Assessment of urinary total testosterone production by a highly sensitive time-resolved fluorescence immunoassay

Abstract Background: A rapid and highly sensitive assay for tumor-associated trypsinogen-2 (TAT-2) based on the time-resolved fluorescence immunoassay (TRFIA) detection technique was developed for the determination of serum TAT-2 levels in cancers. Results: The measurement range of TAT-2-TRFIA was 1.53-300 ng/mL. The within-run and between-run coefficients of variation of TAT-2-TRFIA were 4.38% and 7.82%, respectively. The recovery rate of TAT-2-TRFIA was 103.0%. The cross-reaction rates of trypsin and T-cell immunoglobulin mucin 3 were 0.02% and 0.82%, respectively. The TAT-2-positive rates in lung cancer, liver cancer, nasopharyngeal cancer, cholangiocarcinoma, brain cancer, and pancreatic cancer were 45.9%, 50.0%, 45.0%, 64.3%, 50.0%, and 41.7%, respectively, with the areas under ROC curves of 0.788, 0.734, 0.862, 0.720, 0.887, and 0.585, respectively. In patients with lung cancer, the positive rate of the single indicator CEA was 28.4%, which increased to 60.6% after combined use with TAT-2. In patients with cholangiocarcinoma, the positive rate of CA-199 was 35.7%, which increased to 71.4% after combined use with TAT-2. Conclusions: TAT-2 is expected to be used as an auxiliary diagnostic indicator for the combined use of tumor markers to improve the positive rate and accuracy of detection.

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Ultrasensitive Thyrotropin Immunoassay Based on Enzymatically Amplified Time-Resolved Fluorescence with a Terbium Chelate

Clinical Chemistry ◽

10.1093/clinchem/38.4.545 ◽

1992 ◽

Vol 38 (4) ◽

pp. 545-548 ◽

Cited By ~ 24

Author(s):

A Papanastasiou-Diamandi ◽

T K Christopoulos ◽

E P Diamandis

Keyword(s):

Alkaline Phosphatase ◽

Detection Limit ◽

Solid Phase ◽

Thyroid Stimulating Hormone ◽

Fluorescence Immunoassay ◽

Time Resolved Fluorescence ◽

Time Resolved ◽

Highly Sensitive ◽

Assay Time ◽

Highly Sensitive Detection

Abstract We describe an ultrasensitive, enzymatically amplified time-resolved fluorescence immunoassay of thyrotropin (thyroid-stimulating hormone) in serum with use of a terbium chelate as the detectable moiety. In this assay, thyrotropin is first simultaneously reacted with a solid-phase (microtiter well) monoclonal antibody and a soluble biotinylated monoclonal detection antibody. After washing, a streptavidin-alkaline phosphatase conjugate is added, followed by another washing. Alkaline phosphatase acts on the substrate 5-fluorosalicyl phosphate (FSAP) to produce 5-fluorosalicylic acid (FSA). FSA, but not FSAP, can then form with Tb3+ and EDTA a highly fluorescent ternary complex of long fluorescence lifetime. This complex is quantified with time-resolved fluorometry. The thyrotropin assay is highly sensitive (detection limit approximately 0.003 milli-int. unit/L when a total assay time of 85 min is used), precise, and accurate. The thyrotropin assay can also be completed in less than 30 min (detection limit 0.013 milli-int. unit/L), thus making this procedure a candidate technology for high-throughput automated analyzers.

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Multiple labeling and time-resolvable fluorophores

Clinical Chemistry ◽

10.1093/clinchem/37.9.1486 ◽

1991 ◽

Vol 37 (9) ◽

pp. 1486-1491 ◽

Cited By ~ 34

Author(s):

E P Diamandis

Keyword(s):

Dicarboxylic Acid ◽

Detection Limits ◽

Alpha Fetoprotein ◽

Macromolecular Complex ◽

Fluorescence Immunoassay ◽

Time Resolved Fluorescence ◽

Time Resolved ◽

Highly Sensitive ◽

Europium Chelate ◽

New Time

Abstract A new time-resolved fluorescence immunoassay system involving use of the europium chelate of 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9-dicarboxylic acid as label is reviewed. This stable chelate by itself is not very fluorescent but, used in multiple labeling strategies, improves the achievable detection limits. By using multiple labeling, streptavidin tailing, and Eu3+ activation, one can create a very stable, easy-to-use reagent that is suitable for devising highly sensitive immunoassays and other biotechnological assays. This reagent, a streptavidin-based macromolecular complex, is able to detect approximately 300,000 molecules (approximately 0.5 amol) of alpha-fetoprotein in a model noncompetitive immunoassay.

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