scholarly journals Detection of immunogenic protein from salivary gland of Aedes albopictus

2021 ◽  
Vol 40 (3) ◽  
pp. 234-242
Author(s):  
Rike Oktarianti ◽  
Rochmatul Nuryu Khasanah ◽  
Syubbanul Wathon ◽  
Kartika Senjarini
Keyword(s):  
Salivary Gland ◽  
Western Blot ◽  
Salivary Glands ◽  
Aedes Albopictus ◽  
Blot Analysis ◽  
Gel Electrophoresis ◽  
Western Blot Analysis ◽  
Sodium Dodecyl ◽  
Immunogenic Proteins ◽  
Healthy Persons

BackgroundDengue virus is transmitted by several species of Aedes mosquitoes, with Aedes albopictus as secondary vector. During blood feeding, these vectors inject saliva into the vertebrate hosts. The saliva contains anticoagulant, anti-inflammatory and immunogenic factors. The objective of this research was to detect immunogenic proteins from Ae.albopictus salivary glands reacting with sera of people living in dengue endemic areas. MethodsThe identification of immunogenic proteins of Ae. albopictus salivary gland used one-dimensional gel electrophoresis (sodium dodecyl sulfate polyacrylamide gel electrophoresis), and western blot analysis, respectively. To determine the immunogenic nature of the candidate proteins, the antigens from the salivary gland of Ae. albopictus were reacted with sera from healthy persons, dengue hemorrhagic fever (DHF) patients, and neonates, each of the groups comprising 10 samples. ResultsThe protein profiles of Ae. albopictus salivary glands showed 13 bands with molecular weights from 16 kDa up to 97 kDa, i.e. 16, 17, 26, 28, 31, 32, 45, 55, 60, 67, 73, 76, and 97 kDa. According to western blot analysis result, the 31 kDa proteins were recognized in all endemic population sera, both in DHF patients and healthy persons. In contrast, protein bands of 47 and 67 kDa were only recognized by the sera of DHF patients. ConclusionThree immunogenic proteins of 31, 47 and 67 kDa were detected from Ae. albopictus salivary glands. These immunogenic proteins may be developed as candidate biomarkers for bite exposure to Ae. albopictus and as vector-based DHF vaccines.

scholarly journals Identification of Immmunogenic Salivary Proteins of Anopheles vagus based on Mass Spectrometry Analysis

2017 ◽  
Vol 18 (2) ◽  
pp. 73
Author(s):  
Dwi Esti Febriyantiningsih ◽  
Kartika Senjarini ◽  
Rike Oktarianti
Keyword(s):  
Mass Spectrometry ◽  
Salivary Gland ◽  
Western Blot ◽  
Salivary Glands ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Mass Spectrometry Analysis ◽  
Salivary Proteins ◽  
Immunogenic Proteins ◽  
Anopheles Vagus

Malaria has been prevalent for a long time in tropical developing regions causing great morbidity and mortality. Among the malaria vectors, Anopheles vagus has been known as secondary malaria vector in East Java. Salivary glands of mosquitoes perform various functions for survival of the vectors and also conducive for blood feeding, harbouring of malaria parasites, and eventual parasite transmission. The salivary gland proteomes of An. vagus have not been carried out yet. The aim of our study was to identify and characterize the immunogenic proteins of salivary glands proteins of An. vagus. A proteomic approach combining one-dimensional electrophoresis (1DE) followed by western blot analysis using human sera from healthy people living in an endemic area (Kendal); liquid chromatography mass spectrometry (LC-MS/MS) and bioinformatic analysis was adopted to provide the first direct insight into identification and characterization of salivary proteins of An. vagus. Identification of immunogenic proteins using western blot analysis has revealed three immunogenic bands which had molecular weights of 69, 75 and 232 kDa. Among those proteins analysed by LC-MS/MS, there were alpha,1-4 glucan phosphorylase, putative myosin class I heavy chain which have the highest number of total spectrum count peptide. Other proteins like vitellogenin and heat shock protein 82 (Hsp82) were also identified. The majority of proteins were scrutinized marked for their role in metabolism, cytoskeleton protein and stress response. Keywords: Anopheles vagus, salivary gland, immunogenic, proteomics

scholarly journals Investigations on ornithobacterium rhinotracheale in broiler flocks in elazig province located in the east of turkey

2012 ◽  
Vol 49 (No. 8) ◽  
pp. 305-311 ◽  
Author(s):  
G. Ozbey ◽  
H. Ongor ◽  
D. T Balik ◽  
V. Celik ◽  
A. Kilic ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Sodium Dodecyl ◽  
Enzyme Linked Immunosorbent Assay ◽  
Rrna Gene ◽  
Major Band ◽  
Cell Extracts ◽  
Sds Page ◽  
Ornithobacterium Rhinotracheale

In the present study, lung, trachea and serum samples from broiler flocks slaughtered at an abattoir in Elazig province located in the East of Turkey were examined for the presence of Ornithobacterium rhinotracheale using culture and enzyme-linked immunosorbent assay (ELISA). The identity was latter proved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), western blot analysis, and polymerase chain reaction (PCR) assays. A total of 324 serum and 250 lung and trachea samples were collected from 10 commercially reared chicken flocks showing respiratory manifestations. The samples were obtained from different flocks. The causative agent (ORT) was isolated from trachea (1.5%) of five chickens and from both lung and trachea (0.4%) of only one chicken in the bacteriological examination of tissues. The presence of antibodies against ORT was detected in 33 (10.2%) of the 324 sera by ELISA. A 784 bp fragment of the 16S rRNA gene was amplified using specific primers in the PCR. All ORT isolates that were positive by culture were also detected to be positive by the PCR. SDS-PAGE protein profiles of whole cell extracts showed a high similarity for all the isolates with a major band of the molecular weight of 33 kDa (kiloDalton). Results of Western blot analysis indicate four antigenic fractions predominantly with molecular weights of 33, 42, 52 and 66 kDa.

A Proteomic Approach to Discovery of Candidate Proteins Involved in the Transformation of Follicular Lymphoma.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 984-984
Author(s):  
Guang Fan ◽  
Yanping Zhong ◽  
Cristina Smith ◽  
James Huang ◽  
Rita Braziel
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Gel Electrophoresis ◽  
Western Blot Analysis ◽  
Protein Identification ◽  
Differentially Expressed ◽  
Signal Transduction Pathways ◽  
Differentially Expressed Proteins ◽  
2D Gel ◽  
2D Gel Analysis

Abstract Background: Follicular lymphoma (FL) undergoes transformation to a high grade diffuse large B-cell lymphoma (tr-DLBCL) in about 50% of patients. During transformation, a more virulent subclone of tumor cells emerges, leading to a rapidly progressive clinical course and resistance to therapy. The identification of proteins involved in transformation is critical for understanding the mechanism of transformation and developing molecularly targeted therapy. In this study, we compared protein expression between grade 1- FL (G1-FL) and tr-DLBCL using 2D-gel electrophoresis and Western blot analysis. Design: Frozen tissue and frozen cells were obtained from the Department of Pathology, Oregon Health and Science University tumor bank. The protein expression profiles of 3 G1-FL and 3 tr-DLBCL were compared using 2D-gel electrophoresis. Protein identification was done using a MALDI mass spectrometer. Frozen cells of an additional 11 non-paired GI-FL and 11 non-paired tr-DLBCL, and 2 pairs of G1-FL and tr-DLBCL specimens were used for Western blot confirmation of the initial 2D-gel findings. Results: 2D-gel analysis and MALDI protein identification revealed 14 differentially expressed proteins between G1-FL and tr-DLBCL (figure 1), all of which are known to play important roles in cellular energy/metabolic pathways, signal transduction pathways, and protein and nuclear synthesis. The two most differentially expressed proteins on 2D-gel analysis were superoxide dismutase (MnSOD2) and growth factor receptor bound protein 2 (Grb2). Western blot analysis of MnSOD2 and Grb2 confirmed their relative over- or under-expression in frozen cells from multiple additional clinical lymphoma samples, including 2 paired- and 22 non-paired G1-FL and tr-DLBCL. Both 2D-gel analysis and Western Blot showed a significantly higher level of expression of MnSOD2 and a lower expression of Grb2 expression in tr-DLBCL (figure 2). Summary: Using proteomic profiling, confirmed by Western blot analysis of clinical G1-FL and tr-DLBCL samples, we have confirmed 2 proteins (MnSOD2 and Grb2) that are expressed at significantly different levels in G1-FL and DLBCL. MnSOD2 is capable of protecting cells from reactive oxygen species and regulating signal transduction pathways to influence cell growth and apoptosis. Inhibition of MnSOD2 has been shown in studies of several cancer cell lines to render cancer cells more susceptible to apoptosis. Grb2 is a member of a critical signaling pathway leading to Ras activation in hematopoietic cells. Both proteins may play a critical role in FL transformation. These proteins have the potential to be therapeutic drug targets, diagnostic and/or prognostic markers, or biomarkers for monitoring therapeutic response. Summary of Differentially Expressed Spots Summary of Differentially Expressed Spots Figure Figure

scholarly journals Subcellular Localization of the Intracellular Survival-Enhancing Eis Protein of Mycobacterium tuberculosis

2001 ◽  
Vol 69 (7) ◽  
pp. 4295-4302 ◽  
Author(s):  
John L. Dahl ◽  
Jun Wei ◽  
James W. Moulder ◽  
Suman Laal ◽  
Richard L. Friedman
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Amino Acid ◽  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Intracellular Survival ◽  
Terminal Amino Acid ◽  
Terminal Amino

ABSTRACT Mycobacterium tuberculosis is a facultative intracellular pathogen that has evolved the ability to survive and multiply within human macrophages. It is not clear how M. tuberculosis avoids the destructive action of macrophages, but this ability is fundamental in the pathogenicity of tuberculosis. A gene previously identified in M. tuberculosis, designatedeis, was found to enhance intracellular survival ofMycobacterium smegmatis in the human macrophage-like cell line U-937 (J. Wei et al., J. Bacteriol. 182:377–384, 2000). Wheneis was introduced into M. smegmatis on a multicopy vector, sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed the appearance of a unique 42-kDa protein band corresponding to the predicted molecular weight of the eisgene product. This band was electroeluted from the gel with a purity of >90% and subjected to N-terminal amino acid sequencing, which demonstrated that the 42-kDa band was indeed the protein product ofeis. The Eis protein produced by M. tuberculosis H37Ra had an identical N-terminal amino acid sequence. A synthetic polypeptide corresponding to a carboxyl-terminal region of the deduced eis protein sequence was used to generate affinity-purified rabbit polyclonal antibodies that reacted with the 42-kDa protein in Western blot analysis. Hydropathy profile analysis showed the Eis protein to be predominantly hydrophilic with a potential hydrophobic amino terminus. Phase separation of M. tuberculosis H37Ra lysates by the nonionic detergent Triton X-114 revealed the Eis protein in both the aqueous and detergent phases. After fractionation of M. tuberculosis by differential centrifugation, Eis protein appeared mainly in the cytoplasmic fraction but also in the membrane, cell wall, and culture supernatant fractions as well. Forty percent of the sera from pulmonary tuberculosis patients tested for anti-Eis antibody gave positive reactions in Western blot analysis. Although the function of Eis remains unknown, evidence presented here suggests it associates with the cell surface and is released into the culture medium. It is produced during human tuberculosis infection and therefore may be an important M. tuberculosis immunogen.

Isolation of an aspartic proteinase precursor from the egg of a hard tick, Boophilus microplus

Parasitology ◽  
1998 ◽  
Vol 116 (6) ◽  
pp. 525-532 ◽  
Author(s):  
C. LOGULLO ◽  
I. DA SILVA VAZ ◽  
M. H. F. SORGINE ◽  
G. O. PAIVA-SILVA ◽  
F. S. FARIA ◽  
...  
Keyword(s):  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Fat Body ◽  
Sodium Dodecyl ◽  
Aspartic Proteinase ◽  
Boophilus Microplus ◽  
Proteolytic Processing ◽  
Protein Sequencing ◽  
Hard Tick

An aspartic proteinase precursor, herein named BYC (Boophilus Yolk pro-Cathepsin) was isolated from eggs of the hard tick, Boophilus microplus. As judged by electrophoresis on sodium dodecyl sulfate polyacrylamide slab gel (SDS–PAGE), purified BYC presented 2 bands of 54 and 49 kDa, bearing the same NH2-terminal amino acid sequence. By Western blot analysis, BYC was also found in the haemolymph, indicating an extraovarian site of synthesis. Several organs were incubated in culture medium with [35S] methionine, and only the gut and fat body showed synthesis of BYC polypeptides. Protein sequencing of both the NH2-terminal and an internal sequence obtained after cyanogen bromide (CNBr) cleavage of BYC revealed homology with several aspartic proteinase precursors. Incubation at pH 3·5 resulted in autoproteolysis of BYC, which produced the mature form of the enzyme, that displayed pepstatin-sensitive hydrolytic activity against haemoglobin. Western blot analysis using anti-BYC monoclonal antibodies showed proteolytic processing of BYC during embryogenesis and suggested activation of the enzyme during development. A role of BYC in degradation of vitellin, the major yolk protein of tick eggs, is discussed.

scholarly journals Serological cross-reactivity of environmental Escherichia coli strains with Shigella-specific antisera

2018 ◽  
Vol 33 (1-2) ◽  
pp. 29-33 ◽  
Author(s):  
Nafisa Azmuda ◽  
Rabeya Bilkis ◽  
Humaira Akter ◽  
Anowara Begum ◽  
Sirajul Islam Khan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Western Blot ◽  
Blot Analysis ◽  
Western Blot Analysis ◽  
Surface Antigens ◽  
Biochemical Properties ◽  
Cross Reactivity ◽  
Freshwater Ecosystems ◽  
Shigella Species ◽  
Immunogenic Proteins

Many bacteria of clinical and environmental origin show evidence of sharing common surface antigens. The present study aimed for isolation of Escherichia coli strains that were serologically cross-reactive with Shigella species from freshwater ecosystems in Bangladesh by conventional cultural methods. Among twenty eight isolates, two isolates, termed 12(35) and 6(50) showed cross-reactivity with four polyvalent serogroup-specific Shigella antisera using slide agglutination assay. The isolates were identified and charcterized by cultural and biochemical properties and Western blot analysis. The isolates showed typical Escherichia coli cell morphology and cultural and biochemical properties and were identified as Escherichia coli by API 20E tests. Western blot analysis confirmed the isolates as cross-reactive with all the four group-specific Shigella antisera due to presence of immunogenic proteins and LPS. One of the isolates also showed cross-reactivity with multiple type-specific Shigella boydii antisera (monovalent) because of immunogenic proteins. Both the isolates were identified as nonpathogenic due to absence of virulence marker genes of diarrheagenic E. coli variants.This study revealed that a number of bacteria present in the environment could share important Shigella species surface antigens. Naturally occurring nonpathogenic environmental bacteria expressing surface antigens specific for certain types of Shigella could be a good choice for vaccine candidates against shigellosis. Bangladesh J Microbiol, Volume 33, Number 1-2, June-Dec 2016, pp 29-33

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