Detection of serum uric acid using the optical polymeric enzyme biochip system.

An optical polymeric biochip system based on the complementary metal oxide semiconductor (CMOS) photo array sensor and polymeric enzyme biochip for rapidly quantitating uric acid in a one-step procedure was developed. The CMOS sensor was designed with N(+)/P-well structure and manufactured using a standard 0.5 microm CMOS process. The polymeric enzyme biochip was immobilized with uricase-peroxidase and used to fill the reacting medium with the sample. This study encompasses the cloning of the Bacillus subtilis uricase gene and expression in Escherichia coli, as well as the purification of uricase and measurement of its activity. The cloned uricase gene included an open reading frame of 1491 nucleotides that encodes a protein of approximately 55 kDa. The expression of the putative MBP-fusion protein involved approximately 98 kDa of the protein. The CMOS sensor response was stronger at a higher temperature range of 20-40 degrees C, with optimal pH at 8.5. The calibration curve of purified uric acid was linear in the concentration range from 2.5 to 12.5 mg/dL. The results obtained for serum uric acid correlated quite closely with those obtained using the Beckman Synchron method.

[1]  T. Ferenci,et al.  Maltose-binding protein from Escherichia coli. , 1982, Methods in enzymology.

[2]  T. Ichikawa,et al.  Cloning, sequence analysis, and expression in Escherichia coli of the gene encoding the Candida utilis urate oxidase (uricase). , 1996, Journal of biochemistry.

[3]  M. Ericson,et al.  DNA biochip using a phototransistor integrated circuit. , 1999, Analytical chemistry.

[4]  H. Mottola,et al.  Determination of uric acid at the microgram level by a kinetic procedure based on a "pseudo-induction" period. , 1974, Analytical chemistry.

[5]  L. Wallrath,et al.  Species differences in the temporal pattern of Drosophila urate oxidase gene expression are attributed to trans-acting regulatory changes. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[6]  M. Takashio,et al.  Nucleotide sequence of the uricase gene from Bacillus sp. TB-90. , 1996, Journal of Biochemistry (Tokyo).

[7]  G. Berti,et al.  Use of 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone chromogenic system in direct enzymic assay of uric acid in serum and urine. , 1980, Clinical chemistry.

[8]  J. Caballero,et al.  Uricase from leaves: its purification and characterization from three different higher plants , 1997, Planta.

[9]  H Inouye,et al.  Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. , 1988, Gene.

[10]  R. Haeckel.  The Use of Aldehyde Dehydrogenase to Determine H2O2-Producing Reactions. I. The Determination of the Uric Acid Concentration , 1976, Journal of clinical chemistry and clinical biochemistry. Zeitschrift fur klinische Chemie und klinische Biochemie.

[11]  J. Keilin.  THE BIOLOGICAL SIGNIFICANCE OF URIC ACID AND GUANINE EXCRETION , 1959 .

[12]  T. Vo‐Dinh,et al.  Detection of E. coli using a microfluidics-based antibody biochip detection system , 2001, Fresenius' journal of analytical chemistry.

[13]  G. Guilbault,et al.  Enzyme electrode sensing oxygen for uric acid in serum and urine. , 1974, Analytical chemistry.

[14]  B. Slatko,et al.  An Escherichia coli vector to express and purify foreign proteins by fusion to and separation from maltose-binding protein. , 1988, Gene.

[15]  Y. Hongoh,et al.  Cloning, sequence analysis and expression in Escherichia coli of the gene encoding a uricase from the yeast-like symbiont of the brown planthopper, Nilaparvata lugens. , 2000, Insect biochemistry and molecular biology.

[16]  M. Pontet,et al.  Comparative activity of peroxidase-antibody conjugates with periodate and glutaraldehyde coupling according to an enzyme immunoassay. , 1995, Annales de biologie clinique.

[17]  C. Pundir,et al.  Discrete analysis of serum uric acid with immobilized uricase and peroxidase. , 1999, Journal of biochemical and biophysical methods.

[18]  H. Ihara,et al.  Effects of serum bilirubin on determination of uric acid by the uricase-peroxidase coupled reaction. , 1992, Clinical chemistry.

[19]  M. Pineda,et al.  Isolation and characterization of uricase from bean leaves and its comparison with uredospore enzymes , 1999 .

[20]  Chu di Guana,et al.  Vectors that facilitate the expression and purification of foreign peptides in Escherichia coli by fusion to maltose-binding protein. , 1988 .

[21]  M. Smyth,et al.  Poly(o-aminophenol)-modified bienzyme carbon paste electrode for the detection of uric acid. , 1996, Talanta.

[22]  P. Caliceti,et al.  Therapeutic proteins: a comparison of chemical and biological properties of uricase conjugated to linear or branched poly(ethylene glycol) and poly(N-acryloylmorpholine). , 2000, Farmaco.

[23]  Use of 3,5-dichloro-2-hydroxybenzenesulfonic acid/4-aminophenazone chromogenic system in direct enzymic assay of uric acid in serum and urine. , 1980 .

[24]  Y. Hasebe,et al.  Chemical Amplification of Uric Acid Sensor Responses by Dithiothreitol , 1994 .