Biomedical subjects

V Hari

Publications and source records attributed to V Hari.

18 recordsLinked to original sources

PubMed1998

Immunodominant peptide epitopes of allergen, Asp f 1 from the fungus Aspergillus fumigatus.

Aspergillus fumigatus ribotoxin Asp f 1 is a major allergen with IgE binding activity to serum of a majority of patients with allergic bronchopulmonary aspergillosis (ABPA). The IgE binding epitopes or the T-cell stimulatory peptides of this molecule have not been studied. In the present investigation, we have synthesized linear decapeptides spanning the whole molecule of Asp f 1 and analyzed their IgE binding properties. We have also synthesized peptides based on their possible T-cell stimulatory properties and studied the stimulation of peripheral blood mononuclear cells from ABPA patients and normal controls. Several peptides demonstrated distinct IgE antibody binding response against sera from ABPA patients and proliferative response against peripheral blood mononuclear cells from the patients. From the results, it can be concluded that the carboxy-terminal region of Asp f 1 representing amino acid residues 115-149 involved in both humoral and cell mediated immunoresponses in ABPA.

Allergens↗

PubMed1996

Aspergillus fumigatus peptides differentially express Th1 and Th2 cytokines.

Relevant allergens from Aspergillus fumigatus associated with allergic bronchopulmonary aspergillosis (ABPA) have been cloned and expressed. The pathogenesis of ABPA probably depends on specific cytokines and immunoglobulins secreted by lymphocytes on stimulation with different epitopes of those allergens. In the present study, we synthesized peptides of 12-16 amino acids from the sequence of Asp fI and compared their immunological responses in four mice strains (BALB/c, C57BL/6, AKR, and CBA). Of the five peptides studied for their cytokine profile, one showed a clear Th1, whereas another showed a Th2 response. The remaining three peptides varied in their immunoreactivity. The results suggest that a number of epitopes of diverse activities are present in individual molecules and may be involved in the pathogenesis of ABPA through differential cytokine secretions.

Allergens↗

PubMed1995

Electron microscopic localization of ATPase activity in tobacco cells infected by tobacco etch potyvirus and tobacco mosaic virus.

Thin sections of leaves of plants infected by tobacco etch potyvirus (TEV) or tobacco mosaic virus (TMV) were examined for the presence of ATPase activity by electron microscopy. ATPase activity was found as expected in mitochondria, chloroplasts and plasmalemma of both uninfected as well as cells infected by either TEV or TMV. In the TEV-infected cells, ATPase activity was localized to virus-induced vesicles, endoplasmic reticulum and in some cells, to ribosomes attached to the ER. In TMV-infected cells, ATPase activity was found in vesicles as well as in tubular membranes closely associated with the X-bodies.

Adenosine Triphosphatases↗

PubMed1994

In situ localization of ATPase activity in cells of plants infected by maize dwarf mosaic potyvirus.

Cells of healthy maize plants as well as those infected by maize dwarf mosaic potyvirus were examined by electron microscopy for the location of ATPase activity. In healthy and virus infected plants, ATPase activity was found in plasma membranes, chloroplast thylakoid membranes, nuclear membranes and in mitochondria. In virus-infected cells, ATPase activity was also observed in cytoplasmic vesicles which were found in close proximity to the virus-specific cytoplasmic inclusion bodies (CI), at the ends of the arms of the CI and in plasmodesmata.

Adenosine Triphosphatases↗

PubMed1994

Monoclonal antibodies reactive with specific amino acid sequences of the 126 K protein of tobacco mosaic virus.

The 126 K protein of tobacco mosaic virus (TMV) is an NTP binding protein that has guanylyl transferase activity and is predicted to be an ATPase/helicase. In this paper we report the generation of monoclonal antibodies (Mabs) that react with specific amino acid sequences of the 126K protein. The Mabs were generated after immunizing mice with a partially purified preparation of the 126 K protein (native) obtained by centrifugal fractionation of the infected tissue extracts. The Mabs were assayed for specific reactivity by western blotting and by their reactivity with non-overlapping decapeptides corresponding to the entire amino acid sequence of the 126 K protein of TMV. A total of 11 Mabs reactive with specific peptides and three other Mabs that did not react with any peptide but reacted with the 126 K protein were identified.

Amino Acid Sequence↗

PubMed1993

Detection of antibodies to the HIV-1 p25/24 and gp120 proteins by sphere-linked immunodiagnostic assay (SLIDA).

In this paper, we have used the technique known as sphere-linked immunodiagnostic assay (SLIDA) for the detection of antibodies to HIV-1 p25/24 and gp120 proteins. In this technique, the p25/24 and gp120 proteins were covalently bonded to 0.5 micron carboxylated microspheres. They were then exposed to serial dilutions of rabbit antiserum to HIV-1 p25/24, sheep antiserum to gp120 and dilutions of the corresponding normal sera. Specific immunolabeling was monitored by using gold-labeled secondary antibody. The gold particles bound to the microspheres were an indication for the presence of specific antibody in the serum sample. The gold-labeled microspheres were photographed using a transmission electron microscope, and the number of gold particles bound to the microspheres were counted manually using the electron micrographs. The specificity and sensitivity of the technique were then compared to enzyme-linked immunosorbent assay (ELISA) and Western blotting.

Biotechnology↗

PubMed1992

Intracellular distribution of the 126K/183K and capsid proteins in cells infected by some tobamoviruses.

Leaves of plants infected by tobacco mosaic virus (TMV) strain U1, TMV strain M, tomato mosaic virus strain Dahlemense and tobacco mild green mosaic virus strain U2 were examined for the presence and intracellular distribution of their capsid and 126K/183K proteins by immunoblotting and immunogold electron microscopy. The bulk of the capsid protein was found in the virus bundles (crystals), although small amounts were found in the chloroplasts and nuclei of cells infected by some of these tobamoviruses. The 126K protein of TMV-U1 and -M was localized in the X-bodies, whereas in cells infected by the other two viruses which induce no X-bodies, the 126K protein was found to be associated with virus bundles.

Amino Acid Sequence↗

PubMed1992

R-ELISA: repeated use of antigen-coated plates for ELISA and its application for testing of antibodies to HIV and other pathogens.

In this paper we report on the evaluation of several procedures that allow for the repeated use of an antigen-coated, enzyme-linked immunosorbent assay (ELISA) plate for enzyme immunoassay (EIA). We have shown that antigen-coated ELISA plates that were incubated once with an aqueous solution containing 8 M urea, 2% sodium dodecyl sulfate and 2% mercaptoethanol, after an EIA, can be reused again for EIA without loss of antigenic capacity. Thus, in this procedure, after an EIA, the ELISA plates were washed once with the above solution and then in a buffer containing 20 mM Tris-HCl, pH 7.5, 0.1% Tween 20 and 500 mM NaCl. This washing protocol was shown to remove the primary antibody, enzyme-conjugated secondary antibody and substrate without removing the antigen from the ELISA plate microwells. Thus, an antigen-coated ELISA plate previously used for an assay could be reused. We tested this repeat ELISA (R-ELISA) procedure on high antigen-binding ELISA plates coated with two different plant virus proteins, a synthetic peptide, the p25/24 gag and the gp120 proteins of the human immuno-deficiency virus, or the staphylococcus enterotoxin protein. In each case tested, the procedure allowed for the repeated use of the same antigen-coated plates for EIA of the respective antibodies. This procedure should prove to be particularly valuable for mass screening of samples tested for HIV and other disease-causing agents.

Amino Acid Sequence↗

PubMed1991

A temporal study of the expression of the capsid, cytoplasmic inclusion and nuclear inclusion proteins of tobacco etch potyvirus in infected plants.

Young leaves of tobacco, systemically infected by tobacco etch potyvirus (TEV), were examined for the presence and distribution of four virus encoded proteins [capsid, cytoplasmic inclusion (CI) and two nuclear inclusion (NI) proteins] at various time periods after inoculation of expanded leaves of the plants. The analyses were carried out by ELISA and by immunogold electron microscopy of thin sections of the leaves. All four proteins were detected simultaneously in the systemic leaves for the first time on the fifth day after inoculation of the expanded leaves. All four proteins increased in concentration until the seventh day and then showed no further increase with the exception of the capsid protein which continued to accumulate. The CI protein was first detected in association with the plasmalemma/cell wall and was subsequently found mostly in the form of pinwheels in the cytoplasm. The two NI proteins were found at all times after infection within the nucleus, although small concentrations were detected in the cytoplasm. These experiments suggest that both the NIa and NIb proteins are transported into the nucleus immediately after synthesis. At the earliest time periods after infection, high concentrations of these proteins (NIa and NIb) were found in their non-inclusion form in the nucleolus. At 14 days after infection, both proteins were found only as inclusions in the nucleus. The capsid protein was found at all stages of infection only in the cytoplasm.

Capsid↗

PubMed1990

Potato virus Y helper component protein is associated with amorphous inclusions.

The distribution of the helper component (HC) protein of potato virus Y in tissues and cells of infected plants was studied by immunoblotting and immunogold labelling techniques. This HC protein was found in leaf blade and vein tissue but not in the petiole of leaves. In infected cells, the protein was localized in rod-shaped cytoplasmic inclusions known as amorphous inclusions.

Antibodies, Viral↗

PubMed1990

Sphere-linked immunodiagnostic assay (SLIDA): an electron microscopic method for detecting specific antibodies.

We have developed a sensitive method, sphere-linked immunodiagnostic assay, using specific antigens covalently bonded to microspheres for the detection of antibodies in serum. In this method, specific antigens, such as the capsid proteins of tobacco mosaic virus and tobacco etch virus, were independently, covalently bonded to plastic micropheres of 0.5 microns or 0.9 microns in diameter. The antigen-linked spheres were then exposed to normal serum or serum containing specific antibody, followed by treatment with gold-labeled secondary antibodies. The binding of the gold-labeled secondary antibodies to the specific primary antibodies on the spheres acted as an indication of the presence of the specific primary antibodies. The spheres were then examined and photographed by transmission electron microscopy. The number of gold particles bound to the spheres was counted manually using the photographs. The gold labeling was found to be specific and sensitive, enabling detection of antibodies present in highly diluted antisera. The efficiency and sensitivity of the technique for detection of antibodies were compared with those of the enzyme-linked immunosorbent assay and found to be highly sensitive. The technique was also used for testing for the presence of antibodies to herpes simplex virus as well as antibodies to Staphylococcus enterotoxin using microspheres coated with the respective antigens. We believe that this technique could be applied clinically when needed for detection of antibodies to other viruses, such as the Human Immunodeficiency Virus.

Antibodies↗

PubMed1989

Bean mild mosaic virus: genome and its translation products.

Viral RNA from purified preparations of bean mild mosaic virus (BMMV) was resolved into two RNAs of mol. wt 1.5 X 10(6) daltons (R1) and 0.76 X 10(6) daltons (R2). Complementary DNA hybridizations, infectivity tests, and in-vitro translation studies established that R2 is a subgenomic RNA derived from R1. In-vitro translation studies showed that the genomic RNA (R1) coded for two major proteins of mol. wt. 40.5 kD and 25.7 kD. Of these, the 40.5 kD protein was shown to be the capsid protein that is translated in vitro from both the R1 and R2 RNAs. Time-course studies on the appearance of in-vitro translation products showed that the 25.7 kD product is the earliest product made when the genomic R1 RNA is translated. This is not one of the products of R2 RNA. Several of the characteristics of BMMV described in this paper show similarities with turnip crinkle virus (TCV) and carnation mottle virus (CarMV).

Blotting, Northern↗

PubMed1986

Association of tobacco etch virus related RNA with chloroplasts in extracts of infected plants.

The RNA in various subcellular fractions of tobacco etch virus (TEV) infected tissue was analyzed for the presence of complementary viral RNA, and double-stranded viral RNA by hybridization with 32P-labeled viral RNA or cDNA probes. Although viral RNA was detected in several cellular fractions, the complementary RNA of full-length size was found exclusively associated with fractions containing chloroplasts. Treatment of RNAs with RNase before hybridization suggested that the virus-related complementary RNA was present in double-stranded form.

Chloroplasts↗

PubMed1981