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  1. Fulltext Methods for Systematic Identification of Membrane Proteins for Specific Capture of Cancer-Derived Extracellular Vesicles
    Zaborowski MP, Lee K, Na YJ, Sammarco A, Zhang X, Iwanicki M, et al.
    Cell Rep, 2019 Apr 02;27(1):255-268.e6.
    PMID: 30943406 DOI: 10.1016/j.celrep.2019.03.003
    Analysis of cancer-derived extracellular vesicles (EVs) in biofluids potentially provides a source of disease biomarkers. At present there is no procedure to systematically identify which antigens should be targeted to differentiate cancer-derived from normal host cell-derived EVs. Here, we propose a computational framework that integrates information about membrane proteins in tumors and normal tissues from databases: UniProt, The Cancer Genome Atlas, the Genotype-Tissue Expression Project, and the Human Protein Atlas. We developed two methods to assess capture of EVs from specific cell types. (1) We used palmitoylated fluorescent protein (palmtdTomato) to label tumor-derived EVs. Beads displaying antibodies of interest were incubated with conditioned medium from palmtdTomato-expressing cells. Bound EVs were quantified using flow cytometry. (2) We also showed that membrane-bound Gaussia luciferase allows the detection of cancer-derived EVs in blood of tumor-bearing animals. Our analytical and validation platform should be applicable to identify antigens on EVs from any tumor type.
    Matched MeSH terms: Immunoassay/methods
  2. Fulltext Evaluation of the Elecsys(®) Anti-HCV II assay for routine hepatitis C virus screening of different Asian Pacific populations and detection of early infection
    Yoo SJ, Wang LL, Ning HC, Tao CM, Hirankarn N, Kuakarn S, et al.
    J Clin Virol, 2015 Mar;64:20-7.
    PMID: 25728074 DOI: 10.1016/j.jcv.2014.12.015
    Early diagnosis of hepatitis C virus (HCV) infection is essential to allow appropriate treatment and prevent transmission.
    Matched MeSH terms: Immunoassay/methods*
  3. Hev b 5 and Hev b 13 as allergen markers to estimate the allergenic potency of latex gloves
    Yeang HY, Arif SA, Raulf-Heimsoth M, Loke YH, Sander I, Sulong SH, et al.
    J Allergy Clin Immunol, 2004 Sep;114(3):593-8.
    PMID: 15356563 DOI: 10.1016/j.jaci.2004.05.039
    BACKGROUND:
    Sensitization to natural rubber latex has been linked to proteins from medical latex gloves. Various assays to estimate the amount of residual allergenic proteins extractable from latex gloves to assess their potential exposure hazard have inherent weaknesses.

    OBJECTIVE:
    This investigation was aimed at developing 2-site immunoenzymetric assays and identifying appropriate protein markers to assess the allergenic potential of latex gloves.

    METHODS:
    The presence of 6 latex allergens--Hev b 1, 2, 3, 5, 6, and 13--was measured in a cross-section of commercial latex medical gloves by using monoclonal and polyclonal antibody-based 2-site immunoenzymetric assays. The overall allergenic potential of these gloves was assessed by IgE-inhibition assay. Stepwise multiple regression analyses were performed to identify marker allergens that best explained the variation in latex glove allergenicity.

    RESULTS:
    All 6 latex allergens were detected in at least some of the glove samples. Hev b 5 and Hev b 13 were identified as the marker allergens that combined best to explain the variation in the glove allergenicity. The significant multiple correlation (R=0.855) between these 2 markers and glove allergenic potency forms the basis of an assay to gauge latex glove allergenicity.

    CONCLUSION:
    The overall allergenic potential of latex gloves can be estimated by using Hev b 5 and Hev b 13 as indicator allergens. The correlation between glove allergenicity and the level of these allergens was maintained for low-protein gloves (<200 microg/g). This estimation of glove allergenicity was superior to that obtained by using total protein readings.
    Matched MeSH terms: Immunoassay/methods
  4. Achieving enhanced sensitivity and accuracy in carcinoembryonic antigen (CEA) detection as an indicator of cancer monitoring using thionine/chitosan/graphene oxide nanocomposite-modified electrochemical immunosensor
    Yang H, Zhang Z, Zhou X, Binbr Abe Menen N, Rouhi O
    Environ Res, 2023 Dec 01;238(Pt 1):117163.
    PMID: 37722583 DOI: 10.1016/j.envres.2023.117163
    The current study has focused on electrochemical immunosensing of carcinoembryonic antigen (CEA) employing an immobilized antibody on a thionine, chitosan, or graphene oxide nanocomposite modified glassy carbon electrode (anti-CEA/THi-CS-GO/GCE) as an indicator of cancer monitoring. THi-CS-GO nanocomposites were made using ultrasonication, and analyses of their morphology and crystal structure using SEM, FTIR, and XRD showed that thionine and chitosan molecules were intercalated with stacking interactions with both the top and bottom of GO nanosheets. Electrochemical experiments revealed anti-CEA, THi-CS-GO/GCE to have exceptional sensitivity and selectivity towards CEA compounds. The detection limit value was established to be 0.8 pg/mL when it was discovered that variations in the decrease peak current were directly proportional to the logarithm concentration of CEA over a wide range from 10-3 to 104 ng/mL. Results of testing the immunosensor's application capability for detecting CEA in a sample of human serum show that ELISA and DPV results are very congruent. The produced immunosensor demonstrated adequate immunosensor precision in determining CEA in prepared genuine samples of human serum and clinical applications.
    Matched MeSH terms: Immunoassay/methods
  5. Heavy/light chain assay in the monitoring of multiple myeloma
    Ting HY, Sthaneshwar P, Bee PC, Shanmugam H, Lim M
    Pathology, 2019 Aug;51(5):507-511.
    PMID: 31253381 DOI: 10.1016/j.pathol.2019.04.002
    Serum protein (SPE) and immunofixation electrophoresis (IFE) have been extensively validated for the routine use of identifying, characterising and quantifying monoclonal proteins. However, accurate quantitation of IgA monoclonal proteins can be difficult when they migrate in to the β fraction, due to co-migration with transferrin and complement components. The heavy/light chain (HLC) immunoassay is an additional tool for measuring intact immunoglobulin monoclonal proteins. Therefore, we aimed to examine the clinical utility of the HLC assay for the disease monitoring of IgG and IgA multiple myeloma (MM) patients. A total of 177 samples from 30 MM patients (21 IgG and 9 IgA) were analysed retrospectively with median number of six follow up samples per patient (range 3-13). Serum free light chains (sFLC) and HLC were quantified using Freelite and Hevylite immunoassays. Details of M-protein concentration, β-globulin levels, total immunoglobulin levels and disease treatment response were obtained from the laboratory and patient information system. Passing-Bablok regression analysis was performed to compare (i) M-protein quantification with involved HLC (iHLC) and (ii) total immunoglobulin with summated HLC pairs for each immunoglobulin type (e.g., IgGκ+IgGλ). For 127 IgG MM samples, IgG iHLC levels showed a good correlation with SPE quantification (iHLC y=0.96x+4.9; r=0.917) and summated HLC showed a good correlation with total IgG concentration (summated HLC y=0.94x+5.74; r=0.91). In total, 95/127 (75%) IgG MM follow-up samples had an abnormal HLC ratio and 122/127 (96%) had a positive SPE, probably due to the lower sensitivity of HLC assay in detecting clonality in patients with IgG MM. Consistent with this, one patient assigned a very good partial response by International Myeloma Working Group criteria would be assigned a complete response based on HLC measurements. For 50 IgA MM samples, 42/50 (84%) had an abnormal HLC ratio. Conversely, 50/50 (100%) of M-proteins showed β fraction migration and were difficult to accurately quantify by SPE. Therefore, M-protein concentration and iHLC did not correlate as well in IgA MM (y=1.9x-8.4; r=0.8) compared to IgG MM. However, there was good correlation between total IgA and summated IgA HLC (IgAκ+IgAλ y=1.35x-0.33; r=0.95). Of the 8/50 (16%) IgA samples with a normal HLC ratio, 6/8 (75%) were consistent with the disease status being in complete remission. Interestingly, in one IgA MM patient, SPE and IFE were negative, but the serum FLC ratio and involved FLC were highly abnormal, consistent with the presence of light chain escape. Our data suggest HLC measurements could add value to the current disease monitoring of MM patients. In IgG MM patients, the M-protein level correlated well with HLC values. The HLC assay complements the serum FLC assay and is especially useful for monitoring of IgA MM patients who display M-proteins migrating in the β region on SPE.
    Matched MeSH terms: Immunoassay/methods*
  6. Fulltext Production and characterization of a polyclonal antibody of anti-rLipL21-IgG against leptospira for early detection of acute leptospirosis
    Seenichamy A, Bahaman AR, Mutalib AR, Khairani-Bejo S
    Biomed Res Int, 2014;2014:592858.
    PMID: 24860824 DOI: 10.1155/2014/592858
    Leptospirosis is one of the zoonotic diseases in animals and humans throughout the world. LipL21 is one of the important surface-exposed lipoproteins in leptospires and the most effective cross protective immunogenic antigen. It is widely considered as a diagnostic marker for leptospirosis. In this study, we evaluated the serodiagnostic potential of LipL21 protein of Leptospira interrogans serovar Pomona. We have successfully amplified, cloned, and expressed LipL21 in E. coli and evaluated its specificity by immunoblotting. Purified recombinant LipL21 (rLipL21) was inoculated into rabbits for the production of polyclonal antibody. Characterization of the purified IgG antibody against rLipL21 was performed by cross reactivity assay. Only sera from leptospirosis patients and rabbit hyperimmune sera recognized rLipL21 while the nonleptospirosis control sera showed no reaction in immunoblotting. We confirmed that anti-rLipL21-IgG antibody cross reacted with and detected only pathogenic leptospiral species and it did not react with nonpathogenic leptospires and other bacterial species. Results observed showed that anti-rLipL21-IgG antibody has high specificity and sensitivity to leptospires. The findings indicated that the antibody could be used in a diagnostic assay for detection of leptospires or their proteins in the early phase of infection.
    Matched MeSH terms: Immunoassay/methods
  7. Prevalence of Bacillus cereus in selected foods and detection of enterotoxin using TECRA-VIA and BCET-RPLA
    Rusul G, Yaacob NH
    Int J Food Microbiol, 1995 Apr;25(2):131-9.
    PMID: 7547144
    Enterotoxigenic Bacillus cereus was detected in cooked foods (17), rice noodles (3), wet wheat noodles (2), dry wheat noodles (10), spices (8), grains (4), legumes (11) and legume products (3). One hundred ninety-four (42.3%), 70 (15.3%) and 23 (5.2%) of the 459 presumptive B. cereus colonies isolated from PEMBA agar were identified as B. cereus, Bacillus thuringiensis and B. mycoides, respectively. B. cereus isolates were examined for growth temperature, pH profile and enterotoxin production using both TECRA-VIA and BCET-RPLA kits. One hundred seventy-eight (91.8%) and 164 (84%) of the strains were enterotoxigenic as determined using TECRA-VIA and BCET-RPLA, respectively. Eighty-two (50%) of the enterotoxigenic strains were capable of growing at 5 degrees C, and 142 (86.6%) grew at 7 degrees C within 7 days of incubation. The enterotoxigenic strains did not grow at pH 4.0 but 69 (42.0%) of the strains were able to grow at pH 4.5 within 7 days at 37 degrees C. The isolates were resistant to ampicillin (98.8%), cloxallin (100%) and tetracycline (61.0%), and susceptible to chloroamphenicol (87%), erythromycin (77.4%), gentamycin (100%) and streptomycin (98.7%).
    Matched MeSH terms: Immunoassay/methods*
  8. Distinct prevalence of antibodies to the E2 protein of GB virus C/hepatitis G virus in different parts of the world
    Ross RS, Viazov S, Schmitt U, Schmolke S, Tacke M, Ofenloch-Haehnle B, et al.
    J Med Virol, 1998 Feb;54(2):103-6.
    PMID: 9496367
    Since the identification of the new human virus, GB virus C (GBV-C)/hepatitis G-virus (HGV), in 1995/1996, reverse transcription polymerase chain reaction remained the sole available diagnostic tool for GBV-C/HGV infection. Recently, a serologic test based on the detection of antibodies to the putative envelope protein 2 (anti-E2) has been introduced. We used this assay for a seroepidemiological survey including 3,314 healthy individuals from different parts of the world, 123 patients from Germany who were suspected to have an increased risk of acquiring GBV-C/HGV infection, 128 multiple organ donors, and 90 GBV-C/HGV RNA positive persons. In European countries, anti-E2 seropositivity ranged from 10.9% (Germany) to 15.3% (Austria). In South Africa (20.3%) and Brazil (19.5%), even higher anti-E2 prevalence rates were recorded. In Asian countries like Bhutan (3.9%), Malaysia (6.3%), and the Philippines (2.7%), anti-E2 positivity was significantly lower. GBV-C/HGV anti-E2 prevalence in potential "risk groups," i.e., patients on hemodialysis and renal transplant recipients, did not vary significantly from anti-E2 seroprevalence in German blood donors. Anti-E2 and GBV-C/HGV RNA were found to be mutually exclusive, confirming the notion that anti-E2 has to be considered as a marker of past infection.
    Matched MeSH terms: Immunoassay/methods
  9. Evaluation of the cholera spot test: a chromatographic immunoassay for the rapid detection of cholera antigen
    Rohani MY, Hasnidah D, Ong KH
    Malays J Pathol, 1998 Jun;20(1):31-3.
    PMID: 10879261
    A chromatographic immunoassay cholera antigen detection kit, the Cholera Spot test, was evaluated. The test was found to be specific with a sensitivity of 10(6) cfu/ml for the direct detection of V. cholerae in simulated stool specimens and 10 cfu/ml in simulated cotton-tipped swab specimens after overnight incubation in alkaline peptone water. This enables early recognition of cholera cases and their contacts so that prevention and control measures can be promptly instituted.
    Matched MeSH terms: Immunoassay/methods
  10. Specificity and sensitivity of a rapid dipstick test (Brugia Rapid) in the detection of Brugia malayi infection
    Rahmah N, Taniawati S, Shenoy RK, Lim BH, Kumaraswami V, Anuar AK, et al.
    Trans R Soc Trop Med Hyg, 2002 1 31;95(6):601-4.
    PMID: 11816429
    A total of 753 serum samples from 6 institutions in 3 countries (Malaysia, Indonesia and India) were used to evaluate an immunochromatographic rapid dipstick test, Brugia Rapid, for diagnosis of Brugia malayi infection. The samples comprised sera from 207 microfilaria-positive individuals and 546 individuals from filaria non-endemic areas. The latter consisted of 70 individuals with soil-transmitted helminth infections, 68 with other helminth infections, 238 with protozoan infections, 12 with bacterial and viral infections and 158 healthy individuals. The dipstick is prepared with a goat anti-mouse antibody control line and a B. malayi recombinant-antigen test line. First, the dipstick is dipped into a well containing diluted patient serum, thus allowing specific anti-filarial antibody in the serum to react with the recombinant antigen. Then the dipstick is placed into an adjacent well containing reconstituted anti-human IgG4-gold. After 10 min, development of 2 red-purplish lines denotes a positive result and one line indicates a negative reaction. The overall results of the evaluation showed 97% sensitivity, 99% specificity, 97% positive predictive value and 99% negative predictive value. Brugia Rapid is thus a promising diagnostic tool for detection of B. malayi infection, and would be especially useful for the brugian filariasis elimination programme.
    Matched MeSH terms: Immunoassay/methods
  11. A Brugia malayi antigen specifically recognized by infected individuals
    Rahmah N, Anuar AK, A'shikin AN, Lim BH, Mehdi R, Abdullah B, et al.
    Biochem Biophys Res Commun, 1998 Sep 29;250(3):586-8.
    PMID: 9784388
    Western blot analyses were performed on 444 serum specimens: 40 sera from microfilaraemic individuals, 10 sera from elephantiasis patients, 24 treated individuals, 50 sera from residents of endemic areas without anti-filarial IgG4 antibodies (endemic normals), 20 sera from amicrofilaraemic individuals with high anti-filarial IgG4 antibodies, 200 sera from healthy city-dwellers (non-endemic samples), and 100 sera from soil-transmitted helminth-infected individuals. Phast electrophoresis system was used to electrophorese Brugia malayi soluble adult worm antigen on 10-15% SDS-PAGE gradient gels followed by electrophoretic transfer onto PVDF membranes. Membrane strips were then successively incubated with blocking solution, human sera, and monoclonal anti-human IgG4 antibody-HRP, with adequate washings done in between each incubation step. Luminol chemiluminescence detection was then used to develop the blots. An antigenic band with the MW of approximately 37 kDa was found to be consistently present in the Western blots of all microfilaraemic sera, all amicrofilaraemic sera with high titres of anti-filarial IgG4 antibodies, some treated patients, and some elephantiasis patients. The antigen did not occur in immunoblots of individuals with other helminthic infections, normal endemic individuals, and city dwellers. Therefore the B. malayi antigen of with the MW of approximately 37 kDa demonstrated specific reactions with sera of B. malayi-infected individuals and thus may be useful for diagnostic application.
    Matched MeSH terms: Immunoassay/methods
  12. Fulltext Sensitivity and specificity of malaria rapid diagnostic test (mRDT CareStatTM) compared with microscopy amongst under five children attending a primary care clinic in southern Nigeria
    Ogunfowokan O, Ogunfowokan BA, Nwajei AI
    Afr J Prim Health Care Fam Med, 2020 Jun 17;12(1):e1-e8.
    PMID: 32634015 DOI: 10.4102/phcfm.v12i1.2212
    BACKGROUND: Malaria diagnosis using microscopy is currently the gold standard. However, malaria rapid diagnostic tests (mRDTs) were developed to simplify the diagnosis in regions without access to functional microscopy.

    AIM: The objective of this study was to compare the diagnostic accuracy of mRDT CareStatTM with microscopy.

    SETTING: This study was conducted in the paediatric primary care clinic of the Federal Medical Centre, Asaba, Nigeria.

    METHODS: A cross-sectional study for diagnostic accuracy was conducted from May 2016 to October 2016. Ninety-eight participants were involved to obtain a precision of 5%, sensitivity of mRDT CareStatTM of 95% from published work and 95% level of confidence after adjusting for 20% non-response rate or missing data. Consecutive participants were tested using both microscopy and mRDT. The results were analysed using EPI Info Version 7.

    RESULTS: A total of 98 children aged 3-59 months were enrolled. Malaria prevalence was found to be 53% (95% confidence interval [CI] = 46% - 60%), whilst sensitivity and specificity were 29% (95% CI = 20% - 38%) and 89% (95% CI = 83% - 95%), respectively. The positive and negative predictive values were 75% (95% CI = 66.4% - 83.6%) and 53% (95% CI = 46% - 60%), respectively.

    CONCLUSION: Agreement between malaria parasitaemia using microscopy and mRDT positivity increased with increase in the parasite density. The mRDT might be negative when malaria parasite density using microscopy is low.

    Matched MeSH terms: Immunoassay/methods
  13. Fulltext Comparison between AmniSure placental alpha microglobulin-1 rapid immunoassay and standard diagnostic methods for detection of rupture of membranes
    Ng BK, Lim PS, Shafiee MN, Ghani NA, Ismail NA, Omar MH, et al.
    Biomed Res Int, 2013;2013:587438.
    PMID: 24073412 DOI: 10.1155/2013/587438
    Objective. To determine the diagnostic accuracy of placental alpha microglobulin-1 assay and standard diagnostic methods for detecting rupture of membrane. Study Design. Prospective diagnostic study, between June 2011 to November 2011 at a tertiary centre. Initial evaluation included both the standard diagnostic methods for rupture of membranes and placental alpha microglobulin-1 immunoassay. The actual rupture of membranes was diagnosed on review of the medical records after delivery (absence of membrane or a positive pad chart). Main Outcome Measures. Placental alpha microglobulin-1 immunoassay and standard diagnostic methods for diagnosis of rupture of membrane. Results. A total of 211 patients were recruited. At initial presentation, 187 patients (88.6%) had ruptured membranes, while 24 patients (11.4%) had intact membranes. Placental alpha microglobulin-1 immunoassay confirmed rupture of membranes at initial presentation with a sensitivity of 95.7% (179 of 187), specificity of 100% (24 of 24), positive predictive value of 100% (179 of 179), and negative predictive value of 75.0% (24 of 32). By comparison, the conventional standard diagnostic methods had a sensitivity of 78.1% (146 of 187), specificity of 100% (24 of 24), positive predictive value of 100% (146 of 146), and negative predictive value of 36.9% (24 of 65) in diagnosing rupture of membrane. Conclusion. Placental alpha-microglobulin-1 immunoassay is a rapid and accurate method for confirming the diagnosis of rupture of membrane. It was superior to conventional standard diagnostic methods (pooling, nitrazine, and ferning), the nitrazine test alone or fern test alone.
    Matched MeSH terms: Immunoassay/methods*
  14. Performance of selected SARS-CoV-2 antibodies immunoassays
    Mohd Zain R, Shaiful Bahrin NI, Ellan K, Thayan R
    Malays J Pathol, 2021 Dec;43(3):463-464.
    PMID: 34958069
    No abstract available.
    Matched MeSH terms: Immunoassay/methods*
  15. Portable electrochemical immunosensor for detection of Mycobacterium tuberculosis secreted protein CFP10-ESAT6 in clinical sputum samples
    Mohd Azmi UZ, Yusof NA, Abdullah J, Alang Ahmad SA, Mohd Faudzi FN, Ahmad Raston NH, et al.
    Mikrochim Acta, 2021 01 06;188(1):20.
    PMID: 33404779 DOI: 10.1007/s00604-020-04669-x
    An early detection of Mycobacterium tuberculosis is very important to reduce the number of fatal cases and allow for fast recovery. However, the interpretation of the result from smear microscopy requires skilled personnel due to the propensity of the method to produce false-negative results. In this work, a portable, rapid, and simple sandwich-type immunosensor reader has been developed that is able to detect the presence of M. tuberculosis in sputum samples. By using sandwich-type immunosensor, an anti-CFP10-ESAT6 antibody was immobilized onto the graphene/polyaniline (GP/PANI)-modified gold screen-printed electrode. After incubation with the target CFP10-ESAT6 antigen, the iron/gold magnetic nanoparticles (Fe3O4/Au MNPs) conjugated with anti-CFP10-ESAT6 antibody were used to complete the sandwich format. Differential pulse voltammetry (DPV) technique was used to detect the CFP10-ESAT6 antigen at the potential range of 0.0-1.0 V. The detection time is less than 2 h. Under optimal condition, CFP10-ESAT6 antigen was detected in a linear range from 10 to 500 ng mL-1 with a limit of detection at 1.5 ng mL-1. The method developed from this process was then integrated into a portable reader. The performance of the sensor was investigated and compared with the standard methods (culture and smear microscopy). It provides a good correlation (100% sensitivity and 91.7% specificity) with both methods of detection for M. tuberculosis in sputum samples henceforth, demonstrating the potential of the device as a more practical screening tool.Graphical abstract.
    Matched MeSH terms: Immunoassay/methods
  16. Development of monoclonal antibodies against recombinant LipL21 protein of pathogenic Leptospira through phage display technology
    Mohd Ali MR, Sum JS, Aminuddin Baki NN, Choong YS, Nor Amdan NA, Amran F, et al.
    Int J Biol Macromol, 2021 Jan 31;168:289-300.
    PMID: 33310091 DOI: 10.1016/j.ijbiomac.2020.12.062
    Leptospirosis is a potentially fatal zoonosis that is caused by spirochete Leptospira. The signs and symptoms of leptospirosis are usually varied, allowing it to be mistaken for other causes of acute febrile syndromes. Thus, early diagnosis and identification of a specific agent in clinical samples is crucial for effective treatment. This study was aimed to develop specific monoclonal antibodies against LipL21 antigen for future use in leptospirosis rapid and accurate immunoassay. A recombinant LipL21 (rLipL21) antigen was optimized for expression and evaluated for immunogenicity. Then, a naïve phage antibody library was utilized to identify single chain fragment variable (scFv) clones against the rLipL21 antigen. A total of 47 clones were analysed through monoclonal phage ELISA. However, after taking into consideration the background OD405 values, only 4 clones were sent for sequencing to determine human germline sequences. The sequence analysis showed that all 4 clones are identical. The in silico analysis of scFv-lip-1 complex indicated that the charged residues of scFv CDRs are responsible for the recognition with rLipL21 epitopes. The generated monoclonal antibody against rLipL21 will be evaluated as a detection reagent for the diagnosis of human leptospirosis in a future study.
    Matched MeSH terms: Immunoassay/methods
  17. Fulltext IgG4 detection of Echinococcus granulosus paramyosin is a useful diagnostic test for human hydatidosis
    Moghadam ZK, Ghaffarifar F, Khalilpour A, Abdul Aziz F, Saadatnia G, Noordin R
    Clin Vaccine Immunol, 2013 Apr;20(4):501-5.
    PMID: 23365208 DOI: 10.1128/CVI.00019-13
    Hydatidosis is a public health problem in many parts of the world, and improvement in diagnosis of the disease is still being pursued. Protoscoleces of Echinococcus granulosus were isolated from hydatid cysts collected from naturally infected sheep slaughtered in abattoirs in Iran. Sonicated extract of protoscolex was subjected to two-dimensional gel electrophoresis and Western blot analysis. Primary antibodies were from serum samples from 130 hydatidosis patients, 38 individuals infected with other parasitic infections, and 30 healthy people, whereas peroxidase (HRP)-conjugated anti-human IgG and IgG4 were used as secondary antibodies. The recombinant form of the identified protein was produced and tested for its sensitivity and specificity for the detection of human hydatidosis. An antigenic band of ∼60 kDa was found to be sensitive (82%) and specific (100%) for the detection of hydatidosis when probed with anti-human IgG4-HRP, while the sensitivity and specificity were 33 and 100%, respectively, with anti-human IgG-HRP. By mass spectrometry, the band was identified as protoscolex tegument paramyosin. The sensitivity and specificity of full-length paramyosin-recombinant protein in IgG4 blots were found to be 86 and 98%, respectively. In conclusion, IgG4 detection of Echinococcus granulosus paramyosin was found to be useful for the diagnosis of human hydatidosis.
    Matched MeSH terms: Immunoassay/methods
  18. Can fluoride-oxalate and sodium citrate stabilise homocysteine levels after blood collection?
    Lopez JB, Peng CL
    Clin Chem Lab Med, 2003 Oct;41(10):1369-72.
    PMID: 14580168
    The concentration of homocysteine (Hcy) rises rapidly after the collection of blood. This feature requires blood to be collected into the anticoagulants EDTA or heparin and the plasma to then be immediately separated; alternatively, the blood may be kept on ice and centrifuged within 1 hour. The use of chemical preservatives has been proposed as a means of stabilising Hcy levels in whole blood after collection. The objective of this study was to determine whether the commonly available fluoride-oxalate (Fl-Ox) and sodium citrate (Na-Cit) containers could stabilise Hcy levels in blood. Our results showed that when blood was collected into potassium EDTA (K-EDTA) tubes, Hcy levels rose from initial levels, on standing at room temperature (approximately 25 degrees C), by an average of 21% after 3 hours and 32% after 5 hours. The initial Hcy levels of blood collected into Fl-Ox and Na-Cit containers, however, were lower, at averages of 89% and 91%, respectively, compared to that of the same samples when collected into K-EDTA tubes. Hcy in these samples subsequently rose on standing, and after 5 hours was, on the average, 10 and 13% higher, respectively, compared with the initial levels in K-EDTA tubes. We conclude that Fl-Ox and Na-Cit do not stabilise Hcy in blood after collection and should not be used as preservatives.
    Matched MeSH terms: Immunoassay/methods*
  19. IQPA: isothermal nucleic acid amplification-based immunoassay using DNAzyme as the reporter system
    Loh Q, Omar N, Glökler J, Lim TS
    Anal Biochem, 2014 Oct 15;463:67-9.
    PMID: 24972268 DOI: 10.1016/j.ab.2014.06.012
    Immunoassays are often coupled to peroxidase activity for antigen detection. Sensitivity and speed of detection has been increased by the advent of hybrid methods such as immuno-PCR (polymerase chain reaction). However, a more simplified immunoassay that retains both colorimetric peroxidase detection and effective DNA amplification in a setting closer to field application conditions has been nonexistent. Here we describe a method that successfully combines a competitive immunoassay with the new isothermal quadruplex-primed amplification (QPA) to generate excess quadruplex reporter molecules with intrinsic peroxidase DNAzyme activity.
    Matched MeSH terms: Immunoassay/methods*
  20. The specificity of a rapid method of assay for the presence of Legionella species in water
    Lim EW, Meers PD
    Ann Acad Med Singap, 1989 Jul;18(4):348-51.
    PMID: 2679337
    A rapid method of assay, using a monoclonal antibody linked to alkaline phosphatase, was used for the detection of the Pontiac subgroup of Legionella pneumophila serogroup 1. It was tested for its specificity against 53 strains of Legionella recently isolated from the environment in Singapore and Malaysia. The specificity and sensitivity of this method of assay was confirmed, though there is some concern that the specificity was too narrow, and there are reservations about the criteria suggested for interpreting the results.
    Matched MeSH terms: Immunoassay/methods
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