| 1. |
- Jalanko, A, et al.
(författare)
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Screening for defined cystic fibrosis mutations by solid-phase minisequencing
- 1992
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Ingår i: Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 38:1, s. 39-43
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Tidskriftsartikel (refereegranskat)abstract
- We have developed a rapid method for the quantitative detection of point mutations and deletions. In this minisequencing method, enzymatically amplified DNA, 5'-biotinylated in one strand, is bound to a solid phase and denatured. A detection primer, constructed to end immediately before the mutation, is annealed to the immobilized single-stranded template and elongated with a single, labeled deoxynucleoside residue. We have applied the solid-phase minisequencing method to the detection of the major mutation, delta F508, causing cystic fibrosis (CF). In the presence of the allele with the delta F508 mutation, [3H]dTTP is incorporated; with the nonmutated allele, [3H]dCTP is incorporated. Thus, samples from heterozygous individuals allow the incorporation of both labels. The method was evaluated by analyzing 59 coded DNA specimens collected from 20 Finnish CF patients and their parents. The ratio of [3H]C to [3H]T gave unambiguously the allele combination. The solid-phase minisequencing method was also applicable to the analysis of three CF mutations simultaneously, i.e., delta F508, G542X, and G551D. We conclude that the microtiter-plate-based minisequencing test is an accurate method for the screening of defined sequence alterations in the CF gene.
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| 2. |
- Kontula, K, et al.
(författare)
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Apolipoprotein E polymorphism determined by restriction enzyme analysis of DNA amplified by polymerase chain reaction : convenient alternative to phenotyping by isoelectric focusing
- 1990
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Ingår i: Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 36:12, s. 2087-2092
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Tidskriftsartikel (refereegranskat)abstract
- Three common alleles determine six apolipoprotein E (apo E) phenotypes that are associated with variations in serum cholesterol in the population. This genetic variation results from single nucleotide alterations at two DNA loci encoding the amino acid residues 112 and 158 of apo E. We compared results of apo E phenotyping carried out by isoelectric focusing with those of apo E genotyping accomplished by direct DNA analysis. In the latter, the target DNA was amplified by the polymerase chain reaction (PCR) and subsequently analyzed by digestion with the restriction enzyme Hha I, followed by polyacrylamide gel electrophoresis of the cleavage products. With one exception, these two techniques yielded similar results from all 40 samples tested. In addition, a rare variant form of apo E (phenotype E1) was analyzed separately and incorrectly diagnosed as E2 by the Hha I digestion method; the anticipated mutation in the codon 127 was, however, confirmed by demonstration of a new Taq I restriction site in this variant gene. These data confirm that the common isoforms of apo E usually arise from genetic variation of the codons 112 and 158 and demonstrate the feasibility of the PCR technique in apo E genotyping.
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| 3. |
- Lilja, H, et al.
(författare)
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Prostate-specific antigen in serum occurs predominantly in complex with alpha 1-antichymotrypsin
- 1991
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Ingår i: Clinical Chemistry. - : American Association for Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 37:9, s. 25-1618
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Tidskriftsartikel (refereegranskat)abstract
- Immunologic measurements of the serum concentration of prostate-specific antigen (PSA), an abundant prostatic-secreted serine proteinase, are frequently used to monitor patients with prostate cancer, though it has not been ascertained whether this immunoreactivity represents a PSA zymogen, the active proteinase, or PSA complexed to extracellular proteinase inhibitors. To characterize the PSA immunoreactivity in serum, we used monoclonal antibodies produced against PSA and a polyclonal rabbit IgG against alpha 1-antichymotrypsin in the design of three noncompetitive PSA assays: assay T, which detected PSA both when present as the active proteinase and when complexed to alpha 1-antichymotrypsin; assay F, which recognized the active proteinase but most poorly detected PSA complexed to alpha 1-antichymotrypsin; and assay C, which was specific for PSA complexed to alpha 1-antichymotrypsin. We used the three assays to measure PSA immunoreactivity in 64 patients' sera and in the effluent after gel chromatography of sera from four patients. This identified an 80- to 90-kDa complex between PSA and alpha 1-antichymotrypsin as the predominant fraction of the PSA immunoreactivity in blood plasma; an immunoreactive 25- to 40-kDa compound was the minor fraction.
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| 4. |
- Lindqvist, Ulla, et al.
(författare)
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Seven different assays of hyaluronan compared for clinical utility
- 1992
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Ingår i: Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 38:1, s. 127-132
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Tidskriftsartikel (refereegranskat)abstract
- To compare six assays of hyaluronan (hyaluronic acid; HYA) in serum, developed in different laboratories, we analyzed 10 samples from each of three groups: healthy persons, patients with primary biliary cirrhosis, and patients with rheumatoid arthritis. All the assays are based on the use of affinity proteins specific for HYA, prepared from cartilage or brain tissue, and are analogous to RIA or enzyme immunoassay techniques. The assay results were of the same magnitude. Although statistical analysis indicated that the methods in some cases deviated significantly from one another, this variation was less than the physiological variation in the healthy population. Therefore, the results of clinical investigations in which the various methods have been used are comparable. The analyses have high specificity and sensitivity for primary biliary cirrhosis but are somewhat less suitable for detecting rheumatoid arthritis. A seventh laboratory, which obtained antibodies to HYA, used these in an RIA to analyze a separate series of serum specimens. Results were in agreement with those obtained by one of the other assays.
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| 5. |
- Xie, Bin, et al.
(författare)
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Urea and lactate determined in 1-μL whole-blood samples with a miniaturized thermal biosensor
- 1994
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Ingår i: Clinical Chemistry. - : American Association for Clinical Chemistry. - 0009-9147 .- 1530-8561. ; 40:12, s. 2282-2287
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Tidskriftsartikel (refereegranskat)abstract
- A miniaturized flow-injected thermal biosensor was developed for the determination of urea and L-lactate in undiluted blood in 1-μL samples. The sensor employed a small enzyme column constructed of stainless steel tubing and microbead thermistors. Urease and lactate oxidase/catalase were separately immobilized onto controlled-pore glass beads, which, in turn, were charged into the enzyme column. With a flow rate of 70 μL/min, linear analytical ranges from 0.2 to at least 50 mmol/L and 0.2 to 14 mmol/L were obtained for urea and lactate, respectively. The relative standard deviations (CVs) for measurements of analyte in buffer were 0.91% for urea and 1.84% for lactate. For urea in whole blood, the CV for 50 determinations was 4.1%. Contrived samples containing various concentrations of urea and L-lactate in whole blood were determined with this sensor and with a spectrophotometric method. Comparisons of the results gave correlation coefficients of 0.989 and 0.984 for 30 blood urea and 30 blood lactate assays in concentrations ranging from 4 to 20.9 mmol/L and from 1.7 to 12.7 mmol/L, respectively.
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| 6. |
- Agardh, Carl-David, et al.
(författare)
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Association between urinary N-acetyl-beta-glucosaminidase and its isoenzyme patterns and microangiopathy in type 1 diabetes mellitus
- 1991
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Ingår i: Clinical Chemistry. - : American Association for Clinical Chemistry. - 0009-9147. ; 37:10 Pt 1, s. 1696-1699
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Tidskriftsartikel (refereegranskat)abstract
- Urinary N-acetyl-beta-glucosaminidase (NAG) and its isoenzymes (NAG A and NAG B) in samples from 87 type 1 diabetic patients and 40 apparently healthy reference subjects were studied with enzyme immunoassays. The diabetic patients had higher concentrations of urinary NAG than did the control subjects (P less than 0.01), but the isoenzyme pattern did not differ. There was a positive correlation between metabolic control (Hb A1c concentrations) and total NAG (P less than 0.01), NAG A (P less than 0.01), and NAG B (P less than 0.001). The diabetic patients were divided into three groups, depending on the degree of retinopathy. Subjects with severe forms of retinopathy did not have increased concentrations of urinary NAG unless they had concomitant nephropathy. The isoenzyme pattern was similar irrespective of degree of retinopathy or nephropathy. The results indicate that concentrations of urinary NAG are positively correlated to the degree of nephropathy, whereas there is no such correlation to the degree of retinopathy.
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| 7. |
- Kyhse-Andersen, Jan, et al.
(författare)
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Serum cystatin C, determined by a rapid, automated particle-enhanced turbidimetric method, is a better marker than serum creatinine for glomerular filtration rate
- 1994
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Ingår i: Clinical Chemistry. - : American Association for Clinical Chemistry. - 0009-9147. ; 40:10, s. 1921-1926
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Tidskriftsartikel (refereegranskat)abstract
- We describe a fully automated particle-enhanced turbidimetric assay for cystatin C in undiluted serum and EDTA-plasma. The throughput is 90 samples per hour and urgent samples can be analyzed in 7 min. The assay range (0.4-14.1 mg/L) covers the concentration range in health and disease. The within- and between-run imprecision is 0.9% and 2.2%, respectively. Analytical recovery of additions of recombinant cystatin C averaged 98%. Rheumatoid factors (< or = 323,000 IU/L), bilirubin (< or = 150 mumol/L), hemoglobin (< or = 1.2 g/L), and triglycerides (< or = 8.5 mmol/L) do not interfere in the assay. In view of the superior (by ROC analysis) diagnostic accuracy of serum concentrations of cystatin C for reduced glomerular filtration rate (GFR) in comparison with creatinine, cystatin C seems an attractive alternative to creatinine for estimation of GFR.
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| 8. |
- Gislén, Lars, et al.
(författare)
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Rotor Neurons: Basic Formalism and Dynamics
- 1992
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Ingår i: Neural Computation. - : MIT Press. - 1530-888X .- 0899-7667. ; 4:5, s. 737-745
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Tidskriftsartikel (refereegranskat)abstract
- Rotor neurons are introduced to encode states living on the surface of a sphere in D dimensions. Such rotors can be regarded as continuous generalizations of binary (Ising) neurons. The corresponding mean field equations are derived, and phase transition properties based on linearized dynamics are given. The power of this approach is illustrated with an optimization problem—placing N identical charges on a sphere such that the overall repulsive energy is minimized. The rotor approach appears superior to other methods for this problem both with respect to solution quality and computational effort needed.
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