With the development of bio-technological drugs, drug immunogenicity evaluation has become key factor of clarifying safety and efficacy of these drugs. It has become the focus to establish a stable and reliable evaluation system. Due to the advantages such as continuous real-time monitoring, surface plasmon resonance (SPR) technology has been widely used in bio-technological drugs immunogenicity assessments. Our study applied this technology to detect anti-drug antibody (ADA) of a recombinant human anti-rabies monoclonal antibody NM57 in the sera of 48 volunteers admitted in phase I clinical trials. This method could satisfy the basic requirements of detection of ADA.
Antibodies, Anti-Idiotypic ; analysis ; blood ; immunology ; Antibodies, Monoclonal ; analysis ; blood ; immunology ; Antibodies, Viral ; analysis ; blood ; immunology ; Humans ; Rabies virus ; immunology ; Recombinant Proteins ; analysis ; blood ; immunology ; Surface Plasmon Resonance

A 43-year-old man with generalized lichen planus demonstrated serum antibodies against autologous lesional skin. Indirect immunofluorescence using serum and papular lesional skin revealed a lichen planus specific antigen found only in the granular layer. The specific tissue antigen was not detected in normal skin from this patient, in normal skin from patients with skin disorders other than lichen planus or in skin from normal control persons. When titers of the serum antibodies against lichen planus antigen were examined before and after a successful therapy a positive correlation of the titer could be found in this patient.
Adult ; Antibodies/*immunology ; Antigens/*analysis ; Humans ; Lichen Planus/drug therapy/*immunology ; Male ; Skin/*immunology

Antibodies, Antineutrophil Cytoplasmic ; analysis ; immunology ; Antibodies, Antinuclear ; analysis ; immunology ; Antigens, Nuclear ; immunology ; Arthritis, Juvenile ; immunology ; Autoantibodies ; analysis ; immunology ; Humans ; Monitoring, Physiologic ; methods ; Nucleosomes ; immunology ; Phospholipids ; immunology ; Rheumatic Diseases ; immunology ; physiopathology

We evaluated the clinical usefulness of simultaneous LISS/Coombs and NaCl/Enzyme testing using the gel method for screening and identification of unexpected antibodies in 15,014 samples. When unexpected antibodies were detected by either screening test, those antibodies were identified using both the LISS/Coombs and the NaCl/Enzyme gel test. The positive screening rates of the LISS/Coombs, NaCl/Enzyme, and combined tests (excluding 25 autoantibody cases) were 0.48%, 1.29%, and 1.39%, respectively. Among the 57 samples positive by both screening methods, the antibodies in 19.3% could be identified only by the NaCl/Enzyme method. Among the 137 samples positive only by NaCl/Enzyme screening, 74.5% showed positive results in antibody identification only by the NaCl/Enzyme test, although 7.3% were also positive in the LISS/Coombs test. The NaCl/Enzyme method thus showed about threefold higher detection rates than the LISS/Coombs method, especially in screening for Rh antibodies, and higher exact identification rates and discriminatory power for identifying mixed antibodies. Addition of the NaCl/Enzyme method to routine laboratory procedures may detect and identify considerable numbers of significant antibodies that might be missed if only the LISS/Coombs method is used.
Antibodies/*analysis/immunology ; *Coombs' Test ; Erythrocytes/*immunology ; Hemagglutination Tests/*methods ; Humans ; Isoantibodies/analysis ; Reagent Kits, Diagnostic

In order to optimize the fabrication of SiO2 tubes immobilized with antibody for hepatitis C virus antigen (HCAg) detection, we formed the activated amino on the surface of SiO2 tubes by using the activation of aminosilane. Then we immobilized the hepatitis C virus (HCV) monoclonal antibody on the surface of SiO2 tubes by using glutaraldehyde as a chemical cross-linker, followed by detecting HCAg. Sequence tests showed that when the SiO2 tubes were treated in 10% (V/V) aminosilane solution and 3% (V/V) glutaraldehyde solution for 3 hours and 2 hours, respectively, the HCV monoclonal antibody had high immobilization efficiency and low nonspecificity, and the HCAg was detected to 1 ng/mL. This experiment can provide principle and experimental data for establishment of HCAg magnetic immunoassay system.
Antibodies, Immobilized ; immunology ; Antibodies, Monoclonal ; chemistry ; immunology ; Hepatitis C Antibodies ; chemistry ; immunology ; Hepatitis C Antigens ; analysis ; immunology ; Humans ; Silicon Dioxide ; chemistry

BACKGROUNDSkin lesions are common manifestations in systemic lupus erythematosus (SLE). It is still unknown what the definite pathogenesis of skin involvement was and whether DNA participated in it. Our study was designed to explore the pathogenetic role and nature of nuclear antigen (DNA) deposited in the skin lesions of patients with SLE.

METHODSThirty skin samples from patients with SLE and 2 normal skin samples were studied. Extracellular DNA was evaluated by indirect immunofluorescence methods. The deposited immune complexes were extracted by cryoprecipitation, and DNA was then isolated with phenol and chloroform. DNA fragment sizes were detected by agarose gel electrophoresis. Finally, 8 different probes were used to analyze the origin of these DNA molecules using Dot hybridization.

RESULTSExtracellular DNA staining was found only in skin lesions, mainly those located in the basement membrane zone, vascular wall, and hair follicle wall. Normal skin and non-lesion SLE skin showed no fluorescence at locations outside the nuclei. There were no differences in the rate and intensity of extracellular DNA staining when comparing active phase to remission phase patients. No relationship was found between extracellular DNA and circulating anti-dsDNA antibodies. Deposited DNA fragments clustered into four bands of somewhat discrete sizes: 20 000 bp, 1300 bp, 800-900 bp, 100-200 bp. Small sized fragments (100-200 bp) were positively correlated with disease activity (P < 0.05, r = 0.407). Dot hybridization showed significant homology of the various extracellular DNA fragments examined with human genomic DNA, but not with DNA from the microorganisms and viruses we examined. There were also homologies between DNA samples from different individuals.

CONCLUSIONSDNA and its immune complexes may contribute to the pathogenesis of skin lesions in SLE. These DNA molecules range in size from 100 bp to 20 kb and may be endogenous in origin.

Antibodies, Antinuclear ; blood ; Antigen-Antibody Complex ; analysis ; DNA ; analysis ; immunology ; Humans ; Lupus Erythematosus, Systemic ; immunology ; Skin ; immunology ; Staining and Labeling

Antibodies ; Breast Neoplasms ; chemistry ; Female ; Humans ; Immunohistochemistry ; Neoplasm Proteins ; analysis ; Receptors, Estrogen ; analysis ; immunology ; Receptors, Progesterone ; analysis ; immunology

OBJECTIVETo prepare the antibodies against salbutamol (SAL) with high sensitivity and to develop an indirect competitive enzyme-linked immunoassay (ic-ELISA) for fast detection of SAL.

METHODSThe New Zealand white rabbits were immunized with SAL in a small dose and long period mode. The method of ic-ELISA was optimized and adopted for the detection of a series of SAL samples, then the standard curve of SAL was established. The precision and the recoveries of the method were determined.

RESULTSThe antibodies with high sensitivity towards SAL were prepared with a IC50 of 12.21 ng/ml. The ic-ELISA method for SAL measurement was established, the recoveries of measurement was between 95%-105% and the CV was <3%.

CONCLUSIONThe antibodies against salbutamol have been prepared and an indirect competitive enzyme-linked immunoassay for fast and specific detection of SAL has been developed.

Albuterol ; analysis ; immunology ; Animals ; Antibodies ; immunology ; Enzyme-Linked Immunosorbent Assay ; methods ; Male ; Rabbits

Heavy metal leftover on farm and stock products has become a big threat to human. It is necessary to develop some fast and efficient detection methods. Heavy metal immunoassays are new methods for detection of heavy metal ions. Compared to the traditional chemical methods, immunoassays are not only fast, cheap, simple, but also reasonably portable, highly sensitive and selective. It can be used as preliminary screening for rapid determination of heavy metal ions. Except chemical chelators, phytochelatin and metallothionein can also be used for preparing immunogen, both of them can chelate heavy metal ions to carrier protein. There are two prototype assays: polyclonal antibody immunoassay and monoclonal antibody immunoassay. The former includes fluorescence polarization immunoassay; the latter includes indirectly competitive ELISA, one-step competitive immunoassay and KinExA immunoassay. Among these assays, indirectly competitive ELISA which was used for determining heavy metal ions in the early days was easy to be interfered and showed false positive. Fluorescence polarization immunoassay which used polyclonal antibody for determining heavy metal ions was simple and cheap. KinExA instrument could be functioned as an immunosensor for environmental samples. One-step immunoassay which avoided to the addition of second antibody and chromogenic substrate was simple and sensitive. Colloidal gold enhanced immunochromatography assay is a semi-quantitation for determining heavy metal ions. As an adjunctive way for chemical methods, it has the potential application in rapid determination of heavy metal ions.
Animals ; Antibodies, Monoclonal ; immunology ; Gold ; chemistry ; Humans ; Immunoassay ; methods ; Metals, Heavy ; analysis ; immunology

Avidin layer was bound on the substrate surface of Silicon wafer modified with aldehyde. The interaction between avidin and biotin was adopted for the immobilization of mouse monoclonal biotin-anti-M13 (antibody GP3)-labeled biotin. The surface was incubated in a solution containing phage M13KO7, which was trapped by the antibody GP3 with the interaction between phage M13KO7 and antibody GP3, resulting in a variation of layer thickness that was detected by imaging ellipsometry. The results showed a saturated layer of antibody GP3 with a thickness about 6.9 nm on the surface of the silicon wafer. The specific interaction between phage M13KO7 and antibody GP3 resulted in a variation of the layer thickness. The layer of phage M13KO7 bound with antibody GP3 was 17.5 nm in the concentration of 1.1 x 10(10) pfu/mL. Each variation of layer thickness corresponded to a concentration of phage M13KO7 in the range of 0.1 x 10(10) approximately 2.5 x 10(10) pfu/mL, with the sensitivity of 10(9) pfu/mL. Compared with other methods, the optical protein-chip requires only short measurement time, is label free, is a quantitative test, and can be visualized. This study could be significant on the investigation of interactions between the antibody and virus, and shows potential in the early diagnosis of virosis.
Animals ; Antibodies, Viral ; immunology ; Bacteriophage M13 ; immunology ; isolation & purification ; Mice ; Protein Array Analysis ; methods