Bioluminescence immunoassay for angiotensin II using aequorin as a label

Qu, Xiaoge; Deo, Sapna K.; Dikici, Emre; Ensor, Mark; Poon, Michael; Daunert, Sylvia

doi:10.1016/j.ab.2007.08.038

Cited by 13 publications

(3 citation statements)

References 31 publications

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“…In 2007, Qu and associates have improved the detection limit of endogenous Ang II by developing a bioluminescence immunoassay. They obtained a detection limit of 1 pM, allowing for the determination of Ang II in human plasma 50. Although detection limit of our method is higher than that of their method, our method needs very small amount of sample and short analysis time.…”

Section: Resultsmentioning

confidence: 82%

A Facile Synthetic Route to 2 H‐Chromenes: Reconsideration of the Mechanism of the DBU‐Catalyzed Reaction between Salicylic Aldehydes and Ethyl 2‐Methylbuta‐2,3‐dienoate

Dai

Shi

Zhao

et al. 2007

Chemistry A European J

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Reactions of salicylaldehydes with ethyl buta-2,3-dienoate or penta-3,4-dien-2-one catalyzed by a catalytic amount of potassium carbonate produce the corresponding 2 H-chromene products in moderate to good yields under mild conditions. A plausible reaction mechanism is discussed in the light of the results of an 18O-labeling experiment. In addition, in view of these findings, the catalytic function of DBU in reactions of this kind is reconsidered.

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Section: Resultsmentioning

confidence: 82%

A Facile Synthetic Route to 2 H‐Chromenes: Reconsideration of the Mechanism of the DBU‐Catalyzed Reaction between Salicylic Aldehydes and Ethyl 2‐Methylbuta‐2,3‐dienoate

Dai

Shi

Zhao

et al. 2007

Chemistry A European J

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“…Furthermore, our group and that of others have demonstrated that AEQ can be detected at attomole levels . Due to the remarkably low limits of detection achieved when aequorin is used, this photoprotein has been employed in many bioanalytical applications, including, as a calcium indicator in living cells and for in vivo imaging, , as a reporter in DNA hybridization assays , and immunoassays, − and as the signal generating molecule in molecular switch sensing systems. , Herein, we have developed a sensitive, AEQ-based bioluminescence inhibition assay for the detection of OH-PCBs. A unique feature of this inhibition assay is that the AEQ plays a dual role; namely, it is the recognition as well as the signal generating molecule.…”

Section: Resultsmentioning

confidence: 99%

Bioluminescence Inhibition Assay for the Detection of Hydroxylated Polychlorinated Biphenyls

et al. 2012

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Hydroxylated polychlorinated biphenyls (OH-PCBs) are an important class of contaminants that mainly originate from polychlorinated biphenyl metabolism. They may conceivably be as dangerous and persistent as the parent compounds; most prominently, OH-PCBs are endocrine disruptors. Due to increasing evidence of the presence of OH-PCBs in the environment and in living organisms, including humans, and of their toxicity, methods of detection for OH-PCBs are needed in the environmental and medical fields. Herein we describe the development and optimization of a protein+based inhibition assay for the quantification of OH-PCBs. Specifically, the photoprotein aequorin was utilized for the detection of OH-PCBs. We hypothesized that OH-PCBs interact with aequorin and we established that OH-PCBs actually inhibit the bioluminescence of aequorin in a dose-dependent manner. We took advantage of this phenomenon to develop an assay that is capable of detecting a wide variety of OH-PCBs with a range of detection limits, the best detection limit being 11 nM for the compound 2-hydroxy-2',3,4',5',6-pentachorobiphenyl. The viability of this system for the screening of OH-PCBs in spiked biological and environmental samples was also established. We envision the implementation of this novel bioluminescence inhibition assay as a rapid, sensitive and cost-effective method for monitoring OH-PCBs. Furthermore, to the best of our knowledge this is the first time aequorin has been employed to detect an analyte by the inhibition of its bioluminescence reaction. Hence, this strategy may prove to be a general approach for the development of a new generation of protein-based inhibition assays.

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“…The angiotensin II-aequorin fusion protein was constructed by adding a six amino acid spacer sequence and the eight amino acid sequence of angiotensin II (DRVYIHPF) to the N terminus of the cysteine free aequorin mutant using primers and PCR amplification and purified using ion exchange chromatography (See ref for method details.) . The purified angiotensin II-aequorin fusion protein was then charged with a 5 M excess of ctz i for 16 h at 4 °C.…”

Section: Methodsmentioning

confidence: 99%

Genetically Modified Semisynthetic Bioluminescent Photoprotein Variants: Simultaneous Dual-Analyte Assay in a Single Well Employing Time Resolution of Decay Kinetics

et al. 2008

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Progress in the miniaturization and automation of complex analytical processes depends largely on increasing the sensitivity, diversity, and robustness of current labels. Because of their ubiquity and ease of use, fluorescent, enzymatic, and bioluminescent labels are often employed in such miniaturized and multiplexed formats, with each type of label having its own unique advantages and drawbacks. The ultrasensitive detection limits of bioluminescent reporters are especially advantageous when dealing with very small sample volumes and biological fluids. However, bioluminescent reporters currently do not have the multiplexing capability that fluorescent labels do. In an effort to address this limitation, we have developed a method of discriminating two semisynthetic aequorin variants from one another using time resolution. In this work we paired two aequorin conjugates with different coelenterazine analogues and then resolved the two signals from one another using the difference in decay kinetics and half-life times. Utilizing this time-resolution, we then developed a simultaneous, dual-analyte, single well assay for 6-keto-prostaglandin-FI-alpha and angiotensin II, two important cardiovascular molecules.

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Bioluminescence immunoassay for angiotensin II using aequorin as a label

Cited by 13 publications

References 31 publications

A Facile Synthetic Route to 2 H‐Chromenes: Reconsideration of the Mechanism of the DBU‐Catalyzed Reaction between Salicylic Aldehydes and Ethyl 2‐Methylbuta‐2,3‐dienoate

A Facile Synthetic Route to 2 H‐Chromenes: Reconsideration of the Mechanism of the DBU‐Catalyzed Reaction between Salicylic Aldehydes and Ethyl 2‐Methylbuta‐2,3‐dienoate

Bioluminescence Inhibition Assay for the Detection of Hydroxylated Polychlorinated Biphenyls

Genetically Modified Semisynthetic Bioluminescent Photoprotein Variants: Simultaneous Dual-Analyte Assay in a Single Well Employing Time Resolution of Decay Kinetics

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