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Intensity Histogram-Based Reliable Image Analysis Method for Bead-Based Fluorescence Immunoassay
Lee Sanghyun,Kim Juyeong,Bae Pan Kee,Lee Sangmin,김호진 한국바이오칩학회 2024 BioChip Journal Vol.18 No.1
Bead-based fluorescence immunoassay is drawing attention as a next-generation technology in disease diagnosis owing to its high sensitivity and multiplexing capability. Fluorescence imaging of beads is typically used to determine their mean fluorescence intensity. However, the mean intensity can be evaluated differently depending on the analysis methods [such as the shape and size of the region of interest (ROI)]. To address these problems, this study proposes a highly reliable and reproducible image analysis method utilizing a fluorescence intensity-based effective pixel extraction technique. Various potential sources of defective signals (e.g., fluorescence aggregation, non-specific antigen–antibody reactions, and bead defects) can be prevented from contributing to the average value by selectively extracting pixels representing the specific reactions of antigens and antibodies in the ROI. In this study, we fabricated a microfluidic chip composed of multiple bead- based detection lines, performed fluorescence immunoassay, and then compared the mean fluorescence intensity calculated from the fluorescence images with that of a conventional analysis method. Using the conventional method, the evaluated average mean intensity value of beads varied significantly based on the size of the ROI with the coeffi cients of variation ranging from approximately 29–95%. In contrast, the effective pixel extraction method resulted in a coefficient of variation of approximately 3–7% under varying ROI size. Furthermore, the coefficients of variation for four detection lines containing various types of defective signals signifi cantly decreased from approximately 7.1% to 2.6%. The proposed technique will help in minimizing the analysis deviation caused by different ROI selections or defective signals in fluorescent image-based immunoassays.
Lee, Chang‐,Soo,Chang, Hee Hyun,Bae, Pan‐,Kee,Jung, Juyeon,Chung, Bong Hyun WILEY‐VCH Verlag 2013 Macromolecular bioscience Vol.13 No.3
<P><B>Abstract</B></P><P>Integration of biocompatible silica with a fluorescent polymer (PDDF) and superparamagnetic iron oxide nanoparticles (Fe<SUB>3</SUB>O<SUB>4</SUB>) to form uniform core–shell nanostructures has the great potential to form particles for use in multimodal bioimaging applications. Core–shell nanoparticles (PDDF/Fe<SUB>3</SUB>O<SUB>4</SUB>@SiO<SUB>2</SUB>) exhibit fluorescent and magnetic properties that are favorable for their use in magnetic separation and guiding applications, as well as optical and magnetic resonance (MR) imaging capabilities. With the biological analysis in an in vitro intracellular permeation and cytotoxicity test, chemical conjugation of the surface using folic acid (FA) molecules can provide the nanoparticles with cell‐targeting properties, localizing the nanoparticles to folate receptors (FRs) on target KB cells that over‐express the FRs. </P>
Synthesis of Fe3O4-ZnS/AgInS2 composite nanoparticles using a hydrophobic interaction.
Choi, Kang Sik,Bang, Bo Keuk,Bae, Pan Kee,Kim, Yong-Rok,Kim, Chang Hae American Scientific Publishers 2013 Journal of Nanoscience and Nanotechnology Vol.13 No.3
<P>Magnetic nanoparticles and fluorescent quantum dots (QDs) can make many effective applications in biomedical system. Here, we demonstrated one way of synthetic method and its surface modification to use for biomedical applications. Fe3O4 nanoparticles are well known as magnetic materials and its magnetic property can be used in magnetic resonance imaging (MRI), cell detection. QDs as a fluorescent probes, make cell labeling and in vivo imaging possible. ZnS/AgInS2 QDs have a lower toxicity than other QDs (CdSe, CdTe, CdS). We combined two nanoparticles by hydrophobic interaction in their ligands. The prepared fluorescent magnetic composite particles were modified with CTAB-TEOS. The surface modified composite has a low cytotoxicity and these biocompatible particles will provide many possibilities in biomedical system.</P>