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L O Henderson

Publications and source records attributed to L O Henderson.

18 recordsLinked to original sources
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↗
PubMed1990

Impact of protein measurements on standardization of assays of apolipoproteins A-I and B1.

In 1989 the Committee on Apolipoproteins of the International Federation of Clinical Chemistry and the Centers for Disease Control conducted an international survey of total-protein measurements of isolated low-density lipoproteins (LDL) and delipidated high-density lipoproteins (HDL), and of their relationships to the National Institute of Standards and Technology (NIST) bovine serum albumin (BSA) Standard Reference Material (SRM). Most of the 93 apolipoprotein laboratories surveyed use the Lowry total-protein method. Results reported with the LDL preparations demonstrated a large bias and variation among methods; those with delipidated HDL were not as great, but were similar to those for BSA. Performance improved appreciably with use of the Lowry-sodium dodecyl sulfate method and the NIST BSA SRM for protein measurement. The total CVs, including among-laboratory and within-laboratory errors, averaged approximately 50% and 23% for LDL, 17% and 11% for HDL, and 18% and 11% for BSA solutions by all methods and by the Lowry methods, respectively. Regardless of the methods used, greater variability of the protein measurements was seen with the normally occurring LDL than with the nonlipoprotein BSA or delipidated HDL. The mean CV values for all samples among laboratories averaged between 10% and 15% with the modified Lowry methods; the biuret method gave the highest among-laboratory CV, 34%; the Kjeldahl had the lowest, 7.7%. Use of the same methodology and primary nonapolipoprotein standard is essential for comparability of protein results for apolipoprotein primary standard solutions. This is especially true for apolipoprotein B, because its inherent properties and lability make protein analysis difficult. This study supports the use of a standardized selected Lowry-sodium dodecyl sulfate method traceable to quantitative amino acid analysis as a point of reference for determining the protein concentration of primary calibration reference materials for apolipoproteins.

Apolipoproteins A↗
PubMed1989

Model system evaluating fluorescein-labeled microbeads as internal standards to calibrate fluorescence intensity on flow cytometers.

Fluorescence intensity calibration was evaluated in a model system for flow cytometers using commercially available fluorescein-labeled microbeads as internal standards and stabilized fluoresceinated thymus cell nuclei (Fluorotrol) as surrogates for stained mononuclear cells. Spectrophotometrically determined calibration values for the microbeads were used to generate a standard curve that converted green fluorescence histogram channels into molecular equivalents of soluble fluorescein (MESF). In 19 analyses repeated during a single run, the coefficients of variation (CVs) for the derived MESF values on both dimly and brightly stained Fluorotrol populations were less than 2%. In 26 separate determinations over 14 weeks, the CVs of the derived MESF values were less than 3%. The MESF values of the dim and bright Fluorotrol populations derived from the microbead standard curves were both about 50% lower than those determined by direct spectrophotometric analysis of Fluorotrol. The analytical imprecision of fluorescence intensity measurements in this idealized model system has a CV less than 3%, and the analytical inaccuracy shows that calibration in MESF units remains uncertain over about a two-fold range.

Animals↗
PubMed1988

Preparation of lyophilized human serum based reference materials with graded levels of apolipoproteins A-I and B.

Surveys of national and international laboratories indicate that among-laboratory and between-laboratory sources of variance account for the majority of variability in laboratories measuring apolipoproteins A-I and B. These sources of variance are amenable to correction through the use of common quality control and reference materials. We utilized proven techniques employing ethyl alcohol and acetate buffer to precipitate either apolipoprotein A-I rich or apolipoprotein B rich fractions that were blended with whole or delipidated serum producing five pilot-sized pools containing graded levels of apolipoproteins. After lyophilization the pools were tested and each pool contained levels of analytes similar to frozen serum and contained varied amounts of apolipoproteins A-I and B. Temporal and accelerated thermal stability testing demonstrated stability of the analytes in the pools with time (three years) and temperature (up to 56 degrees C). This technology provides a preparative procedure for apolipoprotein reference materials over the extended range needed in clinical applications.

Apolipoprotein A-I↗
PubMed1988

An evaluation of the trends in analytical performance of international apolipoprotein A-1 and B assays.

Two international apolipoprotein A-1 and B (apoA-1 and apoB) surveys were conducted in 1983 and 1986 with use of a lyophilized serum measured by 55 and 91 participants, respectively. The chief source of variability in both surveys was among laboratories, but among-method variation was significant for apoB. Comparing the 1986 with the 1983 findings, we saw a 60% decrease in apoA-1 variability among laboratories, and a similar decrease (53%) was found for apoB. Evaluation of individual measurements suggests that some collaborators may have adjusted their calibrators toward the consensus values published in 1983.

Apolipoprotein A-I↗
PubMed1988

High-density lipoprotein apolipoproteins in urine: I. Characterization in normal subjects and in patients with proteinuria.

A high-resolution two-dimensional electrophoretic method for protein, with silver staining, has been used to characterize and identify urinary high-density-lipoprotein apolipoproteins (HDL-Apos) and their isoforms in healthy subjects and in patients with kidney disease. Analytical techniques based on both molecular mass and ultracentrifugal flotation properties were used to isolate urinary lipoprotein particles with characteristics identical to those of HDL in plasma. HDL-Apos identified in urine of normal subjects and patients with glomerular proteinuria were Apos A-I, A-II, and C. Five isoforms of Apo A-I were present. Immunostaining of electroblotted proteins further confirmed the presence of HDL-Apos in urine. Creatinine clearance rate was decreased in the patients with proteinuria, and ranged from 32.5 to 40 mL/min. Concentrations of cholesterol and triglycerides in serum were greater in the patients' group, whereas mean HDL-cholesterol (0.68, SD 0.10 mmol/L) and Apo A-I (0.953, SD 0.095 g/L) were significantly (each P less than 0.01) lower. Results of this study suggest that measurement of urinary Apo A-I will reflect excretion of HDL in urine.

Apolipoprotein A-I↗
PubMed1988

High-density lipoprotein apolipoproteins in urine: II. Enzyme-linked immunoassay of apolipoprotein A-I.

We have developed a capture antibody, noncompetitive, enzyme-linked immunoassay for urinary apolipoprotein A-I (Apo A-I) in urine, with use of affinity-purified polyclonal antisera against Apo A-I. A 96-well microtiter plate format is used, with unconcentrated urine as sample and dilutions of serum or high-density lipoprotein (HDL) as standards. The intra- and interassay variation (CV) averaged 7.4% and 9.4%, respectively. The limit of detection is low (1.25 ng/L), and no cross-reactivity with Apo B, C, E, or A-II was detected. The mean (+/- SD) concentrations of Apo A-I in urine of patients with glomerular proteinuria were a thousandfold greater (38.4 +/- 23.1 mg/L) than in normal subjects (16.3 +/- 11.3 micrograms/L in men, 17.97 +/- 7.7 micrograms/L in women, a significant difference, P less than 0.001). Apo A-I measurements correlated very well (r = 0.92) with selectivity index assessment. The diurnal variation of the concentration of Apo A-I in urine appears to result from dilution related to fluid intake. This enzymatic method is easy to perform, can be used with large numbers of samples, and is adaptable for use in the routine clinical laboratory. The method holds promise for discriminating between normal and subclinical kidney disease populations by measuring the concentrations of urinary Apo A-I excreted on HDL particles.

Adolescent↗
PubMed1987

Quantitative differences among various proteins as blocking agents for ELISA microtiter plates.

We tested instantized dry milk, casein, gelatins from pig and fish skin, serum albumin and several other proteins for their abilities to block non-specific binding (NSB) of a peroxidase-conjugated immunoglobulin to polystyrene microtiter plate wells. Each blocking protein was tested across a million-fold concentration range, both in simultaneous incubation with the peroxidase conjugate and as a pretreatment agent where excess protein was washed away before incubation with the conjugate. Overall, instantized milk and casein were the most effective proteins tested: they inhibited NSB by over 90% in both the simultaneous and pretreatment modes at far lower concentrations than most of eight other proteins. Enzymatically hydrolyzed porcine skin gelatin was the least effective protein tested: it did not reduce NSB by more than 90% even at its highest concentrations; its blocking ability fell rapidly upon dilution; and it was almost useless as a pretreatment agent. Fish skin gelatin showed much better blocking activity than hydrolyzed porcine gelatin, and it still had the practical advantage of remaining fluid even under refrigeration. Our results suggest that some proteins (such as casein) block NSB to plastic primarily through protein-plastic interactions, while others (such as porcine skin gelatin) block primarily through protein-protein interactions. Although the optimal blocking agent for any particular ELISA system must be determined by empirical testing, these results should be helpful in selecting the best possible candidate proteins for further evaluation.

Animals↗
PubMed1987

An international collaborative study on standardization of apolipoproteins A-I and B. Part I. Evaluation of a lyophilized candidate reference and calibration material.

We evaluated a lyophilized serum preparation for use as a candidate Reference Material for apolipoproteins (apo) A-I and B. An international collaborative study was conducted with 28 participating laboratories, selected on the basis of participation and demonstrated expertise in a 1983 survey of apolipoproteins A-I and B. The analytical suitability of the material was established by confirming linearity of its dose-response curves over a desired concentration range and demonstrating that its response curves paralleled those for fresh sera. Differences in dilution-adjusted mass units ascribable to the five analytical methods used by the various laboratories constituted only 1% of the total variation for apo A-I, but 32% for apo B. The dominant source of error, however, for both apo A-I and B was the variability among laboratories, rather than variability among methods and antisera. The assigned consensus mass-concentration units based on study data are 1.124 g/L for apo A-I, 0.589 g/L for apo B. For these estimates the coefficients of variation were 13% and 27%, respectively. These findings on the proposed Reference Material meet the requirements suggested by the World Health Organization's Expert Committee on Biological Standards for a candidate WHO Reference Preparation.

Apolipoprotein A-I↗
PubMed1987

An international collaborative study on standardization of apolipoproteins A-I and B. Part II. Evaluation of contributions of antisera to among-laboratory variance components.

Local antisera (LA) were compared with a common interim reference antiserum (IRA) to examine the antiserum component of the among-laboratory variation in an international collaborative study with 28 laboratories evaluating a candidate international Reference Material (apo-RM) for apolipoproteins A-I (apo A-I) and B (apo B). Measurement of the relative concentration of lyophilized preparations differed by less than 1% for LA and IRA. The percentage of the total variation in measurement of the concentration of apo-RM that was contributed by antisera among laboratories was 5% and 8% for apo A-I and B, respectively. Estimated differences from overall mean concentrations for the five different immuno-methods were greater for apo B (range: +22% to -23.5%) than for apo A-I (range +14% to -14%), but were similar within a method for LA and IRA. The results indicated that antisera are not a major source of error among laboratories and, indeed, are responsible for relatively little of the total variability.

Apolipoprotein A-I↗
PubMed1979

Plasma-triglycerides do not regulate high-density lipoprotein concentrations.

Diets high in carbohydrate and diets restricted in fat and calories were used to induce large fluctuations in plasma-triglycerides in patients with severe familial hypertriglyceridaemia. High-density-lipoprotein (HDL) concentrations were estimated by radioimmunoassay of the major HDL apoprotein, apo A-I, and were correlated with plasma cholesterol and triglyceride concentrations. Patients with type-1 hyperlipoproteinaemia had apo A-I concentrations about 50% of normal, and no increase in apo A-1 concentrations was observed even when plasmatriglycerides were reduced to the normal range. Apo A-I concentrations in type-5 hyperlipoproteinaemia were not consistently low and did not correlate with plasma-lipid concentrations. It does not seem that hypertriglyceridaemia reduces HDL in either disorder.

Dietary Carbohydrates↗
PubMed1978

Abnormal concentration and anomalous distribution of apolipoprotein A-I in Tangier disease.

A double-antibody radioimmunoassay was used to determine the concentration and distribution of apolipoprotein A-I (apoA-I) in the plasma of patients with Tangier disease. Obligate heterozygotes for Tangier disease had apoA-I levels that were 50% or less of controls, even when estimates of high-density lipoprotein cholesterol concentration were normal. Plasma apoA-I levels in Tangier homozygotes were at most 3% of normal. Most of the apoA-I in the plasma of homozygotes sedimented in the ultracentrifuge at density 1.21 g/ml, and no more than 20% was recovered in the plasma lipoprotein fraction. Radioimmunoassay demonstrated no immunochemical differences between Tangier and control apoA-I.

Adult↗
PubMedDate not supplied

Chlorinated hydrocarbon levels in human serum: effects of fasting and feeding.

Twenty healthy adult humans had serum samples drawn on four occasions within a 24-hr period: after a 12 hr overnight fast, 4-5 hr after a high fat breakfast, at midafternoon, and the next morning after another 12 hr fast. Nonfasting samples had 22% to 29% higher mean concentrations (p less than 0.05) than did fasting samples for polychlorinated biphenyls (PCBs, 4.81 vs 3.74 ng/g serum wt), hexachlorobenzene (HCB, 0.163 vs 0.134 ng/g serum wt), and p,p'-dichlorodiphenyl-dichloroethylene (p,p'-DDE, 6.74 vs 5.37 ng/g serum wt) measured by electron capture gas liquid chromatography. Total serum lipids were estimated from measurements of total cholesterol, free cholesterol, triglycerides, and phospholipids and were 20% higher in nonfasting samples than in fasting samples (7.05 g/L vs 5.86 g/L). When PCBs, HCB, and p,p'-DDE concentrations were corrected by total serum lipids, results from fasting and non-fasting samples were not statistically different. Because of the differences in these chlorinated hydrocarbon concentrations observed with different sample collection regimens, meaningful comparison of analytical results requires standardizing collection procedures or correcting by total serum lipid levels.

Adult↗