (BE) in DBSs. Extensive efforts were undertaken to assure absolute nonlinkage of BE data to any individual. The positive results found by RIA were confirmed by a mass spectrometry (MS) method specifically developed to detect BE in DBSs. BE was measured in 23,141 DBSs collected during 2 months of routine newborn screening in Georgia. A good correlation was observed for RIA results versus MS results (r2 = 0.97). The estimated minimal statewide BE prevalence was 4.8 per 1000 childbearing women. We demonstrated that immunoassay testing for cocaine without confirmatory testing can yield falsely elevated prevalence rates. When proper confirmatory testing is done, DBSs are a valuable source for population-based monitoring of substance abuse among childbearing women.

Blood Specimen Collection↗
PubMed1996

Isotope dilution--mass spectrometric quantification of specific proteins: model application with apolipoprotein A-I.

An enzymatic hydrolysis isotope dilution-mass spectrometric method was developed for reference quantification of specific proteins. The analytical procedure involved measuring a reproducibly hydrolyzed peptide (serving as the primary standard) unique to a specific protein. This new mass spectrometric method was evaluated by assessing the concentration of apolipoprotein (apo) A-I in the European Community Bureau of Reference (BCR) lyophilized Certified Reference Material (CRM 393). We used the method to make 96 measurements (4 replicate analyses of 4 enzymatic digests of 6 vials of BCR-CRM 393), which gave an average total protein mass of 1.048 mg (+/- 1.0% at 99% confidence limits). The total overall analytical CV was 3.95%. The results of this evaluation of our model approach to determine the concentration of a specific protein in a purified preparation demonstrated that our new mass spectrometric method can be used to measure apolipoproteins and other specific proteins without the use of epitopic immunoassay methods.

Amino Acid Sequence↗
PubMed1995

Two-dimensional protein electrophoresis and multiple hypothesis testing to detect potential serum protein biomarkers in children with fetal alcohol syndrome.

Fetal alcohol syndrome (FAS) surveillance and intervention efforts are hampered by the lack of a specific biochemical test for diagnosis of the syndrome. Based on the hypothesis that abnormalities in growth and development (key features of FAS) involve altered protein metabolism, we analyzed serum proteins by two-dimensional gel electrophoresis and image analysis to search for potential protein biomarkers of FAS. Serum samples from 12 participants in whom FAS had been diagnosed and 8 sex- and age-matched participants whose mothers did not consume alcohol were analyzed in duplicate to determine whether the integrated intensities of matched proteins are significantly altered in children with FAS. Multiple hypothesis testing on 34 of the gels consisting of more than 1700 spots per gel revealed 21 proteins that we classified as potential protein biomarkers of FAS on the basis of significant t-test differences at p < 0.02. We classified 8 of the proteins as candidate biomarkers on the basis of significant concentration differences between case and control subjects at p < 0.01. One of the proteins is clearly an isoform of retinol binding protein; two appear in the area of the gel where alcohol dehydrogenase is expected to appear; one appears to be an isoform of alpha-1-antitrypsin; three appear to be isoforms of the beta-chain of haptoglobin; three may be forms of immunoglobulin light chains; and several others have not been associated with known proteins. No single protein differentiated all case subjects from control subjects, but stepwise canonical discriminant analyses revealed four groups of spots that distinguished between FAS case and control subjects with no misclassifications.(ABSTRACT TRUNCATED AT 250 WORDS)

Biomarkers↗
PubMed1994

International Federation of Clinical Chemistry standardization project for measurements of apolipoproteins A-I and B. IV. Comparability of apolipoprotein B values by use of International Reference Material.

We performed temporal and thermal stability studies on SP3-07, a liquid-stabilized reference material for apolipoprotein (apo) B, selected during the previous phase of the International Federation of Clinical Chemistry project on standardization of apolipoprotein measurements. Results indicate that SP3-07 stored at -70 degrees C has the long-term stability required for a reference material. We assigned an accuracy-based apo B value of 1.22 g/L to SP3-07, using a nephelometric method that was calibrated with freshly isolated low-density lipoprotein for which the apo B mass value was determined by a standardized sodium dodecyl sulfate-Lowry procedure. Using a common protocol, the study participants transferred the assigned mass value from SP3-07 to the individual calibrators of the analytical systems and measured the apo B concentration of 20 fresh-frozen samples obtained from individual donors and covering a clinically relevant range of apo B values. The among-laboratory CV on these samples, analyzed by 25 analytical systems, ranged from 3.1% to 6.7%. These results demonstrate the lack of matrix effects of SP3-07 and its ability to provide accurate and comparable apo B values in a variety of immunochemical methods. On the basis of the outcome of these studies, the World Health Organization has endorsed SP3-07 as the International Reference Material for Apolipoprotein B.

Apolipoprotein A-I↗
PubMed1994

Method-dependent variations in the stability of apolipoprotein B in a stabilized liquid reference material.

Using accelerated Arrhenius-type short-term and long-term temporal studies, we evaluated the storage life of a stabilized, liquid-frozen reference material (SLRM) for human apolipoprotein B (apo B) developed by the International Federation of Clinical Chemistry. As measured by our candidate reference RIA, the concentrations of immunoreactive apo B in the SLRM showed pronounced degradation with exposure to increasing temperatures over time. The SLRM was stable for as long as 1 year when stored at - 70 degrees C, but its immunoreactive apo B declined by < 10% when stored at 4 degrees C for 10 months. Using radial immunodiffusion and an ELISA to assess the equivalency of measured mass for the accelerated thermal stability of the SLRM, we found a loss of immunoreactive apo B similar to that measured by RIA. Analyzing the same samples by liquid immunoprecipitation (nephelometry) resulted in the amount of apo B present being overestimated, especially in samples held for long periods. By using different immunological methods to evaluate this thermally aged SLRM, we demonstrated that its measured behavior varies depending on the method of quantitation.

Apolipoproteins B↗
PubMed1993

International Federation of Clinical Chemistry standardization project for measurements of apolipoproteins A-I and B. III. Comparability of apolipoprotein A-I values by use of international reference material.

In the third phase of the International Federation of Clinical Chemistry (IFCC) study for the standardization of apolipoprotein (apo) measurements, the preparation SP1-01, selected as the candidate international reference material for apo A-I, was investigated for its ability to transfer an accuracy-based value to the immunoassay calibrators and to produce comparability of the values for patients' samples. An apo A-I value of 1.50 g/L (SD 0.08 g/L) was assigned to SP1-01 by a highly standardized RIA calibrated with purified apo A-I for which the mass value had been determined by amino acid analysis. According to a common detailed protocol, the participants transferred the mass value from SP1-01 to the calibrator of each method. To confirm that uniformity of calibration ensures comparability of the values over a wide range of apo A-I values, each laboratory analyzed 50 fresh-frozen samples from individual donors, using an approach similar to that adopted by the Cholesterol Reference Laboratory Network. The consensus mean value for each sample was in excellent agreement with the value assigned by the Northwest Lipid Research Laboratories, with the average absolute bias between assigned and consensus value being 0.01 g/L. The among-laboratory CV on each of the 50 samples ranged from 2.1% to 5.6% (mean 3.6%), demonstrating that comparable apo A-I results can be obtained by a variety of immunochemical methods through the use of certified reference material. Based on the results obtained in these studies, SP1-01 has been approved as Apolipoprotein A-I International Reference Material by the World Health Organization.

Apolipoprotein A-I↗
PubMed1991

Interlaboratory study of cellular fluorescence intensity measurements with fluorescein-labeled microbead standards.

To determine the precision of cellular fluorescence intensity (FI) measurements derived from labeled microbead standards, FI results were compared from 43 different flow cytometers in 34 laboratories. All laboratories analyzed prepared aliquots of fluoresceinated calf thymocyte nuclei (Fluorotrol), human lymphocytes stained with fluoresceinated anti-CD4 antibody, and fluoresceinated microbeads used as both internal and external standards. Measurements were conducted by most laboratories on the third and fourth days after sample preparation. Results for percent of events within the gates and the histograms returned by participants indicated that the samples had remained stable and that gated populations had been properly identified. All standard curves showed strong linearity, and the pooled results from all standards produced a best-fit curve that was in close agreement with the assigned values. Nonetheless, results for cellular FI were highly variable, with CVs of 20-34%. Agreement within lab/instrument was much better, with CVs ranging from 3.0 to 9.9%. The overall variability was not obviously attributable to differences in the types of cytometer, nor could it be explained by attributes of the standard curves or any other single variable examined. However, the application of a corrective factor based on FI results for Fluorotrol allowed a two-fold improvement in the precision of FI measurements on CD4-stained lymphocytes, with an overall CV of 11%. Uncharacterized differences in the operating conditions of flow cytometers can influence cellular FI measurements, but consistent results can be obtained if a stained cellular calibrator is analyzed in addition to the proper microbead standards.

CD4-Positive T-Lymphocytes↗
PubMed1991

Establishment of reference methods for lipids, lipoproteins and apolipoproteins.

Reference methods for lipids, lipoproteins, and apolipoproteins have been developed for use as part of an accuracy base for institutional, national, or international reference systems. A widely accepted reference method exists only for total cholesterol. Well described interim or institutional in-house reference methods have been established for the other lipids, lipoproteins, and apolipoproteins. The major criteria for a reference method are 1) scientific basis, 2) sound principles, 3) available calibration and control materials, 4) traceability to a definitive method or a point of reference, and 5) applicability to reference materials that provide traceability to clinical methods and transferability to other reference laboratories. The total cholesterol reference method of the U.S. National Reference System demonstrates how a reference method can be developed and applied. Reference methods now available can lead to an accepted international accuracy base for the clinically useful lipid, lipoprotein, and apolipoprotein measurements.

Apolipoprotein A-I↗
PubMed1990

Effects of analytical method and lyophilized sera on measurements of apolipoproteins A-I and B: an international survey.

In 1987 a collaborative study was initiated with 140 laboratories worldwide to evaluate the effects of analytical method and lyophilization on the measurement of different concentrations of apolipoproteins (apo) A-I and B in four lyophilized serum pool samples. This survey confirmed that the lyophilized apo Reference Material of the International Union of Immunological Societies (IUIS) is useful for apo A-I assays as an international serum-based reference material, because among-method variation is negligible. The apo A-I concentration value of 1.24 g/L is now assigned to the IUIS Reference Material (CDC 1883) by a Centers for Disease Control RIA in-house reference method. Use of lyophilized serum preparation as a reference material for some modes of apo B measurement is questionable because of lyophilization and matrix effects. Both radial immunodiffusion and liquid immunoprecipitin methods demonstrated bias in measured apo B concentrations, compared with overall method-weighted means values on the IUIS Reference Material. Because of the uncertainty associated with LDL primary standard, protein analysis, and concentration differences among analytical methods, assigning a single apo B concentration value to the IUIS Reference Material appears inadvisable at present.

Analysis of Variance↗