Agglutinin Binding Research Articles

A Two-Hybrid Assay to Study Protein Interactions within the Secretory Pathway

Interactions of transcriptional activators are difficult to study using transcription-based two-hybrid assays due to potent activation resulting in false positives. Here we report the development of the Golgi two-hybrid (G2H), a method that interrogates protein interactions within the Golgi, where transcriptional activators can be assayed with negligible background. The G2H relies on cell surface glycosylation to report extracellularly on protein-protein interactions occurring within the secretory pathway. In the G2H, protein pairs are fused to modular domains of the reporter glycosyltransferase, Och1p, and proper cell wall formation due to Och1p activity is observed only when a pair of proteins interacts. Cells containing interacting protein pairs are identified by selectable phenotypes associated with Och1p activity and proper cell wall formation: cells that have interacting proteins grow under selective conditions and display weak wheat germ agglutinin (WGA) binding by flow cytometry, whereas cells that lack interacting proteins display stunted growth and strong WGA binding. Using this assay, we detected the interaction between transcription factor MyoD and its binding partner Id2. Interfering mutations along the MyoD:Id2 interaction interface ablated signal in the G2H assay. Furthermore, we used the G2H to detect interactions of the activation domain of Gal4p with a variety of binding partners. Finally, selective conditions were used to enrich for cells encoding interacting partners. The G2H detects protein-protein interactions that cannot be identified via traditional two-hybrid methods and should be broadly useful for probing previously inaccessible subsets of the interactome, including transcriptional activators and proteins that traffic through the secretory pathway.

Immobilization of carbohydrate epitopes for surface plasmon resonance using the Staudinger ligation

The detection of carbohydrate–protein interactions is often performed using techniques that require surface immobilization of the lectin or the glycan. A commonly used assay for lectin binding is surface plasmon resonance (SPR). We describe an implementation of the Staudinger ligation as a method to immobilize carbohydrate epitopes to a biosensor surface. This was accomplished by first introducing an azide functionality to a carboxymethyldextran surface, followed by reaction with a phosphane-modified carbohydrate ligand. The chemistry employed is extremely mild and was easily adapted to a commercial biosensor system. Using this approach, we investigated the binding of jacalin and wheat germ agglutinin (WGA) to galactose, lactose, and N-acetyl-lactosamine. We observed that WGA binding shows evidence of multivalent interaction with the surface. Additionally, we found that jacalin binding was influenced by the presence of a flexible and hydrophobic galactosyl aglycone.

Characterization of the Pathogenesis Mechanism after Pseudomonas aeruginosa Infection through Food Consumption Using Chick Embryo Model

This study introduced a chick embryos` infection model to elucidate the pathogenesis mechanism of Pseudomonas aeruginosa, which causes serious diseases in human after ingestion of P. aeruginosa-contaminated animal originated foods. The embryonic chick model is able to give a rapid and relatively inexpensive method to assess bacterial pathogenicity compared to embryos of other vertebrates. Embryos were infected with P. aeruginosa and elastase-deficient P. aeruginosa. After infection with P. aeruginosa cells, total bacterial cell numbers and gelatinase activities in the embryos were compared. Thereafter, precartilage condensation and chondrogenesis were assessed by peanut agglutinin (PNA) binding on day 3 and by Alcian blue staining for sulfated proteoglycans on day 5, respectively. P. aeruginosa significantly increased in embryos, resulting in abnormal limb development, whereas P. aeruginosa defective in elastase activity partly impaired proliferation. In addition, P. aeruginosa-infected chick embryos significantly stimulated the production of matrix metalloproteinases. Several analyses showed that elevated proteases suppressed the proliferation and survival of chondrogenic cells. The results show that this infection model was a useful assay to determine the virulence mechanism of P. aeruginosa in human after intake of microbiologically contaminated foods.

MicroRNA-221 Regulates Chondrogenic Differentiation through Promoting Proteosomal Degradation of Slug by Targeting Mdm2

MicroRNAs (miRNAs) are small RNAs that fulfill diverse functions by negatively regulating gene expression. Here, we investigated the involvement of miRNAs in the chondrogenic differentiation of chick limb mesenchymal cells and found that the expression of miR-221 increased upon chondrogenic inhibition. Blockade of miR-221 via peanut agglutinin-based antisense oligonucleotides reversed the chondro-inhibitory actions of a JNK inhibitor on the proliferation and migration of chondrogenic progenitors as well as the formation of precartilage condensations. We determined that mdm2 is a relevant target of miR-221 during chondrogenesis. miR-221 was necessary and sufficient to down-regulate Mdm2 expression, and this down-modulation of Mdm2 by miR-221 prevented the degradation of (and consequently up-regulated) the Slug protein, which negatively regulates the proliferation of chondroprogenitors. These results indicate that miR-221 contributes to the regulation of cell proliferation by negatively regulating Mdm2 and thereby inhibiting Slug degradation during the chondrogenesis of chick limb mesenchymal cells.

Trypanosoma cruzi Subverts Host Cell Sialylation and May Compromise Antigen-specific CD8+ T Cell Responses

Upon activation, cytotoxic CD8(+) T lymphocytes are desialylated exposing beta-galactose residues in a physiological change that enhances their effector activity and that can be monitored on the basis of increased binding of the lectin peanut agglutinin. Herein, we investigated the impact of sialylation mediated by trans-sialidase, a specific and unique Trypanosoma transglycosylase for sialic acid, on CD8(+) T cell response of mice infected with T. cruzi. Our data demonstrate that T. cruzi uses its trans-sialidase enzyme to resialylate the CD8(+) T cell surface, thereby dampening antigen-specific CD8(+) T cell response that might favor its own persistence in the mammalian host. Binding of the monoclonal antibody S7, which recognizes sialic acid-containing epitopes on the 115-kDa isoform of CD43, was augmented on CD8(+) T cells from ST3Gal-I-deficient infected mice, indicating that CD43 is one sialic acid acceptor for trans-sialidase activity on the CD8(+) T cell surface. The cytotoxic activity of antigen-experienced CD8(+) T cells against the immunodominant trans-sialidase synthetic peptide IYNVGQVSI was decreased following active trans-sialidase-mediated resialylation in vitro and in vivo. Inhibition of the parasite's native trans-sialidase activity during infection strongly decreased CD8(+) T cell sialylation, reverting it to the glycosylation status expected in the absence of parasite manipulation increasing mouse survival. Taken together, these results demonstrate, for the first time, that T. cruzi subverts sialylation to attenuate CD8(+) T cell interactions with peptide-major histocompatibility complex class I complexes. CD8(+) T cell resialylation may represent a sophisticated strategy to ensure lifetime host parasitism.

Morphology, Ultrastructure, and Small Subunit rDNA Phylogeny of the Marine Heterotrophic Flagellate Goniomonas aff. amphinema

Marine goniomonads have a worldwide distribution but ultrastructural information has not been available so far. An isolate of the heterotrophic marine nanoflagellate Goniomonas (G. aff. amphinema) from North Wales (UK) has been studied, providing information on its morphology and cellular structure using video, electron, laser scanning confocal microscopy (LSCM), and atomic force microscopy. Here, we describe a new feature, a granular area, potentially involved in particle capture and feeding. The binding of the lectin wheat germ agglutinin to the granular area of cells with discharged ejectisomes indicates the adhesive nature of this novel feature. The presence of a microtubular intracellular cytopharynx, apparently also used for feeding, has been revealed by LSCM. The small subunit rRNA gene of the isolate has been sequenced (1,788 bp). Phylogenetic results corroborate significant genetic divergence within the marine members of Goniomonas. This work highlights the need for integrated morphological, ultrastructural, and molecular investigation when describing and studying heterotrophic nanoflagellates.

The effects of soybean agglutinin binding on the corneal endothelium and the re-establishment of an intact monolayer following injury – A short review

This short review summarizes the localization and effects of the plant lectin soybean agglutinin (SBA) on the injured and non-injured organ-cultured rat corneal endothelium. Although the tissue exists as a non-cycling monolayer on the posterior corneal surface a circular freeze injury promotes wound repair as cells initiate DNA synthesis, mitosis and migration. As a result, by 24 h post-injury, endothelial cells express a surface protein that binds SBA in a diffuse punctate pattern, which by 48 h after injury, becomes confined to the cell periphery. As healing proceeds, SBA binding dramatically declines, such that, only scattered binding is observed by 72 h after wounding. In non-injured organ-cultured endothelia, weak SBA binding appears 24 h after explanation but becomes prominently detected around the cell periphery by 48 h. Incubating injured or non-injured endothelia in SBA leads to alterations in their cellular appearance due to the fact that lectin exposure results in the disruption of the actin cytoskeleton. Although this does not affect migration, treatment with either SBA or N-acetylgalactosamine (the SBA binding sugar) does interfere with the reestablishment of cell–cell contact. It is postulated that the surface protein that binds SBA is expressed during conditions that are stressful to the tissue. During organ-culture the protein's appearance suggests a cellular response to explantation in order to enhance or maintain monolayer integrity. In wound repair its appearance may serve to establish preliminary cell–cell contact during the restoration of the endothelial monolayer.

Dual chemoarchitectonic lamination of the visual sector of the thalamic reticular nucleus

The chemoanatomical organization of the visual sector of the cat's thalamic reticular nucleus (TRN)—that is at the dorsal lateral geniculate nucleus (dLGN) and at the pulvinar nucleus (Pul)—was investigated with two novel cytoarchitectonic markers. The Wisteria floribunda agglutinin (WFA) binding reaction visualized the extracellular perineuronal net (PN) and the SMI 32 immunoreaction stained intracellular neurofilaments. Two distinct layers of the TRN could be detected, particularly by WFA- but also by SMI 32-staining. The outer tier outlined a canopy of labeling placed a bit detached from the diencephalon dorsolaterally, while the inner TRN tier is very tightly attached to the thalamic lamina limitans externa. The labeled neurons showed typically fusiform morphology with dendrites orienting in the plane of TRN. Additionally, these chemoarchitectural reactions identified a chain of structures in the ventral diencephalon connected to the TRN tiers. One stained string is formed by the subthalamic nucleus bound laterally to the peripeduncular nucleus extending further dorsolateral into the outer TRN tier. The other chain laced up the field of Forel, the zona incerta, the ventral LGN, the perigeniculate nucleus (PGN) and the previously-overlooked peripulvinar nucleus (PPulN) and so formed the inner TRN tier. In the third most distanced TRN tier, in the perireticular nucleus, a very few WFA-binding presenting neuron were found. In addition to the PN possessing TRN neurons, WFA-reactive presumable interneurons were also labeled within the visual thalamus. Following tracer injections into the feline Pul, two stripes of cells were retrogradely labeled in the neighboring visual TRN sector. The location of these reticular neurons coincided precisely with the chemoanatomically identified inner and outer TRN tiers. On the analogy of the PGN-TRN duality at the dLGN, the chemoanatomical and tract tracing findings strongly suggest a similar dual organization in the pulvinoprojecting TRN portion.

Measurement of wheat germ agglutinin binding with a fluorescence microscope

Signal intensity in fluorescence microscopy is often measured relative to arbitrary standards. We propose a calibration method based on a solution of the same fluorophore, whose binding to cells needs to be quantified. The method utilizes the low sensitivity of intensity to the object distance in wide-field imaging of uniform materials. Liquid layers of slowly varying depth were prepared by immersing a spherical lens into a drop of a fluorophore placed on a slide. Flatfield-corrected images of the contact and surrounding areas showed linear dependence of the gray level on the depth of fluorescent liquid. This allowed conversion of the measured intensity into the number of molecules per unit area. The method was applied to different cell types stained by WGA-Alexa 488 and WGA-TRITC. Consistent results were obtained by comparing microscopy with flow cytometry, comparing imaging through different objectives and comparing different WGA conjugates. Reproducibility of calibration was within 97% when low magnification was used. Fluorescence of free and bound WGA was found to be different, however, and therefore precise measurement of the number of cell-bound molecules was problematic in this particular system. We conclude that the method achieves reliable measurement of cellular staining in the units of soluble fluorophore. For probes whose fluorescent properties are unaffected by binding, quantification of staining in true molecular units should be possible.

Electrochemical monitoring of binding between wheat germ agglutinin and cellohexose-modified magnetic microbeads

An electrochemical method that uses glucose labeled with an electroactive compound was developed to evaluate the binding of wheat germ agglutinin (WGA) to cellohexose-modified magnetic microbeads. Cellohexose was attached to amino groups on the magnetic bead surface via formation of a Schiff's base. The labeled glucose acts as an electrochemical probe to monitor binding events between WGA and the cellohexose-modified beads. For a known quantity of cellohexose-modified beads, binding of WGA with cellohexose-modified beads was evaluated based on changes in electrochemical response of the labeled glucose. In particular, the peak current decreased as the concentration of WGA increased. Furthermore, the binding affinities of WGA for beads modified with four different cello-oligosaccharides were systematically compared using a voltammetric method.

Acrosome content release in streptolysin O permeabilized mouse spermatozoa.

Sperm cell plasma membrane and the outer acrosomal membrane fuse profusely during the acrosome reaction. The process is triggered by extracellular signals that elicit several intracellular events leading ultimately to membrane fusion. We have developed a streptolysin O permeabilizing protocol that selectively affects the spermatozoon plasma membrane without causing a significant loss of the acrosomal content. Most of the acrosomal acid phosphatase remained sperm-associated even after a 20 min incubation at 37 degrees C. However, the presence of 100 microM Ca2+ in the incubation buffer stimulates the release of the enzyme. The reaction was followed biochemically, measuring the acid phosphatase activity released to the medium and morphologically by the binding of fluorescein isothiocynate-conjugated peanut agglutinin and by electron microscopy. The results show that the streptolysin O permeabilized spermatozoon is a promising model for studying the complex set of events mediating and regulating the acrosome reaction.

Changing P2X receptor localization on maturing sperm in the epididymides of mice, hamsters, rats, and humans: a preliminary study

To study using immunohistochemistry the localization of P2X receptor subtypes on the head of immature sperm in the human, mouse, hamster, and rat caput epididymidis. Basic research. University-based hospital. Three human epididymides were obtained from patients undergoing orchidectomy for metastatic prostate cancer. P2X(1), P2X(2), P2X(3), and P2X(4) receptor immunolocalization on sperm. In the present study, P2X(1,2, and 3) receptor localization was immunohistochemically demonstrated on the head of immature sperm in the human, mouse, hamster, and rat caput epididymidis. P2X(4) receptor immunostaining was also observed on the head of sperm in the caput epididymidis of mice, hamsters, and humans, but not rats. There was a subsequent loss of receptor staining on sperm in the cauda epididymidis, except in humans where staining of P2X(4) receptors persisted. Comparision with peanut agglutinin (PNA) binding studies suggested the P2X receptors were located on the acrosome membrane. P2X(5-7) receptors were examined but found to be absent. The change in localization of receptor subtypes is coincidental with the functionally essential morphologic and maturational changes seen in sperm as they travel through the epididymis, and is suggestive of a role for purinergic signaling in sperm maturation and possibly fertility.

Characterization of the wheat germ agglutinin binding to self-assembled monolayers of neoglycoconjugates by AFM and SPR

Carbohydrate-protein interactions govern many crucial life processes involved in cell recognition events, but are often difficult to study because the interactions are weak, and multivalent exposure appears to be crucial for their biological function. We have used self-assembled monolayers (SAMs) of neoglycoconjugates as a model system to probe the specific interactions between the lectin wheat germ agglutinin (WGA) and monosaccharides by surface plasmon resonance (SPR) and atomic force microscopy (AFM) force measurements. SAMs presenting N-acetyl-D-glucosamine (GlcNAc) as a neoglycoconjugate were produced on gold surfaces, where the SAM formation was monitored using a quartz crystal microbalance (QCM) and shown to be a very rapid process. In the AFM force measurements WGA was covalently coupled to flexible polyethylene glycol (PEG) molecules at a probe surface using amine coupling. GlcNAc-specific binding events were detected with a WGA-modified probe on the GlcNAc-neoglycoconjugate SAM at bond rupture forces of 47 +/- 15 pN. Additionally, less frequent GlcNAc-specific unbinding events were detected at higher forces (120 +/- 20 pN) which are believed to originate from simultaneous detachment of multiple binding sites from the SAM surface. SPR measurements confirmed that WGA has higher affinity toward the immobilized GlcNAc-SAM than toward the soluble free monosaccharide. The binding constants obtained for soluble chitinoligosaccharides suggested up to three subsites within one carbohydrate-binding site of the WGA molecule and also provided further evidence of the multivalent binding character of the WGA dimer.

Case report: Diffuse splenic metastasis of occult breast cancer with incompatible blood group antigenic determinants

Cancer cells with immunogenic properties having altered protein glycosilation, modified blood group substances have been widely studied [Kannagi R, Miyake M, Zenita KM, Itai S, Hiraiwa N, Shigeta K, et al. Cancer-associated carbohydrate antigens: modified blood group substances and oncodevelopmental antigens on tumor cells. Gann Monogr Cancer Res 1988; 34: p. 15-28; Hakomori S. Antigen structure and genetic basis of histo-blood groups A, B and O their changes associated with human cancer. Biochem Biophys Acta 1999; 1473: p. 247-266; Brooks SA, Carter TM, Royle L, Harvey DJ, Fry SA, Kinch C, et al. Altered glycosilation of proteins in cancer: what is the potential for new anti-tumour strategies. Anticancer Agents Med Chem 2008; 8: p. 2-21]. In the study reported here, a 78-year-old female patient was admitted to the hospital with circulatory failure. At autopsy, the spleen (weight: 420 g) was extremely firm with a diffusely blackberry-colored cut surface. There were no signs of carcinomatous process at autopsy. By histology, the spleen showed diffuse metastatic carcinomatous infiltration. Using immunohistochemistry, an antibody to breast carcinoma antigen (BioGenex) labelled metastatic cells of the spleen and bone marrow. The patient was blood group O. Labelling for binding of lectins with and without blood group antigen specificity and monoclonal antibodies was carried out. The B blood group specific Banderiaea simplicifolia agglutinin I and an anti-B blood group monoclonal antibody labelled all the metastatic cells of spleen and bone marrow intensely. There was no detection of blood group A antigen by either binding of Dolichos biflorus agglutinin or anti-blood group A monoclonal antibodies. These observations raise the possibility that the detected incompatible B blood group antigen determinants on the metastatic cells were immunogenic. The surviving carcinoma cells may have found a place of refuge from immune surveillance in the spleen and in the bone marrow, where the complement-mediated tumor cell lysis immune rejection was not effective.

Soybean agglutinin binding to corneal endothelial cell surfaces disrupts in situ monolayer integrity and actin organization and interferes with wound repair

Rat corneal endothelium demonstrates cell-surface soybean agglutinin (SBA) binding during organ-culture or injury. When organ-cultured in medium containing SBA, the endothelial monolayer is disrupted because of cell-cell and cell-matrix alterations. SBA binding disorganizes the circumferential microfilament bundles (CMBs), an effect that is partially prevented by phallacidin preincubation. This disruption is reversible if tissues are returned to standard culture medium. Serum heightens SBA binding, whereas puromycin prevents it. Neither actinomycin D nor alpha-amanitin inhibits SBA binding, suggesting that SBA-binding protein(s) may be post-transcriptionally regulated. During injury-induced cell migration in the presence of SBA, cellular processes are blunted and fail to extend significantly outward. By 72 h post-injury, cells of SBA-treated tissues repopulate the wound but demonstrate little association with neighboring cells. Cells migrating in the presence of N-acetylgalactosamine appear normal but also fail to reassociate with other cells in the jury zone. Immunofluorescent staining for ZO-1 reveals punctuate patterns in cells of control tissues, whereas neither SBA- nor N-acetylgalactosamine-treated tissues exhibit ZO-1 staining. Terminal N-acetylgalactosamine removal fails to affect cell morphology, actin organization, or migration but prevents lectin binding. Our results suggest that SBA binding reflects the synthesis of a stress-induced protein(s) that may play a role in reestablishing cell-cell relationships during monolayer reorganization following injury.

Peritoneal ectopic lesions from women with endometriosis show abnormalities in progesterone-dependent glycan expression

Examination of 12 paired peritoneal ectopic and eutopic endometria for histochemical binding of Dolichos biflorus agglutinin, normally found in the mid-late secretory part of the cycle, showed a failure of lectin binding in 9 of 11 secretory-phase lesions although the eutopic specimens generally stained normally. This failure of glycan expression in the secretory phase may result from various anomalies, including an inability to respond to progesterone, possibly due to a lack of, or to nonfunctional, progesterone receptors, suggesting that an ectopic environment may produce changes in tissue cell biology and hormonal responsiveness compared with that of eutopic endometrium.

Evaluation of a new method for the diagnosis of alterations of Lens culinaris agglutinin binding of thyroglobulin molecules in thyroid carcinoma

The measurement of serum thyroglobulin (Tg) is widely used as a marker for recurrence of thyroid carcinoma following total thyroidectomy. However, this method cannot differentiate between benign and malignant disease. We focused on the sugar chain in the Tg molecule and investigated the usefulness of Lens culinaris agglutinin (LCA)-reactive Tg ratios in sera and wash fluids obtained during fine-needle aspiration (FNA) for the detection of thyroid carcinoma. The study was performed using 203 serum samples (115 from patients with benign thyroid disease and 88 from patients with thyroid carcinomas) and 176 wash fluid samples (143 benign, 21 malignant, and 12 inconclusive). LCA-reactive Tg ratios were determined using an enzyme-linked immunosorbent assay, and a comparison was made between malignant and benign lesions. In serum, the ratio in patients with malignancy was 79.5+/-6.0 [mean+/-standard deviation (SD)], significantly lower than in patients with benign lesions (84.9+/-3.5). The ratios in wash fluid from malignant lesions (75.8+/-18.9) were also significantly lower than those from benign lesions (85.6+/-3.9). These results suggest that this method could distinguish between benign and malignant lesions and may be useful for screening serum and wash samples.

Porous red cell ultrastructure and loss of membrane asymmetry in a novel case of hemolytic anemia

Transmission electron microscopic study revealed large pores on the erythrocyte ghost membranes, disrupted cytoskeleton and microcytosis of circulating erythrocytes in a novel case of hemolytic anemia. Greater loss of phosphatidylserine (PS) asymmetry was observed in younger erythrocytes compared with the aged ones in contrast to the normal red cells. Levels of sialylated glycoconjugates, such as glycophorin, measured by the binding of wheat germ agglutinin, showed greater loss upon aging. Such drastic loss of PS asymmetry leads to faster eryptosis, mediated by shedding of glycophorin-containing microvesicles leaving highly PS-exposed erythrocytes accessible to the phagocytes.

Significance of glycosylation patterns of rat Zajdela ascitic hepatocellular carcinoma: Effect of enzymatic removal of surface sialic acid on humoral response

Rat Zajdela ascitic hepatocellular carcinoma (ZAH) is a malignant cell type with some properties in common with rat hepatocytes. The aim of this study was to determine the glycosylation patterns of surface glycoproteins of two cell lines C and D of ZAH by lectin binding for identification and characterization of tumor-specific markers. Plasma membrane proteins from these cells were separated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate, and probed using radio-iodinated lectins. We observed a decrease in the binding of concavalin A, and an increase in the binding of wheat germ agglutinin (WGA) to glycoproteins from the tumor cell lines as compared with those from normal liver cells. We showed that the increase in binding of WGA was mainly due to increased sialylation of the surface glycoproteins of the tumor cells. The major sialylated glycoproteins of the tumor cells contained O-linked carbohydrate chains. It was also shown that removal of surface sialic acid by neuraminidase significantly decreased the lethality of the tumor and led to increased survival of tumor-bearing animals. The decreased lethality of the tumor appears to be due to increased antigenicity of the desialylated tumor cells. Taken together, the presence of highly sialylated O-linked glycosylation of gp120 in ZAH tumor cells and its absence in normal liver cells is of significance with respect to the biological properties of this tumor.

Binding of Ricinus communis agglutinin to a galactose-carrying polymer brush on a colloidal gold monolayer

A polymer with many pendent galactose residues was prepared by atom-transfer radical polymerization (ATRP) of galactose-carrying vinyl monomer, 2-lactobionamidoethyl methacrylate (LAMA), with a disulfide-carrying ATRP initiator, 2-(2′-bromoisobutyroyl)ethyl disulfide (DT-Br). The galactose-carrying polymer obtained (DT-PLAMA) was accumulated as a polymer brush via Au–S bond on a colloidal gold monolayer deposited on a cover glass. For comparison, a disulfide which carried one galactose residue at both ends (2-lactobionamidoethyl disulfide, Cys-Lac) was accumulated as a self-assembled monolayer (SAM) on the colloidal gold monolayer, too. The association and dissociation processes of galactose residues on the colloidal gold with a lectin, Ricinus communis agglutinin (RCA 120), were observed by the increase and decrease in absorbance at 550 nm corresponding to localized surface plasmon resonance (LSPR) phenomena. The Cys-Lac SAM-carrying glass chip showed a strong non-specific adsorption of the lectin, whereas the DT-PLAMA brush-carrying one reversibly associated with the lectin, indicating reusability of the latter device. The apparent association constant of the lectin with the galactose residues in the DT-PLAMA brush was much larger than the association constant for free galactose, and the detection limit of RCA 120 by the glycopolymer brush-modified device was satisfactorily low. Furthermore, a microscopic observation clearly indicated that the DT-PLAMA brush could reversibly associate with a HepG2 cell having galactose receptors, though these processes could not be observed spectrophotometrically due to a gigantic size of the cell.