Biomedical subjects
S S Kuan
Publications and source records attributed to S S Kuan.
The cardiolipin antigen: chemistry and composition.
Cardiolipin, the primary lipid hapten in the antigen suspension used for the detection of antitreponemal antibodies in the sera of syphilitic patients, was successfully coupled to glucose oxidase, peroxidase, and some other enzymes using different crosslinking agents. These complexes were used to replace the pure uncomplexed cardiolipin for the preparation of the antigen suspension. When these suspensions were allowed to react with serum that contained anticardiolipin antibodies the activity of the enzyme was inhibited. In the absence of antibody, no enzyme inhibition was observed.
Determination of total protein in serum using a tyrosinase enzyme electrode.
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Measurement of creatine kinase isoenzyme MB in serum with immunoseparation and electrochemical detection.
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High pressure liquid chromatographic determination of xanthomegnin in grains and animal feeds.
A high pressure liquid chromatographic (HPLC) method is described for the determination of xanthomegnin in grains and mixed animal feeds at levels ranging from 150 to 1200 ng/g. This is equivalent to actual amounts of xanthomegnin injected on the HPLC system at from 15 to 120 ng/injection. Xanthomegnin is extracted with chloroform and 0.1M phosphoric acid. An aliquot of the crude extract is purified by column chromatography using a commercially available silica gel cartridge. Xanthomegnin is then separated from the remaining interferences by HPLC with a reverse phase C-8 column, and subsequently determined by absorbance detection at 405 nm. Elapsed time for the method from initial extraction to final HPLC determination is approximately 1 h. Recoveries of xanthomegnin added to grains and animal feeds at levels from 150 to 1200 ng/g averaged 82% with a coefficient of variation of 10.2%.
Amperometric enzymic determination of triglycerides in serum.
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Immunoelectrochemical assay for creatine kinase isoenzyme MB.
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Fluorometric enzymatic determination of serum creatinine on the surface of silicone-rubber pads.
An enzymatic semi-solid surface fluorometric method is described for the determination of serum creatinine on silicone-rubber pads. In this method, the glutamate dehydrogenase, alpha-ketoglutarate, ADP and NADH are mixed, then 30 microliters of diluted serum is added. After the free ammonium ion in serum is consumed, creatininase is added to initiate the assay. The rate of disappearance of NADH fluorescence at 460 nm (excitation wavelength 340 nm) is monitored and is proportional to the serum creatinine concentration. The whole assay uses only 0.41 I.U. creatininase, and takes less than 5 min. The calibration curve is linear up to 82 mg creatinine per liter. The proposed method offers a rapid, simple and inexpensive means for creatinine assay. The results obtained correlate well with the modified Jaffe method applied on Technicon SMA 12/60, with a correlation coefficient of 0.998. The recovery averages 99.3%.
Fluorometric and colorimetric enzymic determination of triglycerides (triacylglycerols) in serum.
We describe two fully enzymic methods, fluorometric and colorimetric, for determination of triglycerides (triacylglycerols) in serum. Samples are incubated with microbial lipase for 10 min, and the glycerol released from the triglycerides is oxidized by NAD+ in the presence of glycerol dehydrogenase. In the fluorometric method, the resulting NADH is in turn oxidized by resazurin as catalyzed by diaphorase to form resorufin, a highly fluorescent compound. In the colorimetric method, the NADH is oxidized by coupling with a tetrazolium salt/diaphorase system to form formazan, a highly colored compound. Calibration curves, constructed by plotting change in fluorescence or absorbance vs concentration of triglycerides, were linear up to 6 and 5 g of triglycerides per liter of serum for the fluorometric and colorimetric methods, respectively. The assays require only 5 and 15 microL of serum for fluorometry and colorimetry, respectively. The CV was 0.59% for the fluorometric method, 0.91% for the colorimetric procedure. The time for analysis for either method is less than 15 min. The results correlate well with those obtained by the Dow Diagnostic Kit method, a colorimetric method in which glycerol kinase and glycerol-1-phosphate dehydrogenase form NADH from ATP and NAD+ in the presence of glycerol and glycerol 1-phosphate.
An immobilized hexokinase enzyme stirrer for a simple and economical assay of plasma glucose.
We have adapted the hexokinase glucose procedure to an immobilized enzyme stirrer for the determination of glucose concentrations in human blood plasma. The procedure is a fluorometric rate method measuring the formation of NADPH catalyzed by immobilized glucose-6-phosphate dehydrogenase and hexokinase held within a tiny stirrer. The enzyme stirrer is stable for at least two months and can be used over eight-hundred assays without any loss of activity.
A specific enzyme electrode for L-phenylalanine.
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Amperometric determination of total cholesterol in serum, with use of immobilized cholesterol ester hydrolase and cholesterol oxidase.
We describe an electrochemical method for simple, rapid, and economical assay of total serum cholesterol with use of immobilized cholesterol esterase (EC 3.1.1.13) and cholesterol oxidase (EC 1.1.3.6). A rotating porous cell was specially designed to hold the immobilized enzymes firmly and to allow the reaction mixture to pass through the enzyme layer easily, thus catalyzing the enzymatic transformation quickly. Hydrogen peroxide resulting from a catalytic reactions was measured amperometrically at +0.60 V cs. a standard calomel electrode. The calibration curve for total serum cholesterol was linear from 0 to 5.00 g/liter. The method is specific, precise, and inexpensive. Our results correlate well with those obtained by the method of Abell et al. [Stand. Methods Clin. Chem. 2, 26 (1958)], the correlation coefficient being 0.992. Ascorbic acid or bilirubin in concentrations up to 700 mg/liter do not interfere. The immobilized enzymes are stable, and the same immobilized-enzyme stirrer can be used for at least 200 accurate, reproducible assays.
Determination of plasma glucose with use of a stirrer containing immobilized glucose dehydrogenase.
A stirrer containing immobilized glucose dehydrogenase has been successfully used for determining glucose in plasma. The device is usable for at least two months and for about 500 assays. The reaction was measured kinetically and linearity was observed to 4 g of glucose per liter. Tested with aqueous glucose and with deproteinized plasma, within-day and day-to-day precision were good. Interference and method-comparison (hexokinase method) were examined. The performance of this system makes the technique useful and attractive for routine use in small-volume clinical laboratories.
Measurement of glucose in plasma, with use of immobilized glucose oxidase and peroxidase.
We describe a novel fluorometric technique for simple, rapid, and economical assay of glucose by use of immobilized glucose oxidase and peroxidase. A cylindrical magnetic stirrer was specially designed to hold the immobilized enzymes firmly and to allow the reaction minzymic transformation quickly. Blood plasma can be assayed directly with no pretreatment. Ascrobic acid, uric acid, and creatinine in concenctrations of 0.25, 0.25, and 0.2 g/liter, respectively, did not interfere. The linearity of the assay was extended to 4.0 g of glucose concentrations. Results by our technique and by the otoluidine or hexokinase methods agreed well. The immobilized enzymes are stable for several months and can be used for several hundred highly accurate and reproducible assays.
An inducible amidase from Pseudomonas striata.
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An alternative method for the determination of uric acid in serum.
A novel fluorometric method for the determination of uric acid based on the coupled reactions of uric acid with uricase and peroxidase to form highly fluorescent 2,2'dihydroxy-3,3'-dimethoxy-biphenyl-5,5'-diacetic acid is described. The calibration curve was constructed from a series of standard uric acid solutions vs. the corresponding relative fluorescence. It was linear up to 15 mg/dl (0.9 mmol/l). The serum or plasma samples must be deproteinated with (absolute) ethanol before assay and its uric acid content can be obtained from the calibration curve. This method is rather simple, having good precision and accuracy. High level ascorbic acid in the sample falsely elevates uric acid concentration. Comparison of the results obtained on the patient sera with a colorimetric phosphotungstate method and a standard enzymatic spectrophotometric method gave coefficients of correlation of 0.908 and 0.986, respectively.
A novel enzyme electrode method for the determination of nitrite based on nitrite reductase.
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An improved electrode for the assay of urea in blood.
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