Displaying publications 1 - 20 of 73 in total

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  1. Fulltext The role of blood transfusion in HCV infection: a study testing Ab: RNA & genotype
    Al-Kubaisy, Waqar A., Niazi, Amjad D.
    Int J Public Health Res, 2011;1(2):72-78.
    MyJurnal
    Introduction Hepatitis C Virus (HCV) recently was identified as a major cause of post transfusion hepatitis world wide. To evaluate the role of blood transfusion on the prevalence of HCV infection, by testing antibody and RNA as well as the genotypes of HCV .Also to detect if Blood transfusion acts as unconfounding risk factor for HCV infection.
    Methods Sera from 3491 pregnant women were investigated for the presence of HCV antibodies (anti-HCV) by using third generation enzyme immunoassay (EIA-3) as screening test, followed by immunoblot assay (Lia Tek-III). In addition 94 sera of studied women were subjected to molecular analysis (at laboratories of Sorin BioMedica - Italy) for the detection of viral RNA and genotypes of HCV. Using RT-PCR & DNA Enzyme immunoassay (DEIA) method.
    Results Our study revealed, that seroprevalence rate of HCV specific Ab & RNA were significantly higher (16.32 %, 80% respectively) among women with a history of blood transfusion, compared to those (2.53%, 56.5%) with no such history P=0.0001, P=0.01. And there is a significant direct linear correlation between number of blood transfused and the seropositive rate of anti-HCV (r=0.7, p=0.046). Based on multivariate analysis, interestingly, this study confirmed that, blood transfusion significantly acting as unconfounding risk factor for acquiring HCV infection (Adjusted OR=1.938,95% C.I=1.646-2.28). And the risk of exposure is increases with increased number of blood transfused. Although, we found no significant association between, HCV genotypic distribution and history of blood transfusion. However, high proportion of women with a history of blood transfusion were harboring HCV genotype -4 or 1b, 50%,40%, resepctively.
    Conclusions Our study shows, evidence that, blood transfusion acts as unconfounding risk factor for acquiring and in a mode of transmission of HCV infection. Therefore strict screening of blood donor for HCV-Abs and / or RNA is highly recommended.
    Matched MeSH terms: Immunoblotting
  2. The protective effect of Mucuna pruriens seeds against snake venom poisoning
    Tan NH, Fung SY, Sim SM, Marinello E, Guerranti R, Aguiyi JC
    J Ethnopharmacol, 2009 Jun 22;123(2):356-8.
    PMID: 19429384 DOI: 10.1016/j.jep.2009.03.025
    The seed, leaf and root of Mucuna pruriens have been used in traditional medicine for treatments of various diseases. In Nigeria, the seed is used as oral prophylactics for snakebite.
    Matched MeSH terms: Immunoblotting
  3. Fulltext The differential expression of aqueous soluble proteins in breast normal and cancerous tissues in relation to stage and grade of patients
    Liang S, Singh M, Gam LH
    J Biomed Biotechnol, 2010;2010:516469.
    PMID: 21197096 DOI: 10.1155/2010/516469
    Breast cancer is a leading cause of female deaths worldwide. In Malaysia, it is the most common form of female cancer while Infiltrating ductal carcinoma (IDC) is the most common form of breast cancer. A proteomic approach was used to identify changes in the protein profile of breast cancerous and normal tissues. The patients were divided into different cohorts according to tumour stage and grade. We identified twenty-four differentially expressed hydrophilic proteins. A few proteins were found significantly related to various stages and grades of IDC, amongst which were SEC13-like 1 (isoform b), calreticulin, 14-3-3 protein zeta, and 14-3-3 protein eta. In this study, we found that by defining the expression of the proteins according to stages and grades of IDC, a significant relationship between the expression of the proteins with the stage or grade of IDC can be established, which increases the usefulness of these proteins as biomarkers for IDC.
    Matched MeSH terms: Immunoblotting
  4. Fulltext The Sec61 translocon limits IRE1α signaling during the unfolded protein response
    Sundaram A, Plumb R, Appathurai S, Mariappan M
    Elife, 2017 05 15;6.
    PMID: 28504640 DOI: 10.7554/eLife.27187
    IRE1α is an endoplasmic reticulum (ER) localized endonuclease activated by misfolded proteins in the ER. Previously, we demonstrated that IRE1α forms a complex with the Sec61 translocon, to which its substrate XBP1u mRNA is recruited for cleavage during ER stress (Plumb et al., 2015). Here, we probe IRE1α complexes in cells with blue native PAGE immunoblotting. We find that IRE1α forms a hetero-oligomeric complex with the Sec61 translocon that is activated upon ER stress with little change in the complex. In addition, IRE1α oligomerization, activation, and inactivation during ER stress are regulated by Sec61. Loss of the IRE1α-Sec61 translocon interaction as well as severe ER stress conditions causes IRE1α to form higher-order oligomers that exhibit continuous activation and extended cleavage of XBP1u mRNA. Thus, we propose that the Sec61-IRE1α complex defines the extent of IRE1α activity and may determine cell fate decisions during ER stress conditions.
    Matched MeSH terms: Immunoblotting
  5. Studies on apolipoprotein(a) phenotypes. Part 1. Phenotype frequencies in a healthy Japanese population
    Abe A, Noma A
    Atherosclerosis, 1992 Sep;96(1):1-8.
    PMID: 1418098
    The frequency distribution for serum lipoprotein(a) (Lp(a)) concentrations in healthy Japanese was highly skewed, with a mean +/- S.D. of 14.6 +/- 13.6 mg/dl and a median of 11.0 mg/dl. The present study provides the first evidence on the frequencies of Lp(a) phenotypes and alleles in healthy Japanese subjects. There was a strong inverse relationship between the apparent molecular weights of apo(a) isoforms and plasma Lp(a) concentrations, as reported previously. However, because of the considerable overlap between the Lp(a) concentrations of the different phenotypes, it was impossible to predict Lp(a) concentration from Lp(a) phenotypes, or vice versa. The present results suggest that the distribution of Lp(a) concentrations, mean and median values and Lp(a) phenotype and allele frequencies in healthy Japanese are not significantly different from the results for Europeans, whereas they are significantly different from other Asian populations, i.e. Chinese, Indians and Malaysians.
    Matched MeSH terms: Immunoblotting
  6. Fulltext Sero-prevalence study of IgE responses to allergens from Malaysian house dust (HDM) and storage mites (SM)
    Chong KT, Wong SF, Mak JW, Loh LC, Ho TM
    Trop Biomed, 2015 Sep;32(3):524-39.
    PMID: 26695214 MyJurnal
    Allergens of Dermatophagoides and Blomia species are well-characterized but not for other species. This study was conducted to determine the prevalence of allergic sensitization to house dust (HDM) and storage mites (SM). One hundred adult subjects (aged ≥ 18) were recruited. The mite specific IgE of all allergic subjects were higher compared with healthy subjetcs despite being not statistically significant except for D. farinae and G. malaysiensis. The mean serum IgE levels against HDM and SM for allergic subjects were significantly higher compared with those in healthy subjects. They were mainly sensitized to Dermatophagoides farinae (35%) and Glycycometus malaysiensis (37%). Immunoblots revealed not all allergic subjects showed positive immuno-reactivity against the mites tested. Single or multiple bands were observed for different species. The subjects were commonly sensitized to Group 2 (9-12 kDa), 10 (38 kDa) and 18 (40-48 kDa) allergens. Twenty-one out of 60 allergic subjects were sensitized to either one or more species. The majority of them (71%) were sensitized to single species. The allergic subjects were mainly sensitized to D. pteronyssinus, followed by Tyrophagus putrecentiae and Aleuroglyphus ovatus. Seven were solely sensitized to HDM while 10 were solely sensitized to SM. Four subjects were sensitized to both. Pre-adsorption study revealed no cross-reactivity. There was difference between the prevalence and reactivity to allergens of HDM and SM in these subjects. Both ELISA and immunoblot did not correlate well but can complement each other in improving the detection of mite allergens to the species level.
    Matched MeSH terms: Immunoblotting
  7. Fulltext Scalable Production of Recombinant Membrane Active Peptides and Its Potential as a Complementary Adjunct to Conventional Chemotherapeutics
    Rothan HA, Ambikabothy J, Abdulrahman AY, Bahrani H, Golpich M, Amini E, et al.
    PLoS One, 2015;10(9):e0139248.
    PMID: 26418816 DOI: 10.1371/journal.pone.0139248
    The production of short anticancer peptides in recombinant form is an alternative method for costly chemical manufacturing. However, the limitations of host toxicity, bioactivity and column purification have impaired production in mass quantities. In this study, short cationic peptides were produced in aggregated inclusion bodies by double fusion with a central protein that has anti-cancer activity. The anticancer peptides Tachiplicin I (TACH) and Latarcin 1 (LATA) were fused with the N- and C-terminus of the MAP30 protein, respectively. We successfully produced the recombinant TACH-MAP30-LATA protein and MAP30 alone in E. coli that represented 59% and 68% of the inclusion bodies. The purified form of the inclusion bodies was prepared by eliminating host cell proteins through multiple washing steps and semi-solubilization in alkaline buffer. The purified active protein was recovered by inclusive solubilization at pH 12.5 in the presence of 2 M urea and refolded in alkaline buffer containing oxides and reduced glutathione. The peptide-fusion protein showed lower CC50 values against cancer cells (HepG2, 0.35±0.1 μM and MCF-7, 0.58±0.1 μM) compared with normal cells (WRL68, 1.83±0.2 μM and ARPE19, 2.5±0.1 μM) with outstanding activity compared with its individual components. The presence of the short peptides facilitated the entry of the peptide fusion protein into cancer cells (1.8 to 2.2-fold) compared with MAP30 alone through direct interaction with the cell membrane. The cancer chemotherapy agent doxorubicin showed higher efficiency and selectivity against cancer cells in combination with the peptide- fusion protein. This study provides new data on the mass production of short anticancer peptides as inclusion bodies in E. coli by fusion with a central protein that has similar activity. The product was biologically active against cancer cells compared with normal cells and enhanced the activity and selective delivery of an anticancer chemotherapy agent.
    Matched MeSH terms: Immunoblotting
  8. Regional and accelerated molecular evolution in group I snake venom gland phospholipase A2 isozymes
    Chuman Y, Nobuhisa I, Ogawa T, Deshimaru M, Chijiwa T, Tan NH, et al.
    Toxicon, 2000 Mar;38(3):449-62.
    PMID: 10669032
    In accordance with detection of a few phospholipase A2 (PLA2) isozyme genes by Southern blot analysis, only two cDNAs, named NnkPLA-I , and NnkPLA-II, encoding group I PLA2s, NnkPLA-I and NnkPLA-II, respectively, were isolated from the venom gland cDNA library of Elapinae Naja naja kaouthia of Malaysia. NnkPLA-I and NnkPLA-II showed four amino acid substitutions, all of which were brought about by single nucleotide substitution. No existence of clones encoding CM-II and CM-III, PLA2 isozymes which had been isolated from the venom of N. naja kaouthia of Thailand, in Malaysian N. naja kaouthia venom gland cDNA library was verified by dot blot hybridization analysis with particular probes. NnkPLA-I and NnkPLA-II differed from CM-II and CM-III with four and two amino acid substitutions, respectively, suggesting that their molecular evolution is regional. The comparison of NnkPLA-I, NnkPLA-II and cDNAs encoding other group I snake venom gland PLA2s indicated that the 5'- and 3'-untranslated regions are more conserved than the mature protein-coding region and that the number of nucleotide substitutions per nonsynonymous site is almost equal to that per synonymous site in the protein-coding region, suggesting that accelerated evolution has occurred in group I venom gland PLA2s possibly to acquire new physiological functions.
    Matched MeSH terms: Immunoblotting
  9. Fulltext Recombinant proteins from new constructs of SAG1 and GRA7 sequences and their usefulness to detect acute toxoplasmosis
    Kotresha D, Poonam D, Muhammad Hafiznur Y, Saadatnia G, Nurulhasanah O, Sabariah O, et al.
    Trop Biomed, 2012 Mar;29(1):129-37.
    PMID: 22543613 MyJurnal
    In this study we have cloned unreported gene fragments of Toxoplasma gondii GRA7 and SAG1 and expressed the corresponding recombinant proteins, followed by evaluation of their usefulness for the serological diagnosis of toxoplasmosis. Both recombinant proteins were expressed efficiently in insoluble form, purified by single step Ni-NTA affinity chromatography and their antigenicity to detect toxoplasma specific IgG antibodies were determined by immunoblotting. A total of 60 serum samples from three groups of individuals based on their anti-toxoplasma antibody profiles were tested, namely (I) IgM+, IgG+ (n=20), (II) IgM-, IgG+ (n=20) and (III) IgM-, IgG- (n=20). Both recombinant proteins exhibited high sensitivity (100%) with sera from Group I. rGRA7 and rSAG1 reacted 40% and 80% respectively with Group II sera. The specificity of the recombinant proteins based on reactivities with Group III sera were 100% and 80% with rGRA7 and rSAG1 respectively. Thus rGRA7 was found to be better at discriminating probable acute from chronic phases of toxoplasmosis, and it also showed higher specificity.
    Matched MeSH terms: Immunoblotting/methods
  10. Recombinant expression of the larval excretory-secretory antigen TES-120 of Toxocara canis in the methylotrophic yeast Pichia pastoris
    Fong MY, Lau YL
    Parasitol Res, 2004 Jan;92(2):173-6.
    PMID: 14655048
    A gene encoding the larval excretory-secretory antigen TES-120 of the dog ascarid worm Toxocara canis was cloned into the methylotrophic yeast Pichia pastoris. Specificity of the recombinant TES-120 antigen produced by the yeast was investigated. Forty-five human serum samples from patients infected with different()parasitic organisms, including 8 cases of toxocariasis, were tested against the recombinant antigen in immunoblot assays. Results from the assays showed that the recombinant TES-120 antigen reacted with sera from toxocariasis patients only. This highly specific recombinant TES-120 antigen can potentially be used for the development of an inexpensive serodiagnostic assay for human toxocariasis.
    Matched MeSH terms: Immunoblotting
  11. Recombinant expression of Toxocara canis excretory-secretory antigen TES-120 in Escherichia coli
    Fong MY, Lau YL, Init I, Jamaiah I, Anuar AK, Rahmah N
    Southeast Asian J. Trop. Med. Public Health, 2003 Dec;34(4):723-6.
    PMID: 15115078
    The gene encoding the excretory-secretory antigen TES-120 of dog ascarid worm Toxocara canis was cloned into the bacterium Escherichia coli. The specificity of the recombinant TES-120 antigen produced by the bacterium was investigated. A total of 45 human serum samples from patients infected with differenthelminthes and protozoa, including 8 cases of toxocariasis, were tested against the recombinant antigens in immunoblot assays. The results from the assays revealed that the recombinant TES-120 antigen reacted with sera from toxocariasis patients only. This highly specific recombinant TES-120 antigen can potentially be used for the development of an inexpensive serodiagnostic assay for human toxocariasis.
    Matched MeSH terms: Immunoblotting
  12. Fulltext Potential use of human hair shaft keratin peptide signatures to distinguish gender and ethnicity
    Mohamed Nasir N, Hiji J, Jayapalan JJ, Hashim OH
    PeerJ, 2020;8:e8248.
    PMID: 32030317 DOI: 10.7717/peerj.8248
    Background: Most human hairs collected at old crime scenes do not contain nuclear DNA and are therefore of less value for forensic investigations. In the present study, hair shaft proteins were extracted from 40 healthy subjects between the ages of 21 to 40 years and profiled using gel electrophoresis-based proteomics to determine if they can be used to distinguish gender and ethnicity.

    Methods: Extraction of the human hair shaft proteins was performed using a newly developed alkaline solubilisation method. The extracts were profiled by 2-dimensional electrophoresis and resolved protein spots were identified by mass spectrometry and queried against the human hair database. The study was then followed-up by immunoblotting of the identified hair shaft keratin of interest using commercially available antibodies.

    Results: Separation of the human hair shaft proteins by 2-dimensional electrophoresis generated improved and highly resolved profiles. Comparing the hair shaft protein profiles of 10 female with 10 male subjects and their identification by mass spectrometry and query of the human hair database showed significant altered abundance of truncated/processed type-II keratin peptides K81 (two spots), K83 (one spot) and K86 (three spots). The 2-dimensional electrophoresis profiling of 30 hair shaft samples taken from women of similar age range but from three distinctive ethnic subpopulations in Malaysia further showed significant altered abundance of one type-I and four type-II truncated/processed keratin peptides including K33b, K81, K83 and K86 (2 spots) between at least two of the ethnic groups. When a followed-up immunoblotting experiment was performed to detect the relative expression of the K86 peptides using commercialised antibodies, similar trends of expression were obtained. The present data, when taken together, demonstrated the potential use of keratin peptide signatures of the human hair shaft to distinguish gender and ethnicity although this needs to be further substantiated in a larger scale study.

    Matched MeSH terms: Immunoblotting
  13. Population genetics of coagulation factor XIIIB in three ethnic groups of Singapore
    Saha N, Tay JS, Low PS, Basair JB
    Ann Hum Biol, 1992 5 1;19(3):277-83.
    PMID: 1616285
    The distribution of plasma coagulation factor XXIIB polymorphism was determined by PAG isoelectric focusing and immunoblotting in a group of 670 subjects comprising 375 Chinese, 110 Malays and 185 Indians. The frequencies of FXIIIB*1, FXIIIB*2, and FXIIIB*3 were found to be 0.27, 0.03 and 0.70 in the Chinese; 0.33, 0.05 and 0.64 in the Malays and 0.58, 0.08 and 0.33 in the Indians. The phenotypic distribution of FXIIIB alleles was at Hardy-Weinberg equilibrium in all three populations. A two-dimensional principal-components analysis on the basis of three common alleles at the FXIIIB locus among 19 populations, so far studied, clearly differentiates the Negroid, Mongoloid and Caucasoid populations into three major groups with the exception of Amerindians (Minnesota) and US Blacks showing some Caucasoid influence.
    Matched MeSH terms: Immunoblotting
  14. Penyaringan domain ADP-ribosilasi gen toksin daripada Burkholderia pseudomallei virulen
    Shahrul Hisham Zainal Ariffin, Rahmah Mohamed, Zulkeflie Zamrod, Mohammad Noor Embi
    Sains Malaysiana, 2008;37.
    Bahagian aktif bagi enzim toksin bakteria daripada Burkholderia pseudomallei, Pseudomonas aeruginosa dan difteria merupakan domain ADP-ribosilasi. Domain ini didapati terpelihara di antara ketiga-tiga mikroorganisme. Di dalam kajian ini, domain ADP-ribosilasi Burkholderia pseudomallei telah diamplifikasi daripada genom B. pseudomallei virulen dengan menggunakan pencetus-pencetus yang dibina berdasarkan kepada jujukan domain ADP ribosilasi Pseudomonas aeruginosa. Hasil DNA amplifikasi ditulenkan dan digunakan sebagai prob (HPCR2) untuk menyaring DNA selitan daripada B. pseudomallei yang diklonkan ke dalam vektor pengekspresan pSport-I. Objektif kajian ini adalah untuk menyaring lapan klon yang positif hasil daripada penyaringan awal melalui pendekatan immunoblot menggunakan antitoksin daripada arnab. Penyaringan ini juga melibatkan tiga klon yang tidak memberikan isyarat positif semasa penyaringan secara immunoblot. Keputusan menunjukkan hanya satu klon (L31) daripada lapan klon immunoblot positif mempunyai domain ADP-ribosilasi. Penjujukan DNA separa klon L31 secara manual melibatkan dua pencetus menghasilkan jujukan sepanjang 450pb. Analisis selanjutnya mendapati daripada enam kemungkinan translasi kepada polipeptida hanya satu polipeptida wujud yang tidak mempunyai sebarang kodon penamat pada jujukan kodonnya.
    Matched MeSH terms: Immunoblotting
  15. Overexpression, purification and validation of antigenic Salmonella enterica serovar Typhi proteins identified from LC-MS/MS
    Chin CF, Teh BA, Anthony AA, Aziah I, Ismail A, Ong EB, et al.
    Appl Biochem Biotechnol, 2014 Nov;174(5):1897-906.
    PMID: 25149461 DOI: 10.1007/s12010-014-1173-y
    In our earlier study, an immunoblot analysis using sera from febrile patients revealed that a 50-kDa band from an outer membrane protein fraction of Salmonella enterica serovar Typhi was specifically recognized only by typhoid sera and not sera from other febrile illnesses. Here, we investigated the identities of the proteins contained in the immunogenic 50-kDa band to pinpoint antigens responsible for its immunogenicity. We first used LC-MS/MS for protein identification, then used the online tool ANTIGENpro for antigenicity prediction and produced recombinant proteins of the lead antigens for validation in an enzyme-linked immunosorbent assay (ELISA). We found that proteins TolC, GlpK and SucB were specific to typhoid sera but react to antibodies differently under native and denatured conditions. This difference suggests the presence of linear and conformational epitopes on these proteins.
    Matched MeSH terms: Immunoblotting
  16. Fulltext Mutant Allele-Specific CRISPR Disruption in DYT1 Dystonia Fibroblasts Restores Cell Function
    Cruz L, György B, Cheah PS, Kleinstiver BP, Eimer WA, Garcia SP, et al.
    Mol Ther Nucleic Acids, 2020 Sep 04;21:1-12.
    PMID: 32502938 DOI: 10.1016/j.omtn.2020.05.009
    Most individuals affected with DYT1 dystonia have a heterozygous 3-bp deletion in the TOR1A gene (c.907_909delGAG). The mutation appears to act through a dominant-negative mechanism compromising normal torsinA function, and it is proposed that reducing mutant torsinA may normalize torsinA activity. In this study, we used an engineered Cas9 variant from Streptococcus pyogenes (SpCas9-VRQR) to target the mutation in the TOR1A gene in order to disrupt mutant torsinA in DYT1 patient fibroblasts. Selective targeting of the DYT1 allele was highly efficient with most common non-homologous end joining (NHEJ) edits, leading to a predicted premature stop codon with loss of the torsinA C terminus (delta 302-332 aa). Structural analysis predicted a functionally inactive status of this truncated torsinA due to the loss of residues associated with ATPase activity and binding to LULL1. Immunoblotting showed a reduction of the torsinA protein level in Cas9-edited DYT1 fibroblasts, and a functional assay using HSV infection indicated a phenotypic recovery toward that observed in control fibroblasts. These findings suggest that the selective disruption of the mutant TOR1A allele using CRISPR-Cas9 inactivates mutant torsinA, allowing the remaining wild-type torsinA to exert normal function.
    Matched MeSH terms: Immunoblotting
  17. Fulltext Moringa oleifera Flower Extract Suppresses the Activation of Inflammatory Mediators in Lipopolysaccharide-Stimulated RAW 264.7 Macrophages via NF-κB Pathway
    Tan WS, Arulselvan P, Karthivashan G, Fakurazi S
    Mediators Inflamm, 2015;2015:720171.
    PMID: 26609199 DOI: 10.1155/2015/720171
    Aim of Study. Moringa oleifera Lam. (M. oleifera) possess highest concentration of antioxidant bioactive compounds and is anticipated to be used as an alternative medicine for inflammation. In the present study, we investigated the anti-inflammatory activity of 80% hydroethanolic extract of M. oleifera flower on proinflammatory mediators and cytokines produced in lipopolysaccharide- (LPS-) induced RAW 264.7 macrophages. Materials and Methods. Cell cytotoxicity was conducted by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Nitric oxide (NO) production was quantified through Griess reaction while proinflammatory cytokines and other key inflammatory markers were assessed through enzyme-linked immunosorbent assay (ELISA) and immunoblotting. Results. Hydroethanolic extract of M. oleifera flower significantly suppressed the secretion and expression of NO, prostaglandin E2 (PGE2), interleukin- (IL-) 6, IL-1β, tumor necrosis factor-alpha (TNF-α), nuclear factor-kappa B (NF-κB), inducible NO synthase (iNOS), and cyclooxygenase-2 (COX-2). However, it significantly increased the production of IL-10 and IκB-α (inhibitor of κB) in a concentration dependent manner (100 μg/mL and 200 μg/mL). Conclusion. These results suggest that 80% hydroethanolic extract of M. oleifera flower has anti-inflammatory action related to its inhibition of NO, PGE2, proinflammatory cytokines, and inflammatory mediator's production in LPS-stimulated macrophages through preventing degradation of IκB-α in NF-κB signaling pathway.
    Matched MeSH terms: Immunoblotting
  18. Monoclonal antibodies specific to heat-treated porcine blood
    Raja Nhari RM, Hamid M, Rasli NM, Omar AR, El Sheikha AF, Mustafa S
    J Sci Food Agric, 2016 May;96(7):2524-31.
    PMID: 26611757 DOI: 10.1002/jsfa.7547
    Porcine blood is potentially being utilized in food as a binder, gelling agent, emulsifier or colorant. However, for certain communities, the usage of animal blood in food is strictly prohibited owing to religious concerns and health reasons. This study reports the development of monoclonal antibodies (MAbs) against heat-treated soluble proteins (HSPs) of autoclaved porcine blood; characterization of MAbs against blood, non-blood and plasma from different animal species using qualitative indirect non-competitive enzyme-linked immunosorbent assay (ELISA); and immunoblotting of antigenic components in HSPs of porcine blood.
    Matched MeSH terms: Immunoblotting
  19. Fulltext Malaysian cockle (Anadara granosa) allergy: Identification of IgE-binding proteins and effects of different cooking methods
    Zailatul HM, Rosmilah M, Faizal B, Noormalin A, Shahnaz M
    Trop Biomed, 2015 Jun;32(2):323-34.
    PMID: 26691261 MyJurnal
    The purpose of this study was to evaluate the effect of different cooking methods on the allergenicity of cockle and to identify proteins most frequently bound by IgE antibodies using a proteomics approach. Raw, boiled, fried and roasted extracts of the cockle were prepared. The protein profiles of the extracts were obtained by separation using sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) and 2-dimensional gel electrophoresis (2-DE). IgE-immunoblotting was then performed with the use of individual sera from patients with cockle allergy and the major IgE-binding proteins were analyzed by mass-spectrometry. SDS-PAGE of raw extract showed 13 protein bands. Smaller numbers of protein bands were detected in the boiled, fried and roasted extracts. The 2-DE gel profile of the raw extract further separated the protein bands to ~50 protein spots with molecular masses between 13 to 180 kDa and isoelectric point (pI) values ranging from 3 to 10. Immunoblotting of raw extract exhibited 11 IgE-binding proteins with two proteins of 36 and 40 kDa as the major IgE-binding proteins, while the boiled extract revealed 3 IgE-binding proteins. Fried and roasted extracts only showed a single IgE-binding protein at 36 kDa. 2-DE immunoblotting of raw extract demonstrated 5 to 20 IgE reactive spots. Mass spectrometry analysis led to identification of 2 important allergens, tropomyosin (36 kDa) and arginine kinase (40 kDa). Heated extracts showed a reduction in the number of IgE-reactive bands compared with raw extract, which suggest that thermal treatment can be used as a tool in attempting to reduce cockle allergenicity. The degree of allergenicity of cockle was demonstrated in the order raw > boiled > fried ≈ roasted. Two important allergens reacting with more than 50% of patients' sera identified using mass spectrometric approaches were tropomyosin and arginine kinase. Thus, allergens found in this study would help in component based diagnosis, management of cockle allergic patients and to the standardisation of allergenic test products as tools in molecular allergology.
    Matched MeSH terms: Immunoblotting
  20. Magnoflorine Enhances LPS-Activated Pro-Inflammatory Responses via MyD88-Dependent Pathways in U937 Macrophages
    Haque MA, Jantan I, Harikrishnan H, Abdul Wahab SM
    Planta Med, 2018 Nov;84(17):1255-1264.
    PMID: 29906814 DOI: 10.1055/a-0637-9936
    Magnoflorine, a major bioactive metabolite isolated from Tinospora crispa, has been reported for its diverse biochemical and pharmacological properties. However, there is little report on its underlying mechanisms of action on immune responses, particularly on macrophage activation. In this study, we aimed to investigate the effects of magnoflorine, isolated from T. crispa on the pro-inflammatory mediators generation induced by LPS and the concomitant NF-κB, MAPKs, and PI3K-Akt signaling pathways in U937 macrophages. Differentiated U937 macrophages were treated with magnoflorine and the release of pro-inflammatory mediators was evaluated through ELISA, while the relative mRNA expression of the respective mediators was quantified through qRT-PCR. Correspondingly, western blotting was executed to observe the modulatory effects of magnoflorine on the expression of various markers related to NF-κB, MAPK and PI3K-Akt signaling activation in LPS-primed U937 macrophages. Magnoflorine significantly enhanced the upregulation of TNF-α, IL-1β, and PGE2 production as well as COX-2 protein expression. Successively, magnoflorine prompted the mRNA transcription level of these pro-inflammatory mediators. Magnoflorine enhanced the NF-κB activation by prompting p65, IκBα, and IKKα/β phosphorylation as well as IκBα degradation. Besides, magnoflorine treatments concentration-dependently augmented the phosphorylation of JNK, ERK, and p38 MAPKs as well as Akt. The immunoaugmenting effects were further confirmed by investigating the effects of magnoflorine on specific inhibitors, where the treatment with specific inhibitors of NF-κB, MAPKs, and PI3K-Akt proficiently blocked the magnoflorine-triggered TNF-α release and COX-2 expression. Magnoflorine furthermore enhanced the MyD88 and TLR4 upregulation. The results suggest that magnoflorine has high potential on augmenting immune responses.
    Matched MeSH terms: Immunoblotting
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