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Fast and ultrafast thermal contrast amplification of gold nanoparticle-based immunoassays.

ABSTRACT: For highly sensitive point-of-care (POC) diagnostics, we explored the limit of thermal contrast amplification (TCA) reading of gold nanoparticles (GNPs/mm²) at test regions in immunoassays. More specifically, we built and compared fast (minute scale) and ultrafast (seconds scale) TCA setups using continuous-wave (CW) and ms pulsed lasers, respectively. TCA improved the limit of detection (LoD) for silica-core gold nanoshells (GNSs) preloaded in nitrocellulose (NC) membrane as model lateral flow immunoassays (LFAs) by 10- to 20-fold over visual reading. While the ultrafast TCA led to higher thermal signals, this came with a twofold loss in LoD vs. fast TCA primarily due to noise within the infrared sensor and a necessity to limit power to avoid burning. To allow higher laser power, and therefore amplification fold, we also explored transparent glass coverslip substrate as a model microfluidic immunoassay (MIA). We found the ultrafast TCA reading of GNS-coated coverslips achieved a maximal signal amplification (57-fold) over visual reading of model LFAs. Therefore, ultrafast TCA-MIA is promising for ultrasensitive and ultrafast diagnostics. Further advantages of using TCA in MIA vs. LFA could include lower sample volume, multiplexed tests, higher throughput, and fast reading. In summary, TCA technology is able to enhance the sensitivity and speed of reading GNPs (GNPs/mm²) within both LFAs and MIAs.

SUBMITTER: Liu Y

PROVIDER: S-EPMC9321340 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Detection of human immunodeficiency virus (HIV) p24 protein at a single pg/ml concentration in point-of-care (POC) settings is important because it can facilitate acute HIV infection diagnosis with a detection sensitivity approaching that of laboratory-based assays. However, the limit of detection (LOD) of lateral flow immunoassays (LFAs), the most prominent POC diagnostic platform, falls short of that of laboratory protein detection methods such as enzyme-linked immunosorbent assay (ELISA). Here, we report the development and optimization of a thermal contrast amplification (TCA) LFA that will allow ultrasensitive detection of 8 pg/ml p24 protein spiked into human serum at POC, approaching the LOD of a laboratory test. To achieve this aim, we pursued several innovations as follows: (a) defining a new quantitative figure of merit for LFA design based on the specific to nonspecific binding ratio (BR); (b) using different sizes and shapes of gold nanoparticles (GNPs) in the systematic optimization of TCA LFA designs; and (c) exploring new laser wavelengths and power regimes for TCA LFA designs. First, we optimized the blocking buffer for the membrane and running buffer by quantitatively measuring the BR using a TCA reader. The TCA reader interprets the thermal signal (i.e., temperature) of GNPs within the membrane when irradiated by a laser at the plasmon resonance wavelength of the particle. This process results in higher detection and quantitation of GNPs than in traditional visual detection (i.e., color intensity). Further, we investigated the effect of laser power (30, 100, 200 mW), GNP size and shape (30 and 100 nm gold spheres, 150 nm gold-silica shells), and laser wavelength (532, 800 nm). Applying these innovations to a new TCA LFA design, we demonstrated that 100 nm spheres with a 100 mW 532 nm laser provided the best performance (i.e., LOD = 8 pg/ml). This LOD is significantly better than that of the current colorimetric LFA and is in the range of the laboratory-based p24 ELISA. In summary, this TCA LFA for p24 protein shows promise for detecting acute HIV infection in POC settings.

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Fast and ultrafast thermal contrast amplification of gold nanoparticle-based immunoassays.

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