Objective:To study the glycosylation of different clusterin isoforms and the alterations of their intracellular localizations in esophageal squamous cell carcinoma(ESCC),so as to better understand the potential mechanisms of clusterin in tumorigenesis of ESCC.Methods: The glycosylation of clusterin proteins was studied after treated with PGNase-F by Western blot analysis in 23 ESCC specimens and their corresponding adjacent tissues.Immunohistochemical staining was used to visualize the intracellular localization of clusterin proteins.Results: Mature clusterin,a 40 000 glycosylated protein in SDS-PAGE gel,(usually) presents in the connective tissues of the lamina propria of esophageal epithelial mucosa,and is secreted by esophageal glands onto the esophageal mucosa and in plasm.The glycosylation of clusterin proteins was abnormal in the 23 ESCC specimens,with the premature clusterin and nuclear clusterin aggregated in the cells and the mature clusterin decreased or disappeared in the stromal layer of esophagus.Conclusion: The deglycosylation and localization alterations of clusterin in ESSC may be closely associated with the tumorigenesis of ESCC.

Heat shock protein (HSP) 105 is a 105x103 stress protein that belongs to the HSP-110 family. It is released by tissues in response to a wide variety of stresses including infection and ischaemia. Studies have shown that the molecule is involved as a biochemical mediator of heat induced apoptosis by binding to p53 at scrotal temperatures and dissociating from it at suprascrotal temperatures in testicular germ cells. Recent studies have shown that HSP-105 is over-expressed in various malignancies besides in normal testicular tissue. HSP-105 may be a candidate of testis antigen.

Mass spectrometry-based quantitative proteomics has become an indispensable tool for tumor marker discovery.Stable isotope labeling with amino acid in cell culture(SILAC),as a representative of in vivo metabolic labeling strategy,is a simple and relatively quantitative assay for comparative proteomics.Two kinds of cultured cells are grown in the medium containing either a light or heavy isotope labeled essential amino acids.After several cell doublings,the heavy amino acids are introduced into the nascent polypeptides in a sequence-specific fashion and the natural amino acid is completely replaced by its isotopically labeled analog.Accuracy quantification would be feasible by analyzing the peak intensities of every same peptide pairs labeled with light or heavy isotopes by the mass spectrometry.Compared with chemical labeling,SILAC requires much less amount of labeled proteins,and it is an efficient,straightforward,reproducible and accurate approach in cell-based systems.This method can not only investigate the dynamics of protein abundance,but also monitor the variation of posttranslational modifications and protein-protein interactions qualitatively and quantitatively.It is a powerful and important quantitative proteomics tool for tumor marker discovery.

Copy number variations (CNVs) refer as a DNA segment that is 1 kb or larger and is presented at a variable copy number in comparison with a reference genome. Classes of CNVs include insertions, deletions, duplications and their complex combinations. Because they widely distributed in the genome with some important characteristics, such as heritable, relative stable and heterogeneity, CNVs are considered as novel genomic polymorphism markers. And the alteration of gene dosage which resulted from CNVs could change phenotype, so a novel CNV genome-wide association analysis (CNV-GWAS) strategy appeared recently and began to used for identifying susceptible genes of complex diseases. It was approved that it could complement the tranditional genome-wide association studies based on single nucleotide polymorphisms. Therefore, genomic structure variances are favorable for revealing the molecular mechanisms and genetic foundation of complex diseases.

Nowadays, proteomics focuses on quantitative analysis rather than qualitative. In the field of quantitative proteomics, Isobaric tags for relative and absolute quantitation (iTRAQ) is one of the most widely used techniques. The advantage of iTRAQ is high throughput, high stability and free of the restriction of sample property. iTRAQ is suitable for almost all kinds of samples, and up to 8 samples can be analyzed simultaneously by commercially available kit. Along with the development of techniques, more and more mass spectrometry (MS) platforms are used in iTRAQ experiments and the accuracy of iTRAQ has been improved. iTRAQ has been applied to studies of microorganism, animal, plant, medical and protein post-translational modification. Here we review the recent progress in the development of iTRAQ and its applications in quantitative proteomics.
Animals ; Humans ; Mass Spectrometry ; Proteomics ; methods

Objective:To clone soluble human anti-digoxin antibodies from large phage antibody library and construct a vector that expresses diabody.Methods:Soluble ScFvs were prepared through infecting E coli.HB2151 with the selected phage antibodies and induced with IPTG.The diabody vector was modified by enzyme digestion and checked by SDS-electrophoretogram.Results:It was showed by ELISA that soluble ScFv had specific binding ability to digoxin.The vector to express a soluble diabody was obtained by genetic modification,which was shown by Western blot.Conclusion:Soluble human anti-digoxin antibodies were successfully obtained from phage antibodies.The vector is efficient in creating diabody.

A new method based on adherence of cellulolytic bacteria to insoluble cellulose for isolation and purification of thermophilic cellulolytic anaerobes was reported, in which Hungate anaerobic operating techniques were used to roll tubes with insoluble cellulose powder as substrate.

Objective To investigate the efficacy and safety of ultrasound-guided polyglycol in patients with scar pregnancy. Methods Thirty-six patients with scarring treated in Third People's Hospital of Hangzhou from November 2014 to November 2016 were selected as subjects. All patients underwent ultrasound-guided polyglycol administration. The results were analyzed and analyzed. The results were analyzed. Gonadotropin (β-HCG) returned to normal time and the occurrence of adverse symptoms. Results The average recovery time, treatment cost, average recovery time and adverse reaction rate of blood β-HCG in the study group were lower than those in the control group, the difference was statistically significant (P<0.05). Conclusion The effect of ultrasound on the injection of polyphenols in patients with scar pregnancy is safe, feasible and easy to be accepted by patients and their families. It has clinical significance.

Objective:To clone human anti-ricin antibodies from large phage antibody library.Methods:Panning for a large phage library against ricin toxin was conducted to select specific antibodies against ricin. The binding activities and specificities were tested by ELISA method. Soluble ScFvs were prepared through infecting E coli. HB2151 with the selected phage antibodies and induction with IPTG. Results:Forty positive clones were obtained after 5 rounds of panning, and 12 clones had specific binding ability to ricin toxin. DNA fingerprinting showed 7 different band patterns indicating 7 different positive clones. DNA sequencing showed that variable regions of these ScFvs belonged to different subgroups.Conclusion:Human anti-ricin antibodies were successfully obtained from large phage antibody library.

In proteomic research, to improve protein solubility of membrane proteins and nuclear proteins, buffers containing high concentration of detergent, such as 4% SDS, were widely used. However, high concentration of detergent might severely interfere with the downstream proteomic analysis, including protein quantitation and trypsin digestion. To improve the proteomic compatibility of buffers with high concentration of detergent, we used short gel method to pretreat buffers containing detergent. Protein samples were first separated by a short (2-2.5 mm) SDS-PAGE electrophoresis, and proteins were quantitated by comparing with bovine serum albumin standards via optical density analysis. The gel was then cut and peptides were recovered using in-gel digestion. The quantitative linearity range of this method was 1 to 8 μg. The quantitation was accurate and reproducible. After short gel analysis, recovered peptides generated high mass spectrometry signals. In conclusion, short gel method eliminated the interference of high concentration detergent in the proteomics analysis, and it was suitable for protein samples' pretreatment, and was worth to apply in proteomic research.
Detergents ; chemistry ; Electrophoresis, Polyacrylamide Gel ; methods ; Mass Spectrometry ; Membrane Proteins ; chemistry ; Nuclear Proteins ; chemistry ; Proteins ; chemistry ; Proteomics ; methods ; Trypsin