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Gold quantum dots sensitively detect proteins

BioPhotonics
Jan 2007
David L. Shenkenberg

More sensitive detection of troponin-1 protein, which increases during the onset of a heart attack, could allow doctors to intercept cardiac attacks early and prevent significant damage to the heart. An assay based on gold quantum dots may be a preliminary step toward more precise diagnostics that can provide early indications of clinical conditions, diseases and emergencies such as heart attacks.

Enzyme-linked immunosorbent assays are commonly used to measure protein levels, but they lack the sensitivity that immunofluorescence assays can offer. For immunofluorescence assays, quantum dots can be ideal fluorophores because of their high quantum yield and photostability. Therefore, researchers at the University of Miami, at MP Biomedicals LLC in Irvine, Calif., at Trinity College in Dublin, Ireland, at Veeco Instruments Inc. in Santa Barbara, Calif., and at the University of California, Irvine, developed an assay that uses gold quantum dots to determine protein levels.

To show that the assay can detect proteins, the scientists joined the quantum dots to an antibody that binds to human immunoglobulin G. Because they envisioned that people who use this assay could monitor changes in fluorescence to ascertain protein levels, they examined whether adding various levels of human immunoglobulin G alters the level of measured fluorescence.

The researchers chose gold quantum dots because they are more economical and less toxic than cadmium ones. They encapsulated the quantum dots with branched polymers, called poly(amido) amine dendrimers, because those polymers enabled the scientist to precisely control the quantum dots’ size. A size of <5 nm allows gold quantum dots to retain their high photoluminescence intensities, tunable emission frequencies and high photostability.

The fact that the dendrimers added to the quantum dots’ size had to be considered. To characterize the size of the dendrimer-gold quantum dot complexes, the researchers employed a Veeco Instruments atomic force microscope, a Philips transmission electron microscope and a PerkinElmer UV-VIS spectrometer. The complexes ranged from 4 to 6 nm in size. Thus, most complexes were small enough to exhibit the desired photophysical properties.

BNImmuno_AFM.jpg
Researchers used gold quantum dots, represented by the bumps in this atomic force microscopy image, to detect nanomolar to micromolar protein concentrations.

For fluorescence measurements, the researchers used a Horiba Jobin Yvon spectrometer. The quantum dots had excitation and emission maxima of 387 and 450 nm, respectively, whether joined to the antibody or separate. Nanomolar to micromolar levels of human immunoglobulin G quenched the fluorescence of the gold quantum dot complexes in a linear fashion, demonstrating that human immunoglobulin G levels can be detected by measuring fluorescence.

Miodrag Micic, one of the study’s authors, said that any antibody other than human immunoglobulin G can be joined to the quantum dots, enabling detection of any protein. The broad dynamic range allows for analysis of various concentrations of proteins. The assay is sensitive to nanomolar concentrations, so it can detect a few protein molecules in samples. Micic said that such sensitivity also can be critical when the sample volume is limited, such as in neonatal medicine and in pharmacological studies using mice. In those cases, low-volume samples such as a finger prick of blood are often taken in lieu of larger samples such as venous blood to minimize invasiveness.

Micic said that the gold quantum dot complexes could be used for making biosensors and for histochemical labeling for confocal and fluorescence microscopy. Their low toxicity could enable them to be used for in vivo imaging.

He noted that the researchers immediately plan to join the gold quantum dot complexes to avidin. Avidin naturally binds to biotin, and biotin can be easily linked to any antibody. Therefore, avidin and biotin allow researchers to construct the gold-quantum-dot-based assay for detecting any protein, without knowledge of complicated antibody conjugation methods.

The scientists also are developing an assay platform, based on the tunable emission property of gold quantum dots, that will allow simultaneous detection of multiple analytes. Finally, they will seek FDA approval for selected protein biomarker detection assays.

Chemical Communications, online, Nov. 21, 2006, 10.1039/b611278a.


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