Recombinant glycoprotein G analog for determination of specific immunoglobulins to herpes simplex virus type 2 by ELISA. 2013

Ludmila Korshun, and Mariya Vudmaska, and Larissa Moysa, and Galina Kovtonjuk, and Svetlana Mikhalap, and Larissa Ganova, and Nikolay Spivak
JSC "Diaproph-Med", Kyiv, Ukraine. Electronic address: KL2004@ukr.net.

In order to the detection of type-specific IgG to herpes simplex virus type 2 (HSV-2) in human serum or plasma the recombinant analog of HSV-2 glycoprotein G (gG2) was created. To construct an expression vector the DNA fragment with a sequence identical to immunodominant regions of HSV-2 gG2 was cloned into modified vector pET28a containing of the glutation-S-transferase sequence (pET28-GST). Escherichia coli BL21 (DE3) were transformed with the recombinant plasmid. The target protein was expressed mainly in soluble form. Chromatographic purification of soluble GST-gG2 protein was performed taking into account the features of its primary structure that are 6His-tag and GST-tag. To determine the affinity constant of the specific IgG to GST-gG2 we used the method proposed by Friguet et al. (1985). The affinity constants were within the range of 10(7)-10(8)M(-1) proving their high-affinity. The purified recombinant HSV-2 antigen was used to design a diagnostic ELISA kit, which was evaluated with referent controls and standard panels of sera containing and/or not containing anti-HSV-2 IgG. Comparative evaluation of this kit and the commercially available "HSV-Type 2 IgG-ELISA" (NovaTec, Dietzenbach, Germany) kit was performed. There was no significant difference (P>0.05). It allows to use developed ELISA kit for clinical diagnosis of HSV-2 infection.

UI MeSH Term Description Entries
D007136 Immunoglobulins Multi-subunit proteins which function in IMMUNITY. They are produced by B LYMPHOCYTES from the IMMUNOGLOBULIN GENES. They are comprised of two heavy (IMMUNOGLOBULIN HEAVY CHAINS) and two light chains (IMMUNOGLOBULIN LIGHT CHAINS) with additional ancillary polypeptide chains depending on their isoforms. The variety of isoforms include monomeric or polymeric forms, and transmembrane forms (B-CELL ANTIGEN RECEPTORS) or secreted forms (ANTIBODIES). They are divided by the amino acid sequence of their heavy chains into five classes (IMMUNOGLOBULIN A; IMMUNOGLOBULIN D; IMMUNOGLOBULIN E; IMMUNOGLOBULIN G; IMMUNOGLOBULIN M) and various subclasses. Globulins, Immune,Immune Globulin,Immune Globulins,Immunoglobulin,Globulin, Immune
D011993 Recombinant Fusion Proteins Recombinant proteins produced by the GENETIC TRANSLATION of fused genes formed by the combination of NUCLEIC ACID REGULATORY SEQUENCES of one or more genes with the protein coding sequences of one or more genes. Fusion Proteins, Recombinant,Recombinant Chimeric Protein,Recombinant Fusion Protein,Recombinant Hybrid Protein,Chimeric Proteins, Recombinant,Hybrid Proteins, Recombinant,Recombinant Chimeric Proteins,Recombinant Hybrid Proteins,Chimeric Protein, Recombinant,Fusion Protein, Recombinant,Hybrid Protein, Recombinant,Protein, Recombinant Chimeric,Protein, Recombinant Fusion,Protein, Recombinant Hybrid,Proteins, Recombinant Chimeric,Proteins, Recombinant Fusion,Proteins, Recombinant Hybrid
D003955 Diagnostic Tests, Routine Diagnostic procedures, such as laboratory tests and x-rays, routinely performed on all individuals or specified categories of individuals in a specified situation, e.g., patients being admitted to the hospital. These include routine tests administered to neonates. Admission Tests, Routine,Hospital Admission Tests,Physical Examination, Preadmission,Routine Diagnostic Tests,Admission Tests, Hospital,Diagnostic Test, Routine,Diagnostic Tests,Examination, Preadmission Physical,Preadmission Physical Examination,Routine Diagnostic Test,Test, Routine Diagnostic,Tests, Diagnostic,Tests, Hospital Admission,Tests, Routine Diagnostic,Admission Test, Hospital,Admission Test, Routine,Diagnostic Test,Examinations, Preadmission Physical,Hospital Admission Test,Physical Examinations, Preadmission,Preadmission Physical Examinations,Routine Admission Test,Routine Admission Tests,Test, Diagnostic,Test, Hospital Admission,Test, Routine Admission,Tests, Routine Admission
D004797 Enzyme-Linked Immunosorbent Assay An immunoassay utilizing an antibody labeled with an enzyme marker such as horseradish peroxidase. While either the enzyme or the antibody is bound to an immunosorbent substrate, they both retain their biologic activity; the change in enzyme activity as a result of the enzyme-antibody-antigen reaction is proportional to the concentration of the antigen and can be measured spectrophotometrically or with the naked eye. Many variations of the method have been developed. ELISA,Assay, Enzyme-Linked Immunosorbent,Assays, Enzyme-Linked Immunosorbent,Enzyme Linked Immunosorbent Assay,Enzyme-Linked Immunosorbent Assays,Immunosorbent Assay, Enzyme-Linked,Immunosorbent Assays, Enzyme-Linked
D004926 Escherichia coli A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc. Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D006558 Herpes Genitalis Infection of the genitals (GENITALIA) with HERPES SIMPLEX VIRUS in either the males or the females. Genital Herpes,Herpes Simplex, Genital,Herpes Simplex Virus Genital Infection,Genital Herpes Simplex,Herpes, Genital
D006801 Humans Members of the species Homo sapiens. Homo sapiens,Man (Taxonomy),Human,Man, Modern,Modern Man
D000914 Antibodies, Viral Immunoglobulins produced in response to VIRAL ANTIGENS. Viral Antibodies
D000956 Antigens, Viral Substances elaborated by viruses that have antigenic activity. Viral Antigen,Viral Antigens,Antigen, Viral
D014759 Viral Envelope Proteins Integral membrane proteins that are incorporated into the VIRAL ENVELOPE. They are glycosylated during VIRAL ASSEMBLY. Envelope Proteins, Viral,Viral Envelope Glycoproteins,Viral Envelope Protein,Virus Envelope Protein,Virus Peplomer Proteins,Bovine Leukemia Virus Glycoprotein gp51,Hepatitis Virus (MHV) Glycoprotein E2,LaCrosse Virus Envelope Glycoprotein G1,Simian Sarcoma Virus Glycoprotein 70,Viral Envelope Glycoprotein gPr90 (Murine Leukemia Virus),Viral Envelope Glycoprotein gp55 (Friend Virus),Viral Envelope Proteins E1,Viral Envelope Proteins E2,Viral Envelope Proteins gp52,Viral Envelope Proteins gp70,Virus Envelope Proteins,Envelope Glycoproteins, Viral,Envelope Protein, Viral,Envelope Protein, Virus,Envelope Proteins, Virus,Glycoproteins, Viral Envelope,Peplomer Proteins, Virus,Protein, Viral Envelope,Protein, Virus Envelope,Proteins, Viral Envelope,Proteins, Virus Envelope,Proteins, Virus Peplomer

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