Development of an ELISA test for determination of the urinary trypsin inhibitor: analytical performance and applications.
Increased urinary excretion of urinary trypsin inhibitor (UTI) has been reported in various inflammatory conditions and in Alzheimer's subjects, but its diagnostic potential remains to be elucidated. A reliable and specific enzyme-linked immunosorbent assay (ELISA) test for the determination of the UTI in human urine was developed. This assay was performed using 96-well microtiter plates. The plate surface is coated with an anti-UTI polyclonal antibody, the urine sample was added in a dilution range, and the detection was achieved using the enzyme-conjugated antibody. The assay was quantified by the build-up of colored product upon the addition of the substrate. Recoveries were 93%, and the intra- and inter-assay CVs were 4.25% and 21%, respectively. The ELISA showed parallelism of standard and urine samples and no significant interference by the biological matrix. The usefulness of the assay has been demonstrated by applying it to urine samples from Alzheimer's disease patients, and comparing with negative controls. UTI urinary levels are significantly increased in Alzheimer's subjects.
D'Ercole
Stefania
S
Department of Molecular, Cellular, and Animal Biology, Post Graduate-School in Clinical Biochemistry, University of Camerino, Camerino (MC), Italy. stefania.dercole@unicam.it
Priori
Anna Maria
AM
Pucciarelli
Stefania
S
Fioretti
Evandro
E
Tacconi
Rosalia
R
Angeletti
Mauro
M
Eleuteri
Anna Maria
AM
Pucci
Eugenio
E
D000368:Aged; D000369:Aged, 80 and over; D000906:Antibodies / Q000276:immunology; D000941:Antigens / Q000276:immunology / Q000652:urine; D002138:Calibration; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D012680:Sensitivity and Specificity; D014361:Trypsin Inhibitors / Q000276:immunology / Q000652:urine*; D016482:Urinalysis / Q000379:methods*
D000906:Antibodies; D000941:Antigens; D014361:Trypsin Inhibitors