Development of Gold Nanomaterials and Microgels Based Hybrid Materials for Bioanalysis Application
Date Issued
2014
Date
2014
Author(s)
Chen, Li-Yi
Abstract
This dissertation focuses on developments of high sensitivity and selectivity optical probes for the detection of proteins and heavy metal ions in biological sample, including photoluminescent gold nanodots and hybrid nanomaterials with microgels. The functional microgel nanomaterials (trypsin/gold nanoparticle/microgels) is further applied to detect proteins through surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS) in conjugation with an internal standard. The dissertation was structured with four chapters. In Chapter 1, the detailed background of photoluminecent gold nanomaterials and microgels, and basic principle of soft ionization techniques of mass spectrometry were widely described. In Chapter 2, the developed approach, using photoluminescent 11-mercaptoundecanoic acid–gold nanodots (11-MUA–Au NDs), was applied for the detection of hemoglobin through photoluminescence (PL) quenching. The mechanism of quenching, which occurred through redox reactions between the 11-MUA–Au NDs and the Fe(II) atoms of hemin units, was supported by an increase in the signals (G 2.0 and 5.9) of high-spin state Fe(III) ions. This approach using bovine serum albumin blocked 11-MUA–Au NDs provided good sensitivity for hemoglobin of 0.5 nM in biological buffer and with great selectivity over other non-heme–containing proteins. Furthermore, we also validated the practicality of this approach through the determination of the concentrations of hemoglobin in diluted human blood samples. In Chapter 3, poly(N-isopropylacrylamide) microgels (PNIPAM MGs) incorporated with photoluminescent Au NDs had been prepared and used for the detection of mercury ions (Hg2+). Based on Hg2+-induced photoluminescence quenching due to the formation of Au–Hg amalgam and formation of Au NDs–PNIPAM MGs aggregates, this approach provided limits of detection for Hg2+ of 1.9 nM in biological buffer solutions containing 500 mM NaCl. This selective and sensitive Au NDs–PNIPAM MG probe has been applied to the determination of the concentration of Hg2+ in a representative fish sample. In Chapter 4, temperature sensitive MGs containing trypsin and gold nanoparticles have been employed for the digestion of cytochrome c (Cyt c) under microwave irradiation. For further quantitation of Cyt c in cell lysates, SALDI-MS was applied for detection by using gold nanoparticles as the matrix and GR-10 peptide as an internal standard. The internal standard SALDI-MS approach provided linearity (R2 = 0.98) of MS signal ratio (I1168.6/I1067.6) of the tryptic digested peptide (m/z 1168.6) to GR-10 peptide (m/z 1067.6), with a limit of detection of 10 nM. This approach has been validated by the analysis of the concentration of cytoplasmic Cyt c the lysates of HeLa cells.
Subjects
photoluminescent gold nanodots (Au NDs)
gold nanoparticles (Au NPs)
poly(N-isopropylacrylamide)
microgels
hemoglobin
cytochrome c
mercury ion
surface-assisted laser desorption/ionization mass spectrometry (SALDI-MS)
Type
thesis
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