Simultaneous Bioluminescent Immunoassay of Serum Total and IgG-Bound Prolactins

详细信息    查看全文

• 作者：Alexander N. Kudryavtsev ; Vasilisa V. Krasitskaya ; Alexei I. Petunin ; Andrey Y. Burakov ; Ludmila A. Frank
• 刊名：Analytical Chemistry
• 出版年：2012
• 出版时间：April 3, 2012
• 年：2012
• 卷：84
• 期：7
• 页码：3119-3124
• 全文大小：322K
• 年卷期：v.84,no.7(April 3, 2012)
• ISSN：1520-6882

文摘

Novel dual-analyte single-well bioluminescence immunoassay (BLIA) for total and IgG-bound prolactins was developed on the base of Ca²⁺-regulated photoprotein obelin mutants with altered color and kinetics of bioluminescence signal as reporters. The mutants W92F-H22E and Y138F were chemically conjugated with monoclonal mouse anti-hPRL and anti-hIgG immunoglobulins and thus displayed signals from total prolactin and IgG-bounded prolactin (macroprolactin) correspondingly. Bioluminescence of the reporters was simultaneously triggered by a single injection of Ca²⁺ solution and discriminated via bioluminescent signal spectral and time resolution. The developed microplate-based immunoassay allows detection of two prolactin forms in crude serum without additional manipulations (e.g., gel chromatography or PEG-precipitation). Total prolactin bioluminescence immunoassay in standard, control, and clinical sera offers high sensitivity and reproducibility. The BLIA results show good correlation with those obtained by RIA and immunoassay after gel chromatography.