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Displaying publications 81 - 100 of 226 in total

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Low prevalence of latex sensitivity in South African spina bifida children in Cape Town

Johar A, Lim DL, Arif SA, Hawarden D, Toit GD, Weinberg EG, et al.

Pediatr Allergy Immunol, 2005 Mar;16(2):165-70.
PMID: 15787875

Spina bifida children have a high prevalence of latex allergy in studies reported from Europe and the USA. This study investigated the prevalence of latex allergy in a cohort of 24 spina bifida children at the Red Cross Children's Hospital from Cape Town, South Africa. The children were investigated using a detailed questionnaire, skin prick tests (ALK-Abello), ImmunoCap RASTs, Western blotting and ELISA, using the purified latex proteins Hev b1 and Hev b3 and whole latex preparation. A low overall prevalence of latex sensitization of 16.7% was found in the children. Children who were sensitive reacted to water insoluble to Hev b1 and Hev b3 proteins. The low prevalence of latex sensitization in the South African children may not be entirely explained by stringent latex avoidance. The children were from a low socioeconomic social status and 'hygiene' and other factors should be considered.

Matched MeSH terms: Blotting, Western
Ongoing outbreak of an acute muscular Sarcocystis-like illness among travellers returning from Tioman Island, Malaysia, 2011-2012

Esposito DH, Freedman DO, Neumayr A, Parola P

Euro Surveill, 2012 Nov 08;17(45).
PMID: 23153473

As of 4 November, 2012, 100 patients with an acute muscular Sarcocystis-like illness associated with travel to Tioman Island, Malaysia, have been identified. Thirty-five travelled there mostly during July and August 2011 and 65 mostly during July and August 2012, suggesting an ongoing outbreak. Epidemiological investigations are ongoing. Public health agencies and practicing clinicians should be aware of this rarely-reported disease in humans and consider it as differential diagnosis in travellers returning from Tioman Island.

Matched MeSH terms: Blotting, Western
Induction and characterization of heat shock proteins of Salmonella typhi and their reactivity with sera from patients with typhoid fever

Tang SW, Abubakar S, Devi S, Puthucheary S, Pang T

Infect Immun, 1997 Jul;65(7):2983-6.
PMID: 9199477

The heat shock protein (HSP) response of Salmonella typhi following exposure to elevated growth temperatures was studied. Three major proteins with molecular sizes of 58, 68, and 88 kDa were abundantly expressed when S. typhi cells were shifted from 37 to 45 degrees C and to 55 degrees C. These proteins were also constitutively expressed at 37 degrees C. Western blotting and immunoprecipitation studies with anti-HSP monoclonal antibodies revealed that the 58- and 68-kDa proteins were analogous to the GroEL and DnaK proteins, respectively, of Escherichia coli. These HSPs are also abundantly present in the outer membrane fraction of disrupted cells and, to a lesser extent, in the cytosol. Immunoblotting experiments with sera from patients with a culture-positive diagnosis of typhoid fever showed the presence of antibodies to these HSPs. Nine of twelve sera reacted with the 58-, 68-, and 88-kDa proteins, while three sera reacted only with the 68- and 88-kDa proteins. All 10 sera from healthy individuals showed no binding to these HSPs. In light of the well-documented roles of HSPs in the pathogenesis of microbial infections and as immunodominant antigens, these findings may be relevant for a better understanding of disease processes and for the future development of diagnostic and preventive strategies.

Matched MeSH terms: Blotting, Western
Inhibitory effects of Tualang Honey on experimental breast cancer in rats: a preliminary study

Kadir EA, Sulaiman SA, Yahya NK, Othman NH

Asian Pac J Cancer Prev, 2013;14(4):2249-54.
PMID: 23725121

The study was conducted to determine the effect of Malaysian jungle Tualang Honey (TH) on development of breast cancer induced by the carcinogen 7,12-dimethylbenz(α)anthracene (DMBA) in rats. Forty nulliparous female Sprague-Dawley rats were given 80 mg/kg DMBA then randomly divided into four groups: Group 1 served as a Control while Groups 2, 3 and 4 received 0.2, 1.0 or 2.0 g/kg bodyweight/day of TH, respectively, for 150 days. Results showed that breast cancers in the TH-treated groups had slower size increment and smaller mean tumor size (≤ 2 cm3) compared to Controls (≤ 8 cm3). The number of cancers developing in TH-treated groups was also significantly fewer (P<0.05). Histological grading showed majority of TH-treated group cancers to be of grade 1 and 2 compared to grade 3 in controls. There was an increasing trend of apoptotic index (AI) seen in TH-treated groups with increasing dosage of Tualang Honey, however, the mean AI values of all TH-treated groups were not significantly different from the Control value (p>0.05). In conclusion, Tualang Honey exerted positive modulation effects on DMBA-induced breast cancers in rats in this preliminary study.

Matched MeSH terms: Blotting, Western
Fulltext Antigenic membrane proteins of virulent variant of Entamoeba histolytica HM-1:IMSS

Kumarasamy G, Abdus Sani AA, Olivos-García A, Noordin R, Othman N

Pathog Glob Health, 2020 09;114(6):333-342.
PMID: 32536281 DOI: 10.1080/20477724.2020.1780402

Amoebiasis, caused by Entamoeba histolytica, is one of the leading parasitic infections in the world. This study was aimed at profiling antigenic membrane proteins of a virulent variant of E. histolytica HM-1:IMSS. The membrane proteins were extracted using ProteoExtract® kit (Merck, Darmstadt, Germany) or conventional method, separated using OFFGEL 3100 fractionator (Agilent Technologies, Santa Clara, California), followed by SDS-PAGE and Western blot analysis. Selected antigenic membrane proteins were identified using LC-ESI-MS/MS. Subsequently, the proteins were classified according to their biological processes and predictions were made on membrane and membrane-associated proteins. When the proteins were probed with pooled sera from amoebic liver abscess (ALA) patients, 10 and 15 antigenic proteins with molecular weights 25 to 200 kDa were identified using the ProteoExtract® kit and conventional method, respectively. LC-ESI-MS/MS identified 13 antigenic proteins, and both extraction methods predicted six of them as membrane and membrane-associated proteins. The topmost biological processes which comprised of six proteins were involved in cellular processes.. These antigenic membrane proteins merit further investigations as potential candidates for vaccine studies.

Matched MeSH terms: Blotting, Western
Fulltext Can We Use 2,3,5-Triphenyltetrazolium Chloride-Stained Brain Slices for Other Purposes? The Application of Western Blotting

Sanchez-Bezanilla S, Nilsson M, Walker FR, Ong LK

Front Mol Neurosci, 2019;12:181.
PMID: 31417355 DOI: 10.3389/fnmol.2019.00181

2,3,5-Triphenyltetrazolium chloride (TTC) staining is a commonly used method to determine the volume of the cerebral infarction in experimental stroke models. The TTC staining protocol is considered to interfere with downstream analyses, and it is unclear whether TTC-stained brain samples can be used for biochemistry analyses. However, there is evidence indicating that, with proper optimization and handling, TTC-stained brains may remain viable for protein analyses. In the present study, we aimed to rigorously assess whether TTC can reliably be used for western blotting of various markers. In this study, brain samples obtained from C57BL/6 male mice were treated with TTC (TTC+) or left untreated (TTC-) at 1 week after photothrombotic occlusion or sham surgery. Brain regions were dissected into infarct, thalamus, and hippocampus, and proteins were extracted by using radioimmunoprecipitation assay buffer. Protein levels of apoptosis, autophagy, neuronal, glial, vascular, and neurodegenerative-related markers were analyzed by western blotting. Our results showed that TTC+ brains display similar relative changes in most of the markers compared with TTC- brains. In addition, we validated that these analyses can be performed in the infarct as well as other brain regions such as the thalamus and hippocampus. Our findings demonstrate that TTC+ brains are reliable for protein analyses using western blotting. Widespread adoption of this approach will be key to lowering the number of animals used while maximizing data.

Matched MeSH terms: Blotting, Western
Site-Directed Mutagenesis of Cytochrome P450 2D6 and 2C19 Enzymes: Expression and Spectral Characterization of Naturally Occurring Allelic Variants

Dong AN, Pan Y, Palanisamy UD, Yiap BC, Ahemad N, Ong CE

Appl Biochem Biotechnol, 2018 Sep;186(1):132-144.
PMID: 29524040 DOI: 10.1007/s12010-018-2728-0

Genetic polymorphism of the cytochrome P450 (CYP) genes particularly affects CYP2D6 and CYP2C19 to a functionally relevant extent, and it is therefore crucial to elucidate the enzyme kinetic and molecular basis for altered catalytic activity of these allelic variants. This study explored the expression and function of the reported alleles CYP2D6*2, CYP2D6*10, CYP2D6*17, CYP2C19*23, CYP2C19*24, and CYP2C19*25 with respect to gene polymorphisms. Site-directed mutagenesis (SDM) was carried out to generate these six alleles. After DNA sequencing, the CYP2D6 and CYP2C19 wild types alongside with their alleles were each independently co-expressed with NADPH-CYP oxidoreductase (OxR) in Escherichia coli. The expressed proteins were analyzed using Western blotting, reduced carbon monoxide (CO) difference spectral scanning, and cytochrome c reductase assay. Results from Western blot revealed the presence of all CYP wild-type and allelic proteins in E. coli membrane fractions. The reduced CO difference spectra scanning presented the distinct peak of absorbance at 450 nm, and the cytochrome c reductase assay has confirmed that spectrally active OxR was expressed in each protein preparation. As a conclusion, the results obtained from this study have proven the CYP variants to be immunoreactive and spectrally active and are suitable for use to examine biotransformation and interaction mechanism of the enzymes.

Matched MeSH terms: Blotting, Western
Antibody and T cell responses induced in chickens immunized with avian influenza virus N1 and NP DNA vaccine with chicken IL-15 and IL-18

Lim KL, Jazayeri SD, Yeap SK, Mohamed Alitheen NB, Bejo MH, Ideris A, et al.

Res Vet Sci, 2013 Dec;95(3):1224-34.
PMID: 23948357 DOI: 10.1016/j.rvsc.2013.07.013

We had examined the immunogenicity of a series of plasmid DNAs which include neuraminidase (NA) and nucleoprotein (NP) genes from avian influenza virus (AIV). The interleukin-15 (IL-15) and interleukin-18 (IL-18) as genetic adjuvants were used for immunization in combination with the N1 and NP AIV genes. In the first trial, 8 groups of chickens were established with 10 specific-pathogen-free (SPF) chickens per group while, in the second trial 7 SPF chickens per group were used. The overall N1 enzyme-linked immunosorbent assay (ELISA) titer in chickens immunized with the pDis/N1+pDis/IL-15 was higher compared to the chickens immunized with the pDis/N1 and this suggesting that chicken IL-15 could play a role in enhancing the humoral immune response. Besides that, the chickens that were immunized at 14-day-old (Trial 2) showed a higher N1 antibody titer compared to the chickens that were immunized at 1-day-old (Trial 1). Despite the delayed in NP antibody responses, the chickens co-administrated with IL-15 were able to induce earlier and higher antibody response compared to the pDis/NP and pDis/NP+pDis/IL-18 inoculated groups. The pDis/N1+pDis/IL-15 inoculated chickens also induced higher CD8+ T cells increase than the pDis/N1 group in both trials (P<0.05). The flow cytometry results from both trials demonstrated that the pDis/N1+pDis/IL-18 groups were able to induce CD4+ T cells higher than the pDis/N1 group (P<0.05). Meanwhile, pDis/N1+pDis/IL-18 group was able to induce CD8+ T cells higher than the pDis/N1 group (P<0.05) in Trial 2 only. In the present study, pDis/NP was not significant (P>0.05) in inducing CD4+ and CD8+ T cells when co-administered with the pDis/IL-18 in both trials in comparison to the pDis/NP. Our data suggest that the pDis/N1+pDis/IL-15 combination has the potential to be used as a DNA vaccine against AIV in chickens.

Matched MeSH terms: Blotting, Western/veterinary
Cell Cycle Modulation of MCF-7 and MDA-MB-231 by a Sub- Fraction of Strobilanthes crispus and its Combination with Tamoxifen

Yaacob NS, Nik Mohamed Kamal NN, Wong KK, Norazmi MN

Asian Pac J Cancer Prev, 2015;16(18):8135-40.
PMID: 26745050

BACKGROUND: Cell cycle regulatory proteins are suitable targets for cancer therapeutic development since genetic alterations in many cancers also affect the functions of these molecules. Strobilanthes crispus (S. crispus) is traditionally known for its potential benefits in treating various ailments. We recently reported that an active sub-fraction of S. crispus leaves (SCS) caused caspase-dependent apoptosis of human breast cancer MCF-7 and MDA-MB-231 cells.
MATERIALS AND METHODS: Considering the ability of SCS to also promote the activity of the antiestrogen, tamoxifen, we further examined the effect of SCS in modulating cell cycle progression and related proteins in MCF-7 and MDA-MB-231 cells alone and in combination with tamoxifen. Expression of cell cycle- related transcripts was analysed based on a previous microarray dataset.
RESULTS: SCS significantly caused G1 arrest of both types of cells, similar to tamoxifen and this was associated with modulation of cyclin D1, p21 and p53. In combination with tamoxifen, the anticancer effects involved downregulation of ERα protein in MCF-7 cells but appeared independent of an ER-mediated mechanism in MDA-MB-231 cells. Microarray data analysis confirmed the clinical relevance of the proteins studied.
CONCLUSIONS: The current data suggest that SCS growth inhibitory effects are similar to that of the antiestrogen, tamoxifen, further supporting the previously demonstrated cytotoxic and apoptotic actions of both agents.

Matched MeSH terms: Blotting, Western
Cloning and expression of malaria and tuberculosis epitopes in mycobacterium bovis bacille calmette-guérin

Suppian R, Zainuddin ZF, Norazmi MN

Malays J Med Sci, 2006 Jan;13(1):13-20.
PMID: 22589585

Mycobacterium bovis bacille Calmette-Guèrin (BCG) represents one of the most promising live vectors for the delivery of foreign antigens to the immune system. A recombinant BCG containing a synthetic gene coding for the malarial epitopes namely, the fragment 2 of region II of EBA-175 (F2R(II)EBA) and the repeat sequence of the circumsporozoite protein NANP generated in favour of mycobacterium codon usage using assembly PCR was constructed. Two T-cell epitopes of the 6-kDa M. tuberculosis early-secreted antigenic target (ESAT-6) antigen were also clone in the same construct. Expression of the synthetic gene was driven by the heat shock protein 65 (hsp65) promoter from M. tuberculosis and the signal peptide from the MPT63 antigen of M. tuberculosis. Expression of the composite epitopes was detected by Western blotting of the cell extract and culture supernatant of the recombinant clones using a specific rabbit polyclonal antibody against F2R(II)EBA. This study demonstrates the possibility of cloning and expressing immunogenic epitopes from causative agents of two important diseases: malaria and tuberculosis (TB) in a single recombinant BCG construct.

Matched MeSH terms: Blotting, Western
Fulltext Vitrification of Blastocyst Murine embryos affects PI3K pathway by modulating the expression of XIAP and S6K1 proteins

Mohd Fazirul, M., Sharaniza, A.R., Norhazlin, J.M.Y., Wan Hafizah, W.J., Razif, D., Froemming, G.R.A., et al.

Pertanika Journal of Science & Technology, 2017;25(108):187-198.
MyJurnal

Cryopreservation by vitrification has been widely used in Assisted Reproductive Technology (ART) to preserve embryos for an extended period of time. However, the effect of vitrification on development of the embryos is lacking. Therefore, understanding on vitrification effects on embryonic proteins, especially those involved in preimplantation development is crucial to provide high quality embryos for further usage. In this study, XIAP and S6K1 protein expressions following vitrification was investigated, since they have been implicated in diverse cellular processes including cell growth, migration, proliferation, differentiation, survival and development of preimplantation embryos via the PI3K pathway. Embryos were obtained from superovulated female ICR mice which were mated with fertile males. The embryos were harvested at the 2-cell stage and cultured until blastocyst stage. Blastocysts were then vitrified in ESF40 cryoprotectant. Western blot was carried out to determine the expression of XIAP and S6K1 proteins. The results showed the expression of XIAP and S6K1 significantly decreased in vitrified blastocyst compared to the control. This indicates that blastocyst vitrification may impact developmental competence through the activation of apoptotic pathways.

Matched MeSH terms: Blotting, Western
Fulltext Helicobacter pylori recombinant UreG protein: cloning, expression, and assessment of its seroreactivity

Khalilpour A, Osman S, Yunus MH, Santhanam A, Vellasamy N, Noordin R

BMC Res Notes, 2014;7:809.
PMID: 25406411 DOI: 10.1186/1756-0500-7-809

Helicobacter pylori is a human pathogen and during the process of infection, antigens from the bacterium elicit strong host humoral immune responses. In our previous report, native H. pylori UreG protein showed good reactivity with sera from H. pylori patients. This study was aimed at producing the recombinant form of the protein (rUreG) and determining its seroreactivities.

Matched MeSH terms: Blotting, Western
Antigenic proteins of Helicobacter pylori of potential diagnostic value

Khalilpour A, Santhanam A, Wei LC, Saadatnia G, Velusamy N, Osman S, et al.

Asian Pac J Cancer Prev, 2013;14(3):1635-42.
PMID: 23679248

Helicobacter pylori antigen was prepared from an isolate from a patient with a duodenal ulcer. Serum samples were obtained from culture-positive H. pylori infected patients with duodenal ulcers, gastric ulcers and gastritis (n=30). As controls, three kinds of sera without detectable H. pylori IgG antibodies were used: 30 from healthy individuals without history of gastric disorders, 30 from patients who were seen in the endoscopy clinic but were H. pylori culture negative and 30 from people with other diseases. OFF-GEL electrophoresis, SDS-PAGE and Western blots of individual serum samples were used to identify protein bands with good sensitivity and specificity when probed with the above sera and HRP-conjugated anti-human IgG. Four H. pylori protein bands showed good (≥ 70%) sensitivity and high specificity (98-100%) towards anti-Helicobacter IgG antibody in culture- positive patients sera and control sera, respectively. The identities of the antigenic proteins were elucidated by mass spectrometry. The relative molecular weights and the identities of the proteins, based on MALDI TOF/ TOF, were as follows: CagI (25 kDa), urease G accessory protein (25 kDa), UreB (63 kDa) and proline/pyrroline- 5-carboxylate dehydrogenase (118 KDa). These identified proteins, singly and/or in combinations, may be useful for diagnosis of H. pylori infection in patients.

Matched MeSH terms: Blotting, Western
Entamoeba histolytica antigenic protein detected in pus aspirates from patients with amoebic liver abscess

Othman N, Mohamed Z, Yahya MM, Leow VM, Lim BH, Noordin R

Exp Parasitol, 2013 Aug;134(4):504-10.
PMID: 23680184 DOI: 10.1016/j.exppara.2013.05.001

Entamoeba histolytica is a causative agent of amoebic liver abscess (ALA) and is endemic in many underdeveloped countries. We investigated antigenic E. histolytica proteins in liver abscess aspirates using proteomics approach. Pus samples were first tested by real-time PCR to confirm the presence of E. histolytica DNA and the corresponding serum samples tested for E. histolytica-specific IgG by a commercial ELISA. Proteins were extracted from three and one pool(s) of pus samples from ALA and PLA (pyogenic liver abscess) patients respectively, followed by analysis using isoelectric focussing, SDS-PAGE and Western blot. Unpurified pooled serum samples from infected hamsters and pooled human amoebic-specific IgG were used as primary antibodies. The antigenic protein band was excised from the gel, digested and analysed by MALDI-TOF/TOF and LC-MS/MS. The results using both primary antibodies showed an antigenic protein band of ∼14kDa. Based on the mass spectrum analysis, putative tyrosine kinase is the most probable identification of the antigenic band.

Matched MeSH terms: Blotting, Western
Toxoplasma gondii excretory secretory antigenic proteins of diagnostic potential

Saadatnia G, Mohamed Z, Ghaffarifar F, Osman E, Moghadam ZK, Noordin R

APMIS, 2012 Jan;120(1):47-55.
PMID: 22151308 DOI: 10.1111/j.1600-0463.2011.02810.x

Infection with Toxoplasma gondii is widespread and important in humans, especially pregnant women and immunosuppressed patients. A panel of tests is usually required for diagnosis toxoplasmosis. Excretory secretory antigen (ESA) is highly immunogenic, and thus it is a good candidate for investigation into new infection markers. ESA was prepared from tachyzoites of RH strain of T. gondii by mice intraperitoneal infection. Sera were obtained from several categories of individuals who differed in their status of anti-Toxoplasma IgM, IgG and IgG avidity antibodies. The ESA was subjected to SDS-PAGE, two-dimensional gel electrophoresis and Western blot analysis. Antigenic bands of approximate molecular weights of 12, 20 and 30 kDa, when probed with anti-human IgM-HRP and IgA-HRP, showed good potential as infection markers. The highest sensitivity of the bands was 98.7% with combination of IgM and IgA blots with sera of patients with anti-Toxoplasma IgM+ IgG+. The specificities were 84% and 70% with sera from other infections and healthy controls in IgM blots and IgA blots respectively. By mass spectrometry, the 12 kDa protein was identified as thioredoxin. The two top proteins identified for 20 kDa molecule were microneme protein 10 and dense granule protein 7; whereas that for 30 kDa were phosphoglycerate mutase 1 and phosphoglycerate mutase.

Matched MeSH terms: Blotting, Western
Detection of immunogenic parasite and host-specific proteins in the sera of active and chronic individuals infected with Toxoplasma gondii

Yeng C, Osman E, Mohamed Z, Noordin R

Electrophoresis, 2010 Dec;31(23-24):3843-9.
PMID: 21080484 DOI: 10.1002/elps.201000038

Toxoplasma gondii infection in pregnant women may result in abortion and foetal abnormalities, and may be life-threatening in immunocompromised hosts. To identify the potential infection markers of this disease, 2-DE and Western blot methods were employed to study the parasite circulating antigens and host-specific proteins in the sera of T. gondii-infected individuals. The comparisons were made between serum protein profiles of infected (n=31) and normal (n=10) subjects. Antigenic proteins were identified by immunoblotting using pooled sera and monoclonal anti-human IgM-HRP. Selected protein spots were characterised using mass spectrometry. Prominent differences were observed when serum samples of T. gondii-infected individuals and normal controls were compared. A significant up-regulation of host-specific proteins, α(2)-HS glycoprotein and α(1)-B glycoprotein, was also observed in the silver-stained gels of both active and chronic infections. However, only α(2)-HS glycoprotein and α(1)-B glycoprotein in the active infection showed immunoreactivity in Western blots. In addition, three spots of T. gondii proteins were detected, namely (i) hypothetical protein chrXII: 3984434-3 TGME 49, (ii) dual specificity protein phosphatase, catalytic domain TGME 49 and (iii) NADPH-cytochrome p450 reductase TGME 49. Thus, 2-DE approach followed by Western blotting has enabled the identification of five potential infection markers for the diagnosis of toxoplasmosis: three are parasite-specific proteins and two are host-specific proteins.

Matched MeSH terms: Blotting, Western
Mature Erythrocyte Surface Antigen Protein Identified in the Serum of Plasmodium falciparum-Infected Patients

Zainudin NS, Othman N, Muhi J, Abdu Sani AA, Noordin R

Am J Trop Med Hyg, 2015 Dec;93(6):1268-73.
PMID: 26392156 DOI: 10.4269/ajtmh.15-0333

This study was performed to identify circulating Plasmodium falciparum proteins in patient serum, which may be useful as diagnostic markers. Depletion of highly abundant proteins from each pooled serum sample obtained from P. falciparum-infected patients and healthy individuals was performed using the Proteoseek Antibody-Based Albumin/IgG Removal Kit (Thermo Scientific, Rockford, IL). In analysis 1, the depleted serum was analyzed directly by NanoLC-MS/MS. In analysis 2, the depleted serum was separated by two-dimensional electrophoresis followed by western blot analysis. Subsequently, the selected band was analyzed by NanoLC-MS/MS. The result of analysis 1 revealed the presence of two mature erythrocyte surface antigen (MESA) proteins and chloroquine resistance transporter protein (PfCRT). In addition, analysis 2 revealed an antigenic 75-kDa band when the membrane was probed with purified IgG from the pooled serum obtained from P. falciparum-infected patients. MS/MS analysis of this protein band revealed fragments of P. falciparum MESA proteins. Thus, in this study, two different analyses revealed the presence of Plasmodium MESA protein in pooled serum from malaria patients; thus, this protein should be further investigated to determine its usefulness as a diagnostic marker.

Matched MeSH terms: Blotting, Western
Production of Toxocara cati TES-120 Recombinant Antigen and Comparison with its T. canis Homolog for Serodiagnosis of Toxocariasis

Zahabiun F, Sadjjadi SM, Yunus MH, Rahumatullah A, Moghaddam MH, Saidin S, et al.

Am J Trop Med Hyg, 2015 Aug;93(2):319-25.
PMID: 26033026 DOI: 10.4269/ajtmh.15-0190

Toxocariasis is a cosmopolitan zoonotic disease caused by the infective larvae of Toxocara canis and T. cati. Diagnosis in humans is usually based on clinical symptoms and serology. Immunoglobulin G (IgG)-enzyme-linked immunosorbent assay kits using T. canis excretory-secretory (TES) larval antigens are commonly used for serodiagnosis. Differences in the antigens of the two Toxocara species may influence the diagnostic sensitivity of the test. In this study, T. cati recombinant TES-120 (rTES-120) was cloned, expressed, and compared with its T. canis homolog in an IgG4-western blot. The diagnostic sensitivity and specificity of T. cati rTES-120 were 70% (33/47) and 100% (39/39), respectively. T. canis rTES-120 showed 57.4% sensitivity and 94.4% specificity. When the results of assays using rTES-120 of both species were considered, the diagnostic sensitivity was 76%. This study shows that using antigens from both Toxocara species may improve the serodiagnosis of toxocariasis.

Matched MeSH terms: Blotting, Western
Fulltext Identification, production and assessment of two Toxoplasma gondii recombinant proteins for use in a Toxoplasma IgG avidity assay

Teh AY, Amerizadeh A, Osman S, Yunus MH, Noordin R

Pathog Glob Health, 2016 Oct-Dec;110(7-8):277-286.
PMID: 27697019

The IgG avidity assay is an important tool in the management of suspected toxoplasmosis in pregnant women. This study aimed to produce new Toxoplasma gondii recombinant proteins and to assess their usefulness in an IgG avidity assay. Toxoplasma positive and negative serum samples were used, the former were categorized into low (LGA) and high (HGA) IgG avidity samples. Immunoblots were performed on 30 T. gondii cDNA clones to determine the reactivity and IgG avidity to the expressed proteins. Two of the clones were found to have diagnostic potential and were analyzed further; AG12b encoded T. gondii apical complex lysine methyltransferase (AKMT) protein and AG18 encoded T. gondii forkhead-associated (FHA) domain-containing protein. The His-tagged recombinant proteins, rAG12b and rAG18, were expressed and tested with LGA and HGA samples using an IgG avidity western blot and ELISA. With the IgG avidity western blot, rAG12b identified 86.4% of LGA and 90.9% of HGA samples, whereas rAG18 identified 81.8% of both LGA and HGA samples. With the IgG avidity ELISA, rAG12b identified 86.4% of both LGA and HGA samples, whereas rAG18 identified 77.3% of LGA and 86.4% of HGA serum samples. This study showed that the recombinant antigens were able to differentiate low avidity and high avidity serum samples, suggesting that they are potential candidates for use in the Toxoplasma IgG avidity assay.

Matched MeSH terms: Blotting, Western
Fulltext Identification and Preliminary Evaluation of a Novel Recombinant Protein for Serodiagnosis of Strongyloidiasis

Arifin N, Yunus MH, Nolan TJ, Lok JB, Noordin R

Am J Trop Med Hyg, 2018 04;98(4):1165-1170.
PMID: 29436335 DOI: 10.4269/ajtmh.17-0697

Strongyloides stercoralis is a human parasite that can cause a long-term infection. In immunosuppressed patients, strongyloidiasis may be fatal when there is overwhelming autoinfection resulting in the migration of large numbers of larvae through many organs. Definitive diagnosis is still a challenge, and a combination of symptoms, microscopic identification, and serology test results are often used to arrive at a clinical decision. However, intermittent larval excretion, low parasite burden, and occult infections are challenges with parasitological diagnosis of infection with S. stercoralis. Meanwhile, serologic tests using immunoglobulin G and parasite antigen extract have problems of cross-reactivity with other helminthic infections. Recombinant antigen-based serodiagnosis is a good alternative to overcome the laboratory diagnostic issues. Herein, we report on the isolation of cDNA clone encoding an antigen of potential diagnostic value identified from immunoscreening of a S. stercoralis cDNA library. The translated protein had highest similarity to Strongyloides ratti immunoglobulin-binding protein 1. The recombinant antigen produced, rSs1a, was assessed using western blot and enzyme-linked immunosorbent assay. The latter showed 96% diagnostic sensitivity and 93% specificity; thus, rSs1a has good potential for use in serodiagnosis of human strongyloidiasis.

Matched MeSH terms: Blotting, Western

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