Locus-Specific Bisulfate NGS Sequencing of GSTP1, RNF219, and KIAA1539 Genes in the Total Pool of Cell-Free and Cell-Surface-Bound DNA in Prostate Cancer: A Novel Approach for Prostate Cancer Diagnostics.

The locus-specific methylation of three genes (GSTP1, RNF219, and KIAA1539, also known as FAM214B) in the total pool of blood cell-free DNA, including cell-free DNA from plasma and cell-surface-bound DNA, of patients with prostate cancer and healthy donors was studied on the MiSeq platform. Our study found a higher methylation index of loci for total cell-free DNA compared with cell-free DNA. For total cell-free DNA, the methylation of GSTP1 in each of the 11 positions provided a complete separation of cancer patients from healthy donors, whereas for cell-free DNA, there were no positions in the three genes allowing for such separation.

Locus-Specific Methylation of GSTP1, RNF219, and KIAA1539 Genes with Single Molecule Resolution in Cell-Free DNA from Healthy Donors and Prostate Tumor Patients: Application in Diagnostics.

The locus-specific methylation of three genes (GSTP1, RNF219, and KIAA1539 (also known as FAM214B)) in the blood plasma cell-free DNA (cfDNA) of 20 patients with prostate cancer (PCa), 18 healthy donors (HDs), and 17 patients with benign prostatic hyperplasia (BPH) was studied via the MiSeq platform. The methylation status of two CpGs within the same loci were used as the diagnostic feature for discriminating the patient groups. Many variables had good diagnostic characteristics, e.

Promising New Tools for Targeting p53 Mutant Cancers: Humoral and Cell-Based Immunotherapies.

Transcription factor and oncosuppressor protein p53 is considered as one of the most promising molecular targets that remains a high-hanging fruit in cancer therapy. gene encoding the p53 protein is known to be the most frequently mutated gene in human cancers. The loss of transcriptional functions caused by mutations in p53 protein leads to deactivation of intrinsic tumor suppressive responses associated with wild-type (WT) p53 and acquisition of new pro-oncogenic properties such as enhanced cell proliferation, metastasis and chemoresistance.

Fluorescent tagging the NS1 protein in yellow fever virus: Replication-capable viruses which produce the secretory GFP-NS1 fusion protein.

Yellow fever virus, the prototype in the genus Flavivirus, was used to develop viruses in which the nonstructural protein NS1 is genetically fused to GFP in the context of viruses capable of autonomous replication. The GFP-tagging of NS1 at the amino-terminus appeared possible despite the presence of a small and functionally important domain at the NS1's amino-terminus which can be distorted by such fusing. GFP-tagged NS1 viruses were rescued from DNA-launched molecular clones.

Towards development of plasmacytoma cells-based expression systems utilizing alphavirus vectors: An NS0-VEE model.

Plasmacytoma (myeloma) cells have a large protein expression capacity, although their industrial use is confined to stable expression systems. Vectors derived from genomes of viruses from the genus Alphavirus allow obtaining of high yields of target proteins but their use is limited to transient expression. Little information has been published to date on attempts to combine the myeloma cells as hosts with alphaviruses as expression vectors.

Cell-Free miRNA-141 and miRNA-205 as Prostate Cancer Biomarkers.

Expression levels of five miRNAs (miR-19b, miR-21, miR-126, miR-141, miR-205) were measured in the plasma of healthy donors and prostate cancer patients. It was shown that miR-141 expression level efficiently discriminates early stage prostate cancer patients and correlates with the Gleason score.

Comparative Study of Extracellular Vesicles from the Urine of Healthy Individuals and Prostate Cancer Patients.

Recent studies suggest that extracellular vesicles may be the key to timely diagnosis and monitoring of genito-urological malignancies. In this study we investigated the composition and content of extracellular vesicles found in the urine of healthy donors and prostate cancer patients. Urine of 14 PCa patients and 20 healthy volunteers was clarified by low-speed centrifugation and total extracellular vesicles fraction was obtain by high-speed centrifugation.

Updates on the Production of Therapeutic Antibodies Using Human Hybridoma Technique.

Therapeutic antibodies are implicated into the very promising and fast growing area of pharmaceutics. Human hybridoma technology, allowing generation of natural human antibodies in a native form, seems to be the most direct way that require no additional modifications for production of therapeutic antibodies. However, technical difficulties in human hybridoma creation discovered in the 80s of the last century have switched the mainstream therapeutic antibody development into new directions like display and transgenic mice techniques.

[A New Simple Technique for Producing Labeled Monoclonal Antibodies for Antibody Pair Screening in Sandwich-ELISA].

A simple and fast method for obtaining biotin-labeled monoclonal antibodies was developed usingcontent of hybridoma culture supernatant sufficient to select antibody pairs in sandwich ELISA. The method consists in chemical biotinylation of antigen-bound antibodies in a well of ELISA plate. Using as an example target Vaccinia virus A27L protein it was shown that the yield of biotinylated reactant is enough to set comprehensive sandwich ELISA for a moderate size panel of up to 25 monoclonal antibodies with an aim to determine candidate pairs.

[Immunological and functional characteristics of epitopes and regions of gB glycoprotein of Aujeszky's disease virus].

The role of different gB epitopes and regions at some stages of virus replication in cell cultures and in the formation of immunity to Aujeszky's disease virus (ADV) was studied using a panel of 13 monoclonal antibodies (MAB) that recognize glycoprotein gB (gB) of ADV and antisera against fusion recombinant proteins expressing gB fragments. Productive infection following virion attachment was prevented by antibodies to the N-terminal domain of gB. Three MABs against the N-terminal domain of gB and 5 MABs directed against the immunodominant region located in the gBc-subunit of gB inhibited the cell-to-cell spread of viral infection.

Distribution of B-cell epitopes on the pseudorabies virus glycoprotein B.

In order to map antigenically important regions of glycoprotein B (gB) of pseudorabies virus (PrV), a panel of recombinant fragments of gB expressed in E. coli and truncated fragments of gB generated by cleavage of purified native gB with trypsin and cyanogen bromide was analysed by using 26 monoclonal antibodies directed against gB. Three continuous epitopes were localized in the vicinity of the N terminus of gB, between amino acids (aa) 59 and 126.

Glycoprotein B of Aujeszky's disease virus: topographical epitope mapping and epitope-specific antibody response.

A panel of 26 monoclonal antibodies (mAbs) against glycoprotein B (gB) of Aujeszky's disease (pseudorabies) virus (ADV), a glycoprotein complex consisting of three glycoproteins, gBa, gBb, and gBc, was produced by two research groups and was used for the topographical epitope mapping of gB. An epitope map was constructed in which the identified epitopes of gB were situated in 14 topologically distinct antigenic domains; ten antigenic domains represented by 22 mAbs were localized on gBc, while four antigenic domains represented by four mAbs resided on gBb of the gB complex. All the epitopes located on gBc appeared to be conformation-dependent, whereas all the epitopes on gBb were conformation-independent.