Orientation of Antibody Modified and Reacted on Carboxy-Functionalized Polystyrene Particle Revealed by Zeta Potential Measurement.

The comprehensive understanding of the orientation of antibodies on a solid surface is crucial for affinity-based sensing mechanisms. In this study, we demonstrated that the orientation of primary antibodies modified on carboxy-functionalized polystyrene (PS) particles can be analyzed using zeta potential behavior at different pH based on the combined Gouy-Chapman-Stern model and the acid dissociation of carboxy groups and antibodies. We observed that at low surface concentrations of the primary antibody, a side-on orientation was predominant.

Effect of Organic Solvent on the Mass Transfer Mechanism of Coumarin 102 in a Single Octadecylsilyl Silica Gel/Organic Solvent-Water System by Laser Trapping and Fluorescence Microspectroscopy.

Understanding mass transfer kinetics within individual porous particles is crucial for theoretically explaining the retention and elution behaviors in chromatography and drug delivery. Using laser trapping and fluorescence microspectroscopy, we investigated the diffusion mechanism of coumarin 102 (C102) into single octadecylsilyl particle in acetonitrile (ACN)/water, ,-dimethylformamide (DMF)/water, and 1-butanol (BuOH)/water solutions. The intraparticle diffusion behavior of C102 was evaluated using the spherical diffusion equation, allowing us to determine the intraparticle diffusion coefficients (): (8-10) × 10 cm s for ACN, (10-16) × 10 cm s for DMF, and (4-6) × 10 cm s for BuOH.

Diffusion behavior of rhodamine 6G in single octadecylsilyl-functionalized silica particle revealed by fluorescence correlation spectroscopy.

We investigated the diffusion behavior of rhodamine 6G (Rh6G) within single octadecylsilyl-functionalized (ODS) silica particle in an acetonitrile (ACN)/water system using fluorescence correlation spectroscopy (FCS). FCS measurements were conducted at the center of the particle to exclusively determine the intraparticle diffusion coefficient (D). The obtained D values were analyzed based on a pore and surface diffusion model, the results of which indicate that surface diffusion primarily governs the intraparticle diffusion of Rh6G.

Evolution of myoglobin diffusion mechanisms: exploring pore and surface diffusion in a single silica particle.

This study elucidates the mass transfer mechanism of myoglobin (Mb) within a single silica particle with a 50 nm pore size at various pH levels (6.0, 6.5, 6.

Dynamic Interfacial Tension Measurement for the Kinetic Study of Eu(III) Extraction in the NaNO/Tributyl Phosphate System Based on an Increase in Interfacial Tension.

We demonstrated for the first time that interfacial tension measurements can be used to evaluate the kinetics of the solvent extraction of metal ions. The Eu(III) extraction mechanism in the nitrate ion/tributyl phosphate (TBP) system was investigated on the basis of dynamic interfacial tension. Interestingly, the interfacial tension of the TBP droplet () increased with Eu(III) extraction.

Kinetic analysis of mass transfer of Eu(III) in extractant-impregnated polymer-layered silica particle in multiple-ion distribution system.

The Eu(III) distribution mechanism in single extractant-impregnated polymer-layered silica particle in a complex solution containing multiple lanthanide ions was investigated using fluorescence microspectroscopy, which was compared with the single-ion distribution system. The rate-determining step of the Eu(III) distribution was the reaction of Eu(III) with the two extractant molecules in the particle. The distribution mechanism and rate constants obtained in the multiple lanthanide ions-distribution system agreed with those of the single-ion distribution system.

Biosensing of DNA through difference in interaction between microparticle and glass plate based on particle dissociation in a coupled acoustic-gravitational field.

A novel approach for detecting DNA without labeling the target DNA was developed based on the particle dissociation behavior in a combined acoustic-gravitational field. The particles, which are tethered on a glass plate via intermolecular interactions (F), are dissociated by the resultant force of the acoustic radiation force (F), which is a function of the applied voltage (V), and the sedimentation force. In this system, V required for particle dissociation is dependent on F.

Eu(III) transfer in single N,N,N',N'-tetraoctyldiglycolamide-impregnated polymer-coated silica particle using fluorescence microspectroscopy: transfer mechanism and effect of polymer crosslinking degree.

A microcapillary manipulation system combined with fluorescence microspectroscopy enabled us to analyze mass transfer in a single particle. In this study, we revealed the Eu(III) distribution in a single diglycolamide-derivative extractant (TODGA)-impregnated polymer-coated silica particle. The reaction of Eu(III) with two TODGA molecules in the polymer layer was the rate-limiting process, which was revealed by the relationship between the rate constants (k and k) and concentrations of Eu(III) and HNO.

Pore Size Dependence of Mass Transfer of Zinc Myoglobin in a Single Mesoporous Silica Particle.

This study investigated the pore size dependence of the mass transfer of zinc myoglobin (ZnMb) in a single mesoporous silica particle through confocal fluorescence microspectroscopy. The ZnMb's fluorescence depth profile in the particle was analyzed by a spherical diffusion model, and the intraparticle diffusion coefficient was obtained. The intraparticle diffusion coefficient in the silica particle with various pore sizes (10, 15, 30, and 50 nm) was furthermore analyzed based on a pore and surface diffusion model.

Semi-quantification of the binding constant based on bond breaking in a combined acoustic-gravitational field.

In this study, we propose a semi-quantification method based on breaking bonds between microparticles and glass plates in a combined acoustic-gravitational field. The semi-quantified binding constant values for BSA-ibuprofen, BSA-ciprofloxacin, ConA-glycogen, ConA-mannan, and BSA-naproxen calculated using this method were 7.5 × 10, 1.

Distribution Behavior of Single-Stranded DNA Molecules in an Amino-Group-Functionalized Silica Microparticle.

In this study, we investigated the distribution behavior of single-stranded DNA molecules with 20 bases in silica particles (particle size: ∼30 μm) using confocal fluorescence microspectroscopy. The distribution kinetics was investigated under various conditions, such as the type of base (adenine, thymine, guanine, and cytosine), pore size of the particle (30 and 50 nm), and salt concentration (100, 200, and 500 mM), which changed the distribution behavior. At high salt concentrations, we observed sigmoidal kinetic behavior, which does not occur in the general distribution of small organic molecules but is often observed in protein aggregation and nuclear growth.

Kinetics and mechanism of Eu(III) transfer in tributyl phosphate microdroplet/HNO aqueous solution system revealed by fluorescence microspectroscopy.

In this study, we reveal an Eu(III) extraction mechanism at the interface between HNO and tributyl phosphate (TBP) solutions using fluorescence microspectroscopy. The mass transfer rate constant at the interface is obtained from the analysis of fluorescence intensity changes during the forward and backward extractions at various HNO and TBP concentrations to investigate the reaction mechanism. This result indicates that one nitrate ion reacts with Eu(III) at the interface, whereas TBP molecules are not involved in the interfacial reaction, which is different from the results obtained using the NaNO solution in our previous study.

Structural origin of cooperativity in human hemoglobin: a view from different roles of α and β subunits in the αβ tetramer.

This mini-review, mainly based on our resonance Raman studies on the structural origin of cooperative O binding in human adult hemoglobin (HbA), aims to answering why HbA is a tetramer consisting of two α and two β subunits. Here, we focus on the Fe-His bond, the sole coordination bond connecting heme to a globin. The Fe-His stretching frequencies reflect the O affinity and also the magnitude of strain imposed through globin by inter-subunit interactions, which is the origin of cooperativity.

Direct Quantification of Proteins Modified on a Polystyrene Microparticle Surface Based on ζ Potential Change.

The zeta potential (ζ) of a particle is a surface charge density (σ)-dependent parameter. If a change in σ can be induced by surface modification, the number of molecules modified on the particle can be detected as a measurable change in ζ. In this study, we demonstrate protein detection at zmol or pM levels (bovine serum albumin (BSA), myoglobin (Mb), and lysozyme (Lyz)) on carboxy-functionalized polystyrene (PS) microparticles using the ζ change.

Zeptomole detection of DNA based on microparticle dissociation from a glass plate in a combined acoustic-gravitational field.

In this study, we propose a novel detection principle based on the dissociation of microparticles immobilized on a glass plate through weak hybridization involving 4-6 base pairs (bps) in a combined acoustic-gravitational field. Particle dissociation from the glass plate occurs when the resultant of the acoustic radiation force (F) and the sedimentation force (F) exerted on the particle exceeds the binding force owing to the weak hybridization (F). Because F and F can be controlled by the microparticle density, and F is a function of the applied voltage to the transducer (V), an increase in V induces particle dissociation.

Kinetic Analysis of the Mass Transfer of Zinc Myoglobin in a Single Mesoporous Silica Particle by Confocal Fluorescence Microspectroscopy.

The adsorption/desorption mechanisms of biomolecules in porous materials have attracted significant attention because of their applications in many fields, including environmental, medical, and industrial sciences. Here, we employ confocal fluorescence microspectroscopy to reveal the diffusion behavior of zinc myoglobin (ZnMb, 4.4 nm × 4.

Effect of Molecular Crowding on Complexation of Metal Ions and 8-Quinolinol-5-Sulfonic Acid.

The effect of molecular crowding on macromolecular reactions has been revealed by many researchers. In this study, we investigate the complexation of metal ions (Zn, Co, and Cd) with 8-quinolinol-5-sulfonic acid as a model of small-molecular reactions in molecular crowding. The complexation constants for 1:1, 1:2, and total complexation in the presence of polyethylene glycol (PEG, a molecular crowding reagent) are evaluated based on the increase in the reactant activity by volume exclusion and the decrease in the water activity due to the change in osmotic pressure.

Roles of Fe-Histidine bonds in stability of hemoglobin: Recognition of protein flexibility by Q Sepharose.

Using various mutants, we investigated to date the roles of the Fe-histidine (F8) bonds in cooperative O binding of human hemoglobin (Hb) and differences in roles between α- and β-subunits in the αβ tetramer. An Hb variant with a mutation in the heme cavity exhibited an unexpected feature. When the β mutant rHb (βH92G), in which the proximal histidine (His F8) of the β-subunit is replaced by glycine (Gly), was subjected to ion-exchange chromatography (Q Sepharose column) and eluted with an NaCl concentration gradient in the presence of imidazole, yielded two large peaks, whereas the corresponding α-mutant, rHb (αH87G), gave a single peak similar to Hb A.

A role of heme side-chains of human hemoglobin in its function revealed by circular dichroism and resonance Raman spectroscopy.

Structural changes of heme side-chains of human adult hemoglobin (Hb A) upon ligand (O or CO) dissociation have been studied by circular dichroism (CD) and resonance Raman (RR) spectroscopies. We point out the occurrence of appreciable deformation of heme side-chains like vinyl and propionate groups prior to the out-of-plane displacement of heme iron. Referring to the recent fine resolved crystal structure of Hb A, the deformations of heme side-chains take place only in the β subunits.

Heterogeneity between Two α Subunits of αβ Human Hemoglobin and O Binding Properties: Raman, H Nuclear Magnetic Resonance, and Terahertz Spectra.

Following a previous detailed investigation of the β subunit of αβ human adult hemoglobin (Hb A), this study focuses on the α subunit by using three natural valency hybrid α(Fe-deoxy/O)β(Fe) hemoglobin M (Hb M) in which O cannot bind to the β subunit: Hb M Hyde Park (β92His → Tyr), Hb M Saskatoon (β63His → Tyr), and Hb M Milwaukee (β67Val → Glu). In contrast with the β subunit that exhibited a clear correlation between O affinity and Fe-His stretching frequencies, the Fe-His stretching mode of the α subunit gave two Raman bands only in the T quaternary structure. This means the presence of two tertiary structures in α subunits of the αβ tetramer with T structure, and the two structures seemed to be nondynamical as judged from terahertz absorption spectra in the 5-30 cm region of Hb M Milwaukee, α(Fe-deoxy)β(Fe).

Interrelationship among Fe-His Bond Strengths, Oxygen Affinities, and Intersubunit Hydrogen Bonding Changes upon Ligand Binding in the β Subunit of Human Hemoglobin: The Alkaline Bohr Effect.

Regulation of the oxygen affinity of human adult hemoglobin (Hb A) at high pH, known as the alkaline Bohr effect, is essential for its physiological function. In this study, structural mechanisms of the alkaline Bohr effect and pH-dependent O affinity changes were investigated via H nuclear magnetic resonance and visible and UV resonance Raman spectra of mutant Hbs, Hb M Iwate (αH87Y) and Hb M Boston (αH58Y). It was found that even though the binding of O to the α subunits is forbidden in the mutant Hbs, the O affinity was higher at alkaline pH than at neutral pH, and concomitantly, the Fe-His stretching frequency of the β subunits was shifted to higher values.

Heme Orientation of Cavity Mutant Hemoglobins (His F8 → Gly) in Either α or β Subunits: Circular Dichroism, (1) H NMR, and Resonance Raman Studies.

Native human adult hemoglobin (Hb A) has mostly normal orientation of heme, whereas recombinant Hb A (rHb A) expressed in E. coli contains both normal and reversed orientations of heme. Hb A with the normal heme exhibits positive circular dichroism (CD) bands at both the Soret and 260-nm regions, while rHb A with the reversed heme shows a negative Soret and decreased 260-nm CD bands.

Photoinduced Bilayer-to-Nonbilayer Phase Transition of POPE by Photoisomerization of Added Stilbene Molecules.

The photocontrol of a bilayer-to-nonbilayer phase transition (the liquid-crystalline Lα phase to the inverted hexagonal HII phase) of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE) by the photoisomerization of incorporated stilbene molecules was examined by utilizing differential scanning calorimetry, small-angle X-ray diffraction, ultraviolet (UV)/visible absorption, and attenuated total reflectance Fourier transform infrared spectroscopies. cis-Stilbene lowered the transition temperature, Th, to a greater extent than did trans-stilbene, and the difference was at most ca. 10 °C.