The appropriate sample-handling procedure for measuring the plasma β-amyloid level using a fully automated immunoassay.

Plasma β-amyloid (Aβ) assays are a promising tool for Alzheimer's disease diagnosis in clinical practice. To obtain reliable results, establishing an appropriate sample-handling procedure for each analytical platform is warranted. This study proposes an appropriate sample-handling procedure using HISCL analyzer by elucidating the individual/combined effects of pre-analytical parameters on plasma Aβ42/Aβ40 levels.

Hybrid double-spiral microfluidic chip for RBC-lysis-free enrichment of rare cells from whole blood.

Drug selection and treatment monitoring minimally invasive liquid biopsy using circulating tumor cells (CTCs) are expected to be realized in the near future. For clinical applications of CTCs, simple, high-throughput, single-step CTC isolation from whole blood without red blood cell (RBC) lysis and centrifugation remains a crucial challenge. In this study, we developed a novel cancer cell separation chip, "hybrid double-spiral chip", that involves the serial combination of two different Dean flow fractionation (DFF) separation modes of half and full Dean cycles, which is the hybrid DFF separation mode for ultra-high-throughput blood processing at high precision and size-resolution separation.

Fully automated chemiluminescence enzyme immunoassays showing high correlation with immunoprecipitation mass spectrometry assays for β-amyloid (1-40) and (1-42) in plasma samples.

Blood based β-amyloid (Aβ) assays that can predict amyloid positivity in the brain are in high demand. Current studies that utilize immunoprecipitation mass spectrometry assay (IP-MS), which has high specificity for measuring analytes, have revealed that precise plasma Aβ assays have the potential to detect amyloid positivity in the brain. In this study, we developed plasma Aβ40 and Aβ42 immunoassays using a fully automated immunoassay platform that is used in routine clinical practice.

Silinanyl Rhodamines and Silinanyl Fluoresceins for Super-Resolution Microscopy.

Single-molecule localization microscopy (SMLM) enables the visualization of biomolecules at unprecedented resolution and requires control of the fluorescent blinking (ON/OFF) states of fluorophores to detect single-molecule fluorescence without overlapping of the signals. Although SMLM probes based on the intramolecular spirocyclization of Si-xanthene fluorophores have been developed, fluorophores with lower ON/OFF ratios are required for SMLM visualization of high-density structures. Here, we describe a silinane structure that lowers the ON/OFF ratio of Si-xanthene fluorophores.

Quantification of Amyloid-β in Plasma by Simple and Highly Sensitive Immunoaffinity Enrichment and LC-MS/MS Assay.

Background: Numerous immunoassays have been developed to quantify amyloid β1-40 (Aβ40) and amyloid β1-42 (Aβ42). Nevertheless, given the low concentration of Aβ and the high levels of interfering factors in plasma, quantification of plasma Aβ is still challenging. To overcome the problems related to the specificity of Aβ immunoassays, this study aimed to develop an immunoaffinity enrichment and LC-MS/MS (IA-MS) assay.

Integrated system for detection and molecular characterization of circulating tumor cells.

Circulating tumor cells (CTCs) invade blood vessels in solid tumors and promote metastases by circulating in the blood. CTCs are thus recognized as targets for liquid biopsy and can provide useful information for design of treatments. This diagnostic approach must consider not only the number of CTCs but also their molecular and genetic characteristics.

Fluorescent saxitoxins for live cell imaging of single voltage-gated sodium ion channels beyond the optical diffraction limit.

A desire to better understand the role of voltage-gated sodium channels (Na(V)s) in signal conduction and their dysregulation in specific disease states motivates the development of high precision tools for their study. Nature has evolved a collection of small molecule agents, including the shellfish poison (+)-saxitoxin, that bind to the extracellular pore of select Na(V) isoforms. As described in this report, de novo chemical synthesis has enabled the preparation of fluorescently labeled derivatives of (+)-saxitoxin, STX-Cy5, and STX-DCDHF, which display reversible binding to Na(V)s in live cells.

Superresolution imaging of targeted proteins in fixed and living cells using photoactivatable organic fluorophores.

Superresolution imaging techniques based on sequential imaging of sparse subsets of single molecules require fluorophores whose emission can be photoactivated or photoswitched. Because typical organic fluorophores can emit significantly more photons than average fluorescent proteins, organic fluorophores have a potential advantage in super-resolution imaging schemes, but targeting to specific cellular proteins must be provided. We report the design and application of HaloTag-based target-specific azido DCDHFs, a class of photoactivatable push-pull fluorogens which produce bright fluorescent labels suitable for single-molecule superresolution imaging in live bacterial and fixed mammalian cells.

Location-dependent photogeneration of calcium waves in HeLa cells.

The calcium ion (Ca(2+)) concentrations in a cell are responsible for the control of vital cellular functions and have been widely studied as a means to investigate and control cell activities. Here, we demonstrate Ca(2+) wave generation in HeLa cells by femtosecond laser irradiation and show unexpected properties of the Ca(2+) release and propagation. When the laser was focused in the cell cytoplasm, Ca(2+) release was independent of both external Ca(2+) influx and the phosphoinositide-phospholipase C (PLC) signaling pathway.