Flow cytometric analysis with multicolor fluoroprobes is an essential method for detecting biological signatures of cells. Here, we present a new full‐spectral flow cytometer (spectral‐FCM). Unlike conventional flow cytometer, this spectral‐FCM acquires the emitted fluorescence for all probes across the full‐spectrum from each cell with 32 channels sequential PMT unit after dispersion with prism, and extracts the signals of each fluoroprobe based on the spectral shape of each fluoroprobe using unique algorithm in high speed, high sensitive, accurate, automatic and real‐time. The spectral‐FCM detects the continuous changes in emission spectra from green to red of the photoconvertible protein, KikGR with high‐spectral resolution and separates spectrally‐adjacent fluoroprobes, such as FITC (Emission peak (Em) 519 nm) and EGFP (Em 507 nm). Moreover, the spectral‐FCM can measure and subtract autofluorescence of each cell providing increased signal‐to‐noise ratios and improved resolution of dim samples, which leads to a transformative technology for investigation of single cell state and function. These advances make it possible to perform 11‐color fluorescence analysis to visualize movement of multilinage immune cells by using KikGR‐expressing mice. Thus, the novel spectral flow cytometry improves the combinational use of spectrally‐adjacent various FPs and multicolor fluorochromes in metabolically active cell for the investigation of not only the immune system but also other research and clinical fields of use. © 2015 The Authors. Cytometry Part A Published by Wiley Periodicals, Inc. on behalf of ISAC

In order to provide a protein adsorption resistant surface even when the surface was in contact with a protein solution under completely dry conditions, a new phospholipid copolymer, poly (2-methacryloyloxyethyl phosphorylcholine (MPC)- co-2-vinylnaphthalene (vN)) (PMvN), was synthesized. Poly(ethylene terephthalate) (PET) could be readily coated with PMvN by a solvent evaporation method. Dynamic contact angle measurements with water revealed that the surface was wetted very rapidly and had strong hydrophilic characteristics; moreover, molecular mobility at the surface was extremely low. When the surface came in contact with a plasma protein solution containing bovine serum albumin (BSA), the amounts of the plasma protein adsorbed on the dry surface coated with PMvN and that adsorbed on a dry surface coated with poly(MPC-co-n-butyl methacrylate) (PMB) were compared. Substantially lower protein adsorption was observed with PMvN coating. This is due to the rapid hydration behavior of PMvN. We concluded that PMvN can be used as a functional coating material for medical devices without any wetting pretreatment.

No abstract

We have been revealing immune system based of spatiotemporal regulation of immune cells in the entire body by using photoconvertible fluorescent protein (FP), Kaede and KikGR mice. Flow cytometric analysis of green and red signals of non-photoconverted and photoconverted Kaede or KikGR expressing cells with multiple fluorochrome-conjugated mAbs have been difficult and unable to increase color panels, because of the fluorescence signal overlap between FPs and fluorochromes. We have newly developed a spectral flow cytometer (FCM) based on a novel measurement principle. Unlike a polychromatic FCM, the spectral FCM with 32 channel linear array PMT detects the fluorescence derived from every fluorescent probe. The data of acquired spectra is analyzed with a unique algorithm. We detected spectral changes of KikGR color from green to red during photoconversion. We separated EGFP and Venus. One of the feature advantages of this instrument is that it can recognize spectral shape of each fluorescent probes and we could separate EGFP and FITC, their peaks of wavelength are almost same and only spectral shape of GFP is slightly broader than FITC. Finally, we successfully separated 11-colors including kikGR-green and -red simultaneously. Taken together, spectral FCM allow us to detection of FPs with multicolor fluorochromes, and thus, the system which uses KikGR mice with spectral FCM is a powerful tool to investigate spatiotemporal regulation of immune cells in the entire body.