Immunochromatographic analysis of bovine growth hormone releasing factor involving reversed-phase high-performance liquid chromatography-immunodetection
| Autor: | Ira S. Krull, Daniel H. Fisher, Byung-Yun Cho, Hanfa Zou, Richard A. Strong, John L. Nappier |
|---|---|
| Rok vydání: | 1996 |
| Předmět: |
Streptavidin
Metabolite Biotin Succinimides Enzyme-Linked Immunosorbent Assay Growth Hormone-Releasing Hormone Online Systems Sensitivity and Specificity Biochemistry High-performance liquid chromatography Chromatography Affinity Analytical Chemistry Plasma chemistry.chemical_compound Bacterial Proteins Affinity chromatography Animals Trypsin Chromatography High Pressure Liquid Chromatography Chemistry Immune Sera Organic Chemistry Antibodies Monoclonal Reproducibility of Results General Medicine Reversed-phase chromatography Fluorescence Peptide Fragments Spectrometry Fluorescence Calibration Isothiocyanate Cattle Spectrophotometry Ultraviolet Quantitative analysis (chemistry) Fluorescein-5-isothiocyanate |
| Zdroj: | Journal of Chromatography A. 743:181-194 |
| ISSN: | 0021-9673 |
| DOI: | 10.1016/0021-9673(96)00356-1 |
| Popis: | We have developed high-performance immunoaffinity chromatography (HPIAC) methods for the detection and quantitation of bovine growth hormone releasing factor (GHRF), which could also be applicable to its metabolites in biofluids. These approaches have involved a combination of IAC using immobilized antibody (Ab) to GHRF, together with reversed-phase high-performance liquid chromatography (RP-HPLC) separations of initially isolated and concentrated protein, followed by selective detection, involving on-line immunodetection (ID) schemes. ID methods involved HPIAC supports of the Ab, together with synthesized Ab-fluorescein isothiocyanate conjugates. We have demonstrated optimization methods for each step of the entire hyphenated technique (IAC-HPLC-ID), and then actually quantitated GHRF using this overall system. The minimum detectable concentration was about 1 ng/5 ml (200 ppt) with fluorescence detection (excitation wavelength, 490 nm; emission wavelength, 510-650 nm). We have also tested a single blind, spiked biological sample (bovine plasma), spiked with a known level of GHRF. Accuracy (7.4%) and precision (S.D. = +/- 22%) were quite acceptable for a double immunoassay method. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full Text from ScienceDirect