Displaying publications 1 - 20 of 54 in total

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  1. Yuasa I, Umetsu K, Shotake T, Ishida T, Takenaka O, Terao K, et al.
    Electrophoresis, 1990 Oct;11(10):840-5.
    PMID: 2079025
    Genetic variation of orosomucoid (ORM) in the genus Macaca was investigated. Plasma samples were subjected to isoelectric focusing in a pH range of 4-6.5, followed by immunoprinting with anti-human ORM antibodies. A total of 25 alleles were identified in 231 Asian macaques belonging to 13 species from 23 populations and 22 members belonging to a family of M. fascicularis. Family data presented evidence for a codominant mode of inheritance with multi-alleles at a single autosomal locus. A population study revealed enormous intra- and interspecies variations. The heterozygosity values varied from 0.855 in M. fascicularis (Malaysia) to 0.000 in M. radiata (India), M. silenus (India) and M. arctoides (Malaysia).
  2. Bon MC
    Electrophoresis, 1996 Jul;17(7):1248-52.
    PMID: 8855412
    A combination of a modified Feret' (Silvae Genet. 1971, 20, 46-50) extraction buffer and two types of electrophoresis with acrylamide and starch gels were used to characterize allozymes in mature vegetative tissue of a commercially high value species of rattans (Calamus subinermis). From the analysis of allelic segregation from single maternal rattans and their offspring, genetic control of the 16 observed banding zones, which were consistently scorable, was assumed. Seventeen gene loci were identified. The percentage of polymorphic loci within Calamus subinermis was much higher (70.5%) than expected levels of genetic diversity for tropical woody and non-woody species. It is thought that the protocol described may be applied to the analysis of the genetic diversity of all the endangered Calamus species.
    Matched MeSH terms: Electrophoresis/methods*; Electrophoresis, Starch Gel/methods
  3. Doustjalali SR, Yusof R, Yip CH, Looi LM, Pillay B, Hashim OH
    Electrophoresis, 2004 Jul;25(14):2392-401.
    PMID: 15274022
    We have analyzed unfractionated sera of newly diagnosed patients (n=10) with breast carcinoma (BC), prior to treatment, and patients (n=5) with fibrocystic disease of the breast (FDB) by two-dimensional gel electrophoresis (2-DE) and silver staining. The patients' 2-DE serum protein profiles obtained were then subjected to image analysis and compared to similar data generated from sera of normal healthy female controls (n=10) of the same range of age. The relative expression of alpha1-antichymotrypsin (ACT), clusterin, and complement factor B was significantly higher in all BC patients as compared to normal controls. However, the expression of alpha1-antitrypsin (AAT) in BC patients was apparently lower than that of the controls. Similar differential expression of ACT was detected in the FDB patients. The aberrant expression of the serum acute-phase proteins of patients with BC and FDB was confirmed by competitive enzyme-linked immunosorbent assay (ELISA). Similar altered proteins expression was also observed from immunohistochemical studies of malignant (n=5) and benign (n=5) breast lesions of the respective patients performed using antisera to the aberrantly expressed proteins. However, the malignant breast lesions were instead positively stained for AAT. The differential expression of the serum proteins was apparently abrogated when a six-month follow-up study was performed on nine of the BC patients subsequent to treatment.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
  4. Abdul-Rahman PS, Lim BK, Hashim OH
    Electrophoresis, 2007 Jun;28(12):1989-96.
    PMID: 17503403
    The expression of high-abundance serum proteins in newly diagnosed patients with endometrial adenocarcinoma (EACa), squamous cell cervical carcinoma (SCCa) and cervical adenocarcinoma (ACCa), relative to control female subjects, was analyzed by subjecting serum samples to 2-DE followed by image analysis of the silver-stained protein profiles. The three cohorts of cancer patients demonstrated different altered expression of serum high-abundance proteins compared to negative control women. The expression of alpha1-antitrypsin, alpha1-B glycoprotein, cleaved high-molecular-weight kininogen (light chain) and antithrombin III were consistently altered in all the patients. However, clusterin was upregulated only in the patients with EACa, while those with SCCa and ACCa were typically characterized by the upregulated expression of zinc alpha-2-glycoprotein. The aberrant expression of selective serum proteins in the various cohorts of cancer patients was validated by competitive ELISA as well as by lectin detection. Analysis by using the champedak galactose binding lectin further highlighted an unidentified protein that may be differently glycosylated in the sera of the EACa patients that were studied.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional*
  5. Mohamed E, Abdul-Rahman PS, Doustjalali SR, Chen Y, Lim BK, Omar SZ, et al.
    Electrophoresis, 2008 Jun;29(12):2645-50.
    PMID: 18494030 DOI: 10.1002/elps.200700828
    A 35 kDa glycoprotein whose abundance was previously demonstrated to be enhanced in sera of patients with endometrial adenocarcinoma (n = 12), was isolated from pooled sera of three of the cancer patients using champedak galactose-binding lectin affinity chromatography in the present study. Subjecting it to 2-DE and MS/MS, the glycoprotein was identified as the O-glycosylated fragment of inter-alpha-trypsin inhibitor heavy chain H4 (ITIH4). When compared to control sera (n = 17), expression of the 35 kDa ITIH4 cleavage fragment was demonstrated to be significantly enhanced in sera of patients with breast carcinoma (n = 10), epithelial ovarian carcinoma (n = 10), and germ cell ovarian carcinoma (n = 10) but not in patients with nasopharyngeal carcinoma (n = 13) and osteosarcoma (n = 7). The lectin-based electrophoretic bioanalytical method adopted in the present study may be used to assess the physiological relevance of ITIH4 fragmentation and its correlation with different malignancies, their stages and progression.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
  6. Wan Ibrahim WA, Warno SA, Aboul-Enein HY, Hermawan D, Sanagi MM
    Electrophoresis, 2009 Jun;30(11):1976-82.
    PMID: 19517438 DOI: 10.1002/elps.200800499
    An efficient method for the simultaneous enantioseparation of cyproconazole, bromuconazole, and diniconazole enantiomers was developed by CD-modified MEKC using a dual mixture of neutral CDs as chiral selector. Three neutral CDs namely hydroxypropyl-beta-CD, hydroxypropyl-gamma-CD, and gamma-CD were tested as chiral selectors at different concentrations ranging from 10, 20, 30 and 40 mM, but enantiomers of the studied fungicides were not completely separated. The best dual chiral recognition mode for the simultaneous separation of cyproconazole, bromuconazole, and diniconazole enantiomers was achieved with a mixture of 27 mM hydroxypropyl-beta-CD and 3 mM hydroxypropyl-gamma-CD in 25 mM phosphate buffer (pH 3.0) containing 40 mM SDS to which methanol-acetonitrile (10%:5% v/v) was added as organic modifiers. The best separation was based on the appearance of 10 peaks simultaneously, with good resolution (R(s) 1.1-15.9), and peak efficiency (N>200,000). Good repeatabilities in the migration time, peak area, and peak height were obtained in terms of RSD ranging from (0.72 to 1.06)%, (0.39 to 3.49)%, and (1.90 to 4.84)%, respectively.
  7. Chew FN, Tan WS, Ling TC, Tey BT
    Electrophoresis, 2009 Sep;30(17):3017-3023.
    PMID: 19685471 DOI: 10.1002/elps.200900246
    Mechanical and non-mechanical breakages of bacterial cells are usually the preliminary steps in intracellular protein purification. In this study, the recombinant green fluorescent protein (GFP) was purified from intact Escherichia coli cells using preparative PAGE. In this purification process, cells disruption step is not needed. The cellular content of E. coli was drifted out electrically from cells and the negatively charged GFP was further electroeluted from polyacrylamide gel column. SEM investigation of the electrophoresed cells revealed substantial structural damage at the cellular level. This integrated purification technique has successfully recovered the intracellular GFP with a yield of 82% and purity of 95%.
    Matched MeSH terms: Electrophoresis, Polyacrylamide Gel/methods*
  8. See HH, Hauser PC, Ibrahim WA, Sanagi MM
    Electrophoresis, 2010 Jan;31(3):575-82.
    PMID: 20119968 DOI: 10.1002/elps.200900380
    Rapid and direct online preconcentration followed by CE with capacitively coupled contactless conductivity detection (CE-C(4)D) is evaluated as a new approach for the determination of glyphosate, glufosinate (GLUF), and aminophosphonic acid (AMPA) in drinking water. Two online preconcentration techniques, namely large volume sample stacking without polarity switching and field-enhanced sample injection, coupled with CE-C(4)D were successfully developed and optimized. Under optimized conditions, LODs in the range of 0.01-0.1 microM (1.7-11.1 microg/L) and sensitivity enhancements of 48- to 53-fold were achieved with the large volume sample stacking-CE-C(4)D method. By performing the field-enhanced sample injection-CE-C(4)D procedure, excellent LODs down to 0.0005-0.02 microM (0.1-2.2 microg/L) as well as sensitivity enhancements of up to 245- to 1002-fold were obtained. Both techniques showed satisfactory reproducibility with RSDs of peak height of better than 10%. The newly established approaches were successfully applied to the analysis of glyphosate, glufosinate, and aminophosphonic acid in spiked tap drinking water.
    Matched MeSH terms: Electrophoresis, Capillary/methods*
  9. Seriramalu R, Pang WW, Jayapalan JJ, Mohamed E, Abdul-Rahman PS, Bustam AZ, et al.
    Electrophoresis, 2010 Jul;31(14):2388-95.
    PMID: 20575108 DOI: 10.1002/elps.201000164
    The use of lectin affinity chromatography prior to 2-DE separation forms an alternative method to unmask the expression of targeted glycoproteins of lower abundance in serum samples. Reduced expression of alpha-2 macroglobulin (AMG) and complement factor B (CFB) was detected in sera of patients with nasopharyngeal carcinoma (NPC) when pooled serum samples of the patients and those of healthy individuals were subjected to affinity isolation using immobilized champedak mannose-binding lectin and analyzed by 2-DE and densitometry. The AMG and CFB spots were not detected in the 2-DE protein profiles when the same pooled serum samples were subjected to albumin and IgG depletion and neither were they detected when the depleted samples were analyzed by western blotting and lectin detection. Together with other acute-phase response proteins that were previously reported to be altered in expression in NPC patients, AMG and CFB may serve as useful complementary biomarkers for NPC.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional/methods
  10. Al Azzam KM, Saad B, Aboul-Enein HY
    Electrophoresis, 2010 Sep;31(17):2957-63.
    PMID: 20690150 DOI: 10.1002/elps.201000266
    Binding constants for the enantiomers of modafinil with the negatively charged chiral selector sulfated-β-CD (S-β-CD) using CE technique is presented. The calculations of the binding constants employing three different linearization plots (double reciprocal, X-reciprocal and Y-reciprocal) were performed from the electrophoretic mobility values of modafinil enantiomers at different concentrations of S-β-CD in the BGE. The highest inclusion affinity of the modafinil enantiomers were observed for the S-enantiomer-S-β-CD complex, in agreement with the computational calculations performed previously. Binding constants for each enantiomer-S-β-CD complex at different temperatures, as well as thermodynamic parameters for binding, were calculated. Host-guest binding constants using the double reciprocal fit showed better linearity (r(2)>0.99) at all temperatures studied (15-30°C) and compared with the other two fit methods. The linear van't Hoff (15-30°C) plot obtained indicated that the thermodynamic parameters of complexation were temperature dependent for the enantiomers.
    Matched MeSH terms: Electrophoresis, Capillary/methods*
  11. 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: Electrophoresis, Gel, Two-Dimensional
  12. Vellasamy KM, Mariappan V, Hashim OH, Vadivelu J
    Electrophoresis, 2011 Jan;32(2):310-20.
    PMID: 21254130 DOI: 10.1002/elps.201000355
    Bacterial secreted proteins are known to be involved in virulence and may mediate important host-pathogen interactions. In this study, when the stationary phase culture supernatant of Burkholderia pseudomallei was subjected to 2-DE, 113 protein spots were detected. Fifty-four of the secreted proteins, which included metabolic enzymes, transcription/translation regulators, potential virulence factors, chaperones, transport regulators, and hypothetical proteins, were identified using MS and database search. Twelve of these proteins were apparently reactive to antisera of mice that were immunised with B. pseudomallei secreted proteins. These proteins might be excellent candidates to be used as diagnostic markers or putative candidate vaccines against B. pseudomallei infections.
  13. Nolan MJ, Jex AR, Upcroft JA, Upcroft P, Gasser RB
    Electrophoresis, 2011 Aug;32(16):2075-90.
    PMID: 23479788
    We barcoded 25 in vitro isolates (representing 92 samples) of Giardia duodenalis from humans and other animals, which have been assembled by the Upcroft team at the Queensland Institute of Medical Research over a period of almost three decades. We used mutation scanning-coupled sequencing of loci in the triosephosphate isomerase, glutamate dehydrogenase and β-giardin genes, combined with phylogenetic analysis, to genetically characterise them. Specifically, the isolates (n514) of G. duodenalis from humans from Australia (AD113; BRIS/83/HEPU/106; BRIS/87/HEPU/713; BRIS/89/HEPU/1003; BRIS/92/HEPU/1541; BRIS/92/HEPU/1590; BRIS/92/HEPU/2443; BRIS/93/HEPU/1706), Malaysia (KL/92/IMR/1106) and Afghanistan (WB), a cat from Australia (BAC2), a sheep from Canada (OAS1) and a sulphur-crested cockatoo from Australia (BRIS/95/HEPU/2041) represented assemblage A (sub-assemblage AI-1, AI-2 or AII-2); isolates (n510) from humans from Australia (BRIS/91/HEPU/1279; BRIS/92/HEPU/2342; BRIS/92/HEPU/2348; BRIS/93/HEPU/1638; BRIS/93/HEPU/1653; BRIS/93/HEPU/1705; BRIS/93/HEPU/1718; BRIS/93/HEPU/1727), Papua New Guinea (BRIS/92/HEPU/1487) and Canada (H7) represented assemblage B (sub-assemblage BIV) and an isolate from cattle from Australia (BRIS/92/HEPU/1709) had a match to assemblage E. Isolate BRIS/90/HEPU/1229 from a human from Australia was shown to represent a mixed population of assemblages A and B. These barcoded isolates (including stocks and derived lines) now allow direct comparisons of experimental data among laboratories and represent a massive resource for transcriptomic, proteomic, metabolic and functional genomic studies using advanced molecular technologies.
  14. Jayapalan JJ, Ng KL, Razack AH, Hashim OH
    Electrophoresis, 2012 Jul;33(12):1855-62.
    PMID: 22740474 DOI: 10.1002/elps.201100608
    Diagnosis of prostate cancer (PCa) is currently much reliant on the invasive and time-consuming transrectal ultrasound-guided biopsy of the prostate gland, particularly in light of the inefficient use of prostate-specific antigen as its biomarker. In the present study, we have profiled the sera of patients with PCa and benign prostatic hyperplasia (BPH) using the gel- and lectin-based proteomics methods and demonstrated the significant differential expression of apolipoprotein AII, complement C3 beta chain fragment, inter-alpha-trypsin inhibitor heavy chain 4 fragment, transthyretin, alpha-1-antitrypsin, and high molecular weight kininogen (light chain) between the two groups of patients' samples. Our data are suggestive of the potential use of the serum proteins as complementary biomarkers to effectively discriminate PCa from BPH, although this requires further extensive validation on clinically representative populations.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
  15. Yoon KY, Tan WS, Tey BT, Lee KW, Ho KL
    Electrophoresis, 2013 Jan;34(2):244-53.
    PMID: 23161478 DOI: 10.1002/elps.201200257
    Hepatitis B core antigen (HBcAg) expressed in Escherichia coli is able to self-assemble into large and small capsids comprising 240 (triangulation number T = 4) and 180 (triangulation number T = 3) subunits, respectively. Conventionally, sucrose density gradient ultracentrifugation and SEC have been used to separate these capsids. However, good separation of the large and small particles with these methods is never achieved. In the present study, we employed a simple, fast, and cost-effective method to separate the T = 3 and T = 4 HBcAg capsids by using native agarose gel electrophoresis followed by an electroelution method (NAGE-EE). This is a direct, fast, and economic method for isolating the large and small HBcAg particles homogenously based on the hydrodynamic radius of the spherical particles. Dynamic light scattering analysis demonstrated that the T = 3 and T = 4 HBcAg capsids prepared using the NAGE-EE method are monodisperse with polydispersity values of ∼15% and ∼13%, respectively. ELISA proved that the antigenicity of the capsids was not affected in the purification process. Overall, NAGE-EE produced T = 3 and T = 4 capsids with a purity above 90%, and the recovery was 34% and 50%, respectively (total recovery of HBcAg is ∼84%), and the operation time is 15 and 4 times lesser than that of the sucrose density gradient ultracentrifugation and SEC, respectively.
    Matched MeSH terms: Electrophoresis, Agar Gel/methods*
  16. Liew YK, Neela V, Hamat RA, Nordin SA, Chong PP
    Electrophoresis, 2013 Feb;34(3):397-400.
    PMID: 23161123 DOI: 10.1002/elps.201200380
    The typical concentration of protein loaded varies from 0.13 to 1.40 μg/μL for a classical silver staining method in 2DE gel. Here, we present a simple modified classical silver staining method by modifying the silver impregnation and development reaction steps. This modified method detects the protein spots at extremely low loaded concentrations, ranging from 0.0048 to 0.0480 μg/μL. We recommend this modified silver staining as an excellent method for the limited biological samples used for silver-stained 2DE analysis. Altogether, the protocol takes close to two days from first dimension separation to second dimension separation, followed by silver staining, scanning, and analysis.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional/methods*
  17. Yunus NA, Nili H, Green NG
    Electrophoresis, 2013 Apr;34(7):969-78.
    PMID: 23436439 DOI: 10.1002/elps.201200466
    Dielectrophoresis is the movement of particles in nonuniform electric fields and has been of interest for application to manipulation and separation at and below the microscale. This technique has the advantages of being noninvasive, nondestructive, and noncontact, with the movement of particle achieved by means of electric fields generated by miniaturized electrodes and microfluidic systems. Although the majority of applications have been above the microscale, there is increasing interest in application to colloidal particles around a micron and smaller. This paper begins with a review of colloidal and nanoscale dielectrophoresis with specific attention paid to separation applications. An innovative design of integrated microelectrode array and its application to flow-through, continuous separation of colloidal particles is then presented. The details of the angled chevron microelectrode array and the test microfluidic system are then discussed. The variation in device operation with applied signal voltage is presented and discussed in terms of separation efficiency, demonstrating 99.9% separation of a mixture of colloidal latex spheres.
    Matched MeSH terms: Electrophoresis/instrumentation*; Electrophoresis/methods*
  18. Jayapalan JJ, Ng KL, Shuib AS, Razack AH, Hashim OH
    Electrophoresis, 2013 Jun;34(11):1663-9.
    PMID: 23417432 DOI: 10.1002/elps.201200583
    The present study was aimed at the identification of proteins that are differentially expressed in the urine of patients with prostate cancer (PCa), those with benign prostatic hyperplasia (BPH) and age-matched healthy male control subjects. Using a combination of 2DE and MS/MS, significantly lower expression of urinary saposin B and two different fragments of inter-alpha-trypsin inhibitor light chain (ITIL) was demonstrated in the PCa patients compared to the controls. However, only one of the ITIL fragments was significantly different between the PCa and BPH patients. When image analysis was performed on urinary proteins that were transferred onto NC membranes and detected using a lectin that binds to O-glycans, a truncated fragment of inter-alpha-trypsin inhibitor heavy chain 4 was the sole protein found to be significantly enhanced in the PCa patients compared to the controls. Together, these urinary peptide fragments might be useful complementary biomarkers to indicate PCa as well as to distinguish it from BPH, although further epidemiological evidence on the specificity and sensitivity of the protein candidates is required.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
  19. Chang HY, Hor SY, Lim KP, Zain RB, Cheong SC, Rahman MA, et al.
    Electrophoresis, 2013 Aug;34(15):2199-208.
    PMID: 23712713 DOI: 10.1002/elps.201300126
    This study aims to identify cancer-associated proteins in the secretome of oral cancer cell lines. We have successfully established four primary cell cultures of normal cells with a limited lifespan without human telomerase reverse transcriptase (hTERT) immortalization. The secretome of these primary cell cultures were compared with that of oral cancer cell lines using 2DE. Thirty five protein spots were found to have changed in abundance. Unambiguous identification of these proteins was achieved by MALDI TOF/TOF. In silico analysis predicted that 24 of these proteins were secreted via classical or nonclassical mechanisms. The mRNA expression of six genes was found to correlate with the corresponding protein abundance. Ingenuity Pathway Analysis (IPA) core analysis revealed that the identified proteins were relevant in, and related to, cancer development with likely involvements in tumor growth, metastasis, hyperproliferation, tumorigenesis, neoplasia, hyperplasia, and cell transformation. In conclusion, we have demonstrated that a comparative study of the secretome of cancer versus normal cell lines can be used to identify cancer-associated proteins.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
  20. Aan GJ, Hairi HA, Makpol S, Rahman MA, Karsani SA
    Electrophoresis, 2013 Aug;34(15):2209-17.
    PMID: 23712505 DOI: 10.1002/elps.201300086
    Replicative senescence and stress-induced premature senescence (SIPS) cells are known to share certain traits. However, whether these cells are different at the protein level is unclear. Thus, this study has utilized proteomics to identify differences in the proteomes of replicative senescence and SIPS cells compared to normal cells. Replicative senescence was induced by serial passage of normal cells in culture. SIPS was established by exposure to H2 O2 at a subcytotoxic concentration of 20 μM for two weeks. Following 2DE, protein profiles were compared and protein spots that changed in abundance were identified by MALDI-TOF MS. Quantitative real-time RT-PCR was then performed to evaluate the transcript expression of selected altered proteins. A total of 24 and 10 proteins were found to have changed in abundance in replicative senescence and SIPS cells, respectively, when compared to young cells. Quantitative RT-PCR revealed that nine genes showed the same direction of change as observed in the proteomics analysis. Very little overlap was observed between proteins that changed in replicative senescence and SIPS cells, suggesting that although both SIPS and replicative senescence cells share hallmarks of cellular senescence, they were different in terms of proteins that changed in abundance.
    Matched MeSH terms: Electrophoresis, Gel, Two-Dimensional
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