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A competitive enzyme-linked immunosorbent assay (ELISA) for ginkgolic acids (GAs) was developed using monoclonal antibody (MAb) 9F raised against 6-(13-formylheptyl) salicylic acid covalently coupled to bovine serum albumin (BSA). ELISA, at an effective measuring range of 300 ng/ml–1 μg/ml of GA_15:1, was successful in detecting GAs content in ginkgo leaves and standardized extracts due to the lack of cross-reactivity against various related compounds. The sensitive and simple immunoassay developed in this study was validated to be specific for the quantitative determination of total GAs content in ginkgo crude drugs with no interference from the sample matrix. The analytical recovery of spiked GA_15:1 was 103% in a concentration range between 10 and 40 mg/g dry weight of ginkgo leaves.

Enzyme-linked immunosorbent assay (ELISA) systems using anti-ginsenoside Rb1 (G-Rb1) and Rg1 (G-Rg1) monoclonal antibodies (MAbs) were established for pharmacokinetic investigations of G-Rb1 and G-Rg1 in rat serum. The systems not only allowed sensitive detection of G-Rb1 at the level as low as 20 ng/ml and of G-Rg1 at 300 ng/ml, but showed strong capacity for detecting the two agents in a broad concentration range (20 to 400 ng/ml for G-Rb1 and 0.3 to 10 μg/ml for G-Rg1, respectively). In this respect, these assay systems are superior to other methods using thin-layer chromatography (TLC) or high-performance liquid chromatography (HPLC). In addition, another advantage of these immunoassays is the comparably low quantities of specimen required; as little as 5 μl of serum suffices the need for determination of ginsenosides. We report in this article the application of this immunoassay in pharmacokinetic study of G-Rb1.

Aristolochic acid-II (AA-II) conjugated with bovine serum albumin (BSA) was used as an antigen for immunizing BALB/c mice. Isolated splenocytes from the immunized mice were fused with an aminopterin-sensitive mouse myeloma cell line, SP2/0-Ag14, to produce hybridoma cells that secreted a monoclonal antibody (MAb) against AA-II. The selected hybridoma was subsequently cloned by limited dilution method. For MAb, the isotype and an estimated dissociation constant (K_D) of the MAb were determined. The MAb was used to establish an ELISA method. Accuracy and variation assays, as well as determinations of the specificity and sensitivity, were also carried out and the linear range was 0.19–13 μg/ml. The anti-AA-II MAb showed a very high specificity for AA-II and had low cross-reactivities against the other aristolochic acid (AAs) (CR: AA-I, 3.4%; AA-VIIa, 0.86%) or aristololactam-I (AL-I) (CR < 0.07%) except AA-IIIa which has 17% of cross activity. Anti-AA-II MAb also showed negligible cross-reactivity (< 0.5%) toward other natural compounds with different chemical structures including barbaloin, sennoside A, rutin, glycyrrhizin, caffeic acid etc. This is the first time that an ELISA method was successfully established for the application of anti-AA-II MAb.

Aristolochic acids (AAs) are found in herbal medicines of Aristolochiaceae plants, including Aristolochia and Asarum species. AAs are associated with a rapidly progressive interstitial nephritis, which is called aristolochic acid nephropathy (AAN). However, the in-situ localization of AAs in the target organ, the kidney, has not been investigated yet. In the present study, the accumulation of aristolochic acid I (AA-I) in mouse kidney was revealed by immunoperoxidase light microscopy as well as colloidal gold immunoelectron microscopy (IEM) based on an anti-AA-I and AA-II monoclonal antibody (mAb). Male BALB/c mice were treated with 1.25 or 2.50 mg kg^-1 of AA-I per day for 5 days. Paraffin sections and ultra-thin sections of kidney tissue were respectively prepared. Under light microscopy, the apical surface of proximal tubules was strongly stained for AA-I, whereas no obvious immunostaining was found in the distal tubules and glomerulus, which remained relatively intact. Under electron microscopy, epithelial cells of the proximal tubules, distal tubules and collecting tubules were broken to various degrees. Gold labeling in the proximal and distal tubules was stronger than that in the collecting tubules. In renal tubules, immunogold signals of AA-I tended to accumulate in the mitochondria and peroxisomes, though the signals could be observed all over the cell. Gold signals were also found in the erythrocytes of glomeruli. The MAb against AA-I and AA-II provides a clue for the identification of proteins or factors which might interact with AA-I and thus induce targeted damage of kidney.

Aristolochic acid I (AA-I) is a strong nephrotoxin, carcinogen, and mutagen found in plants such as the Aristolochia species. The mechanisms underlying AA-I toxicity in the kidneys are poorly understood. In this study, we aimed to gain insight into the mechanism of AA-I nephrotoxicity by analyzing the uptake, subcellular distribution, and intracellular targets of AA-I in the human kidney cell line HK-2 using immunocytochemistry, immunoprecipitation, and LC-MS/MS. In HK-2 cells incubated with 20$\mu$g/mL AA-I for different periods of time (up to 12h), AA-I was detected by a specific monoclonal antibody (MAb) against AA-I, both in the cytoplasm and nuclei. Nuclear localization depended on the exposure time. A protein with the molecular weight of 100 kDa was immunoprecipitated with the anti-AA-I MAb from the AA-I-treated cell lysates and was identified by LC-MS/MS as $\alpha$-actinin-4 after digestion of the protein, and was confirmed by immunoblotting with a specific anti-$\alpha$-actinin-4 MAb. This evidence shows, for the first time, that $\alpha$-actinin-4 is a protein targeted by AA-I in kidney cells. Our findings strongly suggest an association between $\alpha$-actinin-4 and AA-I nephrotoxic activity.

Computing Functional Repertoire of Systems Scale Omics Data.

Monoclonal Antibody Therapeutics.

This article shortly reveals about the status of Taiwan's biotech and pharmaceutical industries.

GlaxoSmithKline and OncoMed Pharmaceuticals Form Strategic Alliance to Develop Cancer Stem Cell Antibody Therapeutics.

BioArctic Neuroscience and Eisai Enter Into Exclusive Licensing Agreement with Novel Antibody Treatment for Alzheimer's Disease.

Japan Bioventures Today — Japan Animal Referral Medical Center.

Agilent Technologies Collaborates with BioNanomatrix to Develop Genetic Analysis System.

Eisai Opens Regional Clinical Research Center in Singapore.

GE Healthcare and Novavax to Develop and Market Pandemic Infuenza Solution.

Nanostart AG Marks Entry into Asia as Lead Investor in Singapore's Biomed Start-Up Curiox Biosystems.

QIAGEN and Bio*One Capital Enter into Joint Venture to Develop Molecular Diagnostics Products.

Binex and MaxCyte Collaborate on Cancer Immunotherapy Research.

Prana Discovers New Drug Candidates for Parkinson's Research

Cytopia Announces Novel Anticancer Drug Development Collaboration

CSIRO Signs Agreement with Oncaidia to Develop Anticancer Drug

First Commercial Product Launch of Starpharma's DNT Priostar

China Sky One Develops SemiQuantitative Automatic Inspection Device for Diagnostic Kits

Ranbaxy Laboratories Signs Drug Discovery Development Pact with Merck

IAVI and CHAVI Team Up on AIDS Vaccine Development

DxTech and Nicholas Piramal signs Product Development and Licensing Agreement for Point-of-care Diagnostic Testing System

Sanofi-Aventis Launches SoloSTAR Insulin Pen in India

Strategic Alliance between Ranbaxy and Orchid

Piramal Diagnostics Introduces Asia's First High-definition PET/CT Scanner

Ocimum Biosolutions Launches Genowiz 4.0

BIOVET Launches Asia's First and World's Second BSL-4 Manufacturing Facility in India

Intas Plans Filgrastim launch in North America, Europe

Taisho and Chugai enter Co-Development and Co-Marketing Agreement for Activated Vitamin D Derivative, ED-71

Astellas and CoMentis Sign Agreement to Collaborate on the Research, Development and Commercialization of Beta-Secretase Inhibitors

New Hope for Burn Victims

Agendia and Agilent Technologies Announce Plans to Jointly Develop New Diagnostic Tests, Extend Supply Agreement

First CCD 2D Gel Imaging System Installed at Singapore's New SMART Centre will Help Accelerate Pace of Infectious Disease Research in Southeast Asia

High-Throughput Sequencing on a Next Generation Sequencer to Identify Specific Binders from a Phage Library

Antibody Solution Viscosity and Intermolecular Interactions: Considerations for Development of Highly Concentrated Formulations

Display of Membrane Proteins on a Viral Envelope for Antibody Generation

Sequence and Structural Determinants of Antigen Binding in Antibody CDR Loops

Enhancement of the Stability of Single Chain Fv Molecules with the Amino Acid Substitutions Predicted by High-Performance Computer

Thermal Stability of Camelid Single Domain VHH Antibody

Human epidermal growth factor receptor (EGFR) is strongly associated with malignant proliferation and has been established as an attractive therapeutic target of diverse cancers and used as a significant biomarker for tumor diagnosis. Over the past decades, a variety of monoclonal antibodies (mAbs) have been successfully developed to specifically recognize the third subdomain (TSD) of EGFR extracellular domain. Here, the complex crystal structures of EGFR TSD subdomain with its cognate mAbs were examined and compared systematically, revealing a consistent binding mode shared by these mAbs. The recognition site is located on the $\beta$-sheet surface of TSD ladder architecture, from which several hotspot residues that significantly confer both stability and specificity to the recognition were identified, responsible for about half of the total binding potency of mAbs to TSD subdomain. A number of linear peptide mimotopes were rationally designed to mimic these TSD hotspot residues in different orientations and/or in different head-to-tail manners by using an orthogonal threading-through-strand (OTTS) strategy, which, however, are intrinsically disordered in Free State and thus cannot be maintained in a native hotspot-like conformation. A chemical stapling strategy was employed to constrain the free peptides into a double-stranded conformation by introducing a disulfide bond across two strand arms of the peptide mimotopes. Both empirical scoring and fluorescence assay reached an agreement that the stapling can effectively improve the interaction potency of OTTS-designed peptide mimotopes to different mAbs, with binding affinity increase by $>10$-fold. Conformational analysis revealed that the stapled cyclic peptide mimotopes can spontaneously fold into a double-stranded conformation that well threads through all the hotspot residues on TSD $\beta$-sheet surface and exhibits a consistent binding mode with the TSD hotspot site to mAbs.

Background and Aims: Accurate endoscopic detection of premalignant lesions and early cancers in the colon is essential for cure, since prognosis is closely related to lesion size and stage. Although it has great clinical potential, autofluorescence endoscopy has limited tumor-to-normal tissue image contrast for detecting small preneoplastic lesions. We have developed a molecularly specific, near-infrared fluorescent monoclonal antibody (CC49) bioconjugate which targets tumor-associated glycoprotein 72 (TAG72), as a contrast agent to improve fluorescence-based endoscopy of colon cancer. Methods: The fluorescent anti-TAG72 conjugate was evaluated in vitro and in vivo in athymic nude mice bearing human colon adenocarcinoma (LS174T) subcutaneous tumors. Autofluorescence, a fluorescent but irrelevant antibody and the free fluorescent dye served as controls. Fluorescent agents were injected intravenously, and in vivo whole body fluorescence imaging was performed at various time points to determine pharmacokinetics, followed by ex vivo tissue analysis by confocal fluorescence microscopy and histology. Results: Fluorescence microscopy and histology confirmed specific LS174T cell membrane targeting of labeled CC49 in vitro and ex vivo. In vivo fluorescence imaging demonstrated significant tumor-to-normal tissue contrast enhancement with labeled-CC49 at three hours post injection, with maximum contrast after 48 h. Accumulation of tumor fluorescence demonstrated that modification of CC49 antibodies did not alter their specific tumor-localizing properties, and was antibody-dependent since controls did not produce detectable tumor fluorescence. Conclusions: These results show proof-of-principle that our near-infrared fluorescent-antibody probe targeting a tumor-associated mucin detects colonic tumors at the molecular level in real time, and offer a basis for future improvement of image contrast during clinical fluorescence endoscopy.

Since the percutaneous coronary intervention (PCI) was first introduced into China in 1984, this procedure has become widely accepted as an important step in coronary revascularization. This study aims to evaluate intimal hyperplasia and thrombosis after implantation of platelet glycoprotein IIIa monoclonal antibody (mAb)-eluting stent in the New Zealand white rabbit abdominal aorta or iliac artery by comparing CT angiography and pathological experiments. The antibody-eluting stents were prepared by the passive absorption method. Arterial intima in the stented segment and 0.5 cm adjacent to the stented site were observed and analyzed by scanning electron microscopy (SEM) and cross-sectional staining. Endothelialization and thrombosis on the stent surface were visualized by CT angiography and three-dimensional (3D) reconstruction technique in animals surviving from 1 to 12 weeks. Compared to stainless steel stents, the surface of antibody-eluting stents was covered with a complete endothelial layer after four weeks. Both CT angiography and pathological results showed intimal hyperplasia (p < 0.05) after 12 weeks. Therefore, pathological methods are the goldern standard for evaluation of intimal hyperplasia while CT angiography has a higher specificity in demonstrating the intimal changes after stent implantation for 12 weeks. The mAb-eluting stent has the potential to prevent thrombosis formation due to the interaction of stent with blood, and decreases the stenosis ratio by inhibiting neointima proliferation.

A prototype lateral flow device (LFD) serological test kit was evaluated at Central Science Laboratory (CSL), York, England for on-site detection of Clavibacter michiganensis subsp. sepedonicus(Cms), the causative agent of potato ring rot disease. Monoclonal antibody (MAb) was produced against Cms extra-cellular carbohydrate and then was tested for specificity and sensitivity of detection against a comprehensive panel of bacteria composing different isolates of Cms and close relatives from the National Collection of Plant Pathogenic Bacteria (NCPPB) and CSL research (Protect System) collection. When the Mab was used in immunofluorescence antibody staining (IFAS) tests, the detection limit was as few as 1 × 10³ bacterial cells ml^-1. However, in the LFD, the detection limit was recorded as 1 × 10⁴ bacterial cells ml^-1 in naturally infected and artificially spiked potato, tomato and aubergine plant extracts. All mucoid Cms strains were detected using the LFD but the non-mucoid strain of Cms (NCPPB 3898) was not detected. Specificity of the LFD was comparable to IFAS and Polymerase Chain Reaction laboratory-based tests in distinguishing between Cms and closely-related bacteria.