scholarly journals Strongyloides-Specific IgE Phage cDNA Clones and Development of a Novel ELISA for Strongyloidiasis

Diagnostics ◽  
2021 ◽  
Vol 11 (6) ◽  
pp. 985
Author(s):  
Hussain Ahmad ◽  
Norsyahida Arifin ◽  
Thomas J. Nolan ◽  
James B. Lok ◽  
Nor Suhada Anuar ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Western Blot ◽  
Rapid Test ◽  
Strongyloides Stercoralis ◽  
Specific Ige ◽  
Diagnostic Value ◽  
Epidemiological Studies ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cdna Clones ◽  
Diagnostic Potential

Strongyloidiasis, caused mainly by the nematode Strongyloides stercoralis, is prevalent worldwide and potentially fatal in immunosuppressed patients. We report on a new IgE biomarker to diagnose Strongyloides infection. Sera from two groups infected with Strongyloides served as positive samples: Group 1A, in which infection was confirmed by stool-microscopy and/or stool-polymerase chain reaction (PCR) and was seropositive by an IgG-enzyme linked immunosorbent assay (ELISA) and an IgG4 rapid test, and Group 1B in which infection was confirmed by stool-PCR but was seronegative. Negative samples (controls) comprised infections with other parasites (Group II) and healthy donors (Group III). Immunoscreenings of an S. stercoralis complementary DNA (cDNA) library were performed, and the cDNA clone with the highest diagnostic potential (clone A133) was selected for recombinant protein production and then evaluated using IgE Western blot and ELISA. The Western blot showed that the recombinant protein (rA133) was 100% reactive with Group IA (n = 10) and Group IB (n = 5), and 96% non-reactive with Groups II and III (n = 25). Subsequently, the IgE-ELISA was developed and showed 100% diagnostic sensitivity in Groups IA (n = 32) and IB (n = 11); and 99.3% specificity in Groups II and III (n = 144). In conclusion, this study has identified rA133 as a novel recombinant protein with potential diagnostic value, and that the IgE-ELISA incorporating this protein may be useful for patient diagnosis and epidemiological studies.

scholarly journals Component resolved diagnosis of egg yolk is an indispensable part of egg allergy

2021 ◽  
Vol 49 (2) ◽  
pp. 6-14
Author(s):  
Huang Lunhui ◽  
Shao Yanhong ◽  
Li Shaoshen ◽  
Bao Huijing ◽  
Liu Yunde ◽  
...  
Keyword(s):  
Operating Characteristic ◽  
Egg Yolk ◽  
Specific Ige ◽  
Diagnostic Value ◽  
Egg White ◽  
Enzyme Linked Immunosorbent Assay ◽  
Egg Allergy ◽  
Component Resolved Diagnosis ◽  
Allergen Components

Introduction and objectives: It was urgent to explain the role of egg yolk allergen sensitization to the egg allergic population and we would evaluate the diagnostic value of allergen components in whole eggs, including egg white and egg yolk.Materials and methods: Firstly, we collected 99 positive and 21 negative sera against egg allergy. Then we used modified enzyme linked immunosorbent assay (ELISA) to survey specific IgE (sIgE) to all-proven and single component in eggs, Ovomucoid (Gal d 1), Ovalbumin (Gal d 2), Ovotransferrin (Gal d 3), Lysozyme C (Gal d 4), Serum Albumin (Gal d 5), and YGP42(Gal d 6) in allergic and non-allergic populations. Last but not least, we studied the sIgE reactivities to egg allergen components by receiver operating characteristic (ROC) analysis.Results: Among egg-allergic individuals, nearly 10% were sensitized to five of six egg allergen components, and the cross-reaction frequency between two egg yolk allergens with Gal d 1 was about 30% in the groups diagnosed with egg allergy or non-allergy. The best component-combination diagnosis in egg allergy of Gal d 1+ Gal d 6 demonstrated the largest area under curve (AUC) of 0.994.Conclusions: Our results suggested that there were individual differences in allergenicity of different egg allergen components, especially in the samples negative to egg allergy diagnosed but sensitive to egg yolk components. It was indicated that component resolved diagnosis of egg yolk improved the value for egg allergy management indispensably.

SOX Antibodies in Small-Cell Lung Cancer and Lambert-Eaton Myasthenic Syndrome: Frequency and Relation With Survival

2009 ◽  
Vol 27 (26) ◽  
pp. 4260-4267 ◽  
Author(s):  
Maarten J. Titulaer ◽  
Rinse Klooster ◽  
Marko Potman ◽  
Lidia Sabater ◽  
Francesc Graus ◽  
...  
Keyword(s):  
Western Blot ◽  
High Throughput Screening ◽  
Small Cell Lung Carcinoma ◽  
Diagnostic Value ◽  
Small Cell ◽  
Enzyme Linked Immunosorbent Assay ◽  
Small Cell Lung ◽  
Serum Samples ◽  
Cell Lung Carcinoma ◽  
Myasthenic Syndrome

Purpose SOX1 antibodies are common in small-cell lung carcinoma (SCLC) with and without paraneoplastic syndrome (PNS) and can serve as serological tumor marker. Addition of other antibodies might improve its diagnostic power. We validated an enzyme-linked immunosorbent assay (ELISA) to assess the diagnostic value of serum antibodies in SCLC and Lambert-Eaton myasthenic syndrome (LEMS). Clinical outcome with respect to SOX antibodies was evaluated, as the SOX-related antitumor immune response might help to control the tumor growth. Patients and Methods We used recombinant SOX1, SOX2, SOX3, SOX21, HuC, HuD, or HelN1 proteins in an ELISA to titrate serum samples and validated the assay by western blot. We tested 136 consecutive SCLC patients, 86 LEMS patients (43 with SCLC), 14 patients with SCLC and PNS (paraneoplastic cerebellar degeneration or Hu syndrome), 62 polyneuropathy patients, and 18 healthy controls. Results Our ELISA was equally reliable as western blot. Forty-three percent of SCLC patients and 67% of SCLC-LEMS patients had antibodies to one of the SOX or Hu proteins. SOX antibodies had a sensitivity of 67% and a specificity of 95% to discriminate between LEMS with SCLC and nontumor LEMS. No difference in survival was observed between SOX positive and SOX negative SCLC patients. Conclusion SOX antibodies are specific serological markers for SCLC. Our assay is suitable for high throughput screening, detecting 43% of SCLC. SOX antibodies have diagnostic value in discriminating SCLC-LEMS from nontumor LEMS, but have no relation to survival in patients with SCLC.

scholarly journals Allergenic Characterization of Tropomyosin from the Dusky Brown Cockroach, Periplaneta fuliginosa

2004 ◽  
Vol 11 (4) ◽  
pp. 680-685 ◽  
Author(s):  
Kyoung Yong Jeong ◽  
Heeyu Hwang ◽  
Jongweon Lee ◽  
In-Yong Lee ◽  
Dong Soo Kim ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Periplaneta Americana ◽  
Immunoglobulin E ◽  
Expression System ◽  
Specific Ige ◽  
Cross Reactivity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Inhibitor Concentration ◽  
The Cross ◽  
Elisa Inhibition

ABSTRACTHousehold arthropods are one of the most common causes of allergic diseases. Four species of cockroaches are found to reside in Korean homes, but published work deals almost exclusively with the German and American cockroaches. This study was undertaken to investigate the cross-reactive allergenic components of the dusky brown cockroach,Periplaneta fuliginosa. Enzyme-linked immunosorbent assay (ELISA) inhibition and immunoblot analyses for the dusky brown cockroach were performed withBlattella germanicaandDermatophagoides farinaeallergic sera. cDNA encoding tropomyosin, which is a well known cross-reactive pan-allergen, was cloned by reverse transcriptase PCR, and recombinant protein was produced by using a pET-28b expression system. Native tropomyosin was purified by ammonium sulfate fractionation and electroelution. The immunoglobulin E (IgE) reactivities of native and recombinant tropomyosins were compared by an ELISA inhibition study. All 30 sera tested showedP. fuliginosa-specific IgE, and the IgE-binding reactivity of theP. fuliginosaextract was inhibited as much as 79.4% by aB. germanicaextract and as much as 63.3% by aD. farinaeextract. The deduced amino acid sequence of cloned cDNA was identical with that ofPeriplaneta americanatropomyosin (98.5% nucleotide sequence identity). Seven of 26 (26.9%) allergic sera had IgE specific for recombinant protein, and the maximum inhibition ofP. fuliginosa-specific IgE achieved with recombinant tropomyosin was 37.7% at an inhibitor concentration of 10 μg/ml. Native tropomyosin inhibited the binding of IgE to theP. fuliginosa,B. germanica, andD. farinaeextracts by 65.0, 51.8, and 39% at an inhibitor concentration of 1 μg/ml.P. fuliginosaappears to possess allergens that are highly cross-reactive with allergens ofB. germanicaandD. farinae. Tropomyosin was found to be a major allergenic component accounting for the cross-reactivity between cockroaches and dust mites.

scholarly journals Identification of Human Hepatocyte Proliferation Related Gene C2orf69

2014 ◽  
Vol 3 (1) ◽  
pp. 1-9
Author(s):  
Ren-wen Zhang ◽  
Yong Qiao ◽  
Xiao-hua Hao ◽  
Hong-min Li ◽  
Hui Ren ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Recombinant Protein ◽  
Western Blot ◽  
Polyclonal Antibody ◽  
Prokaryotic Expression ◽  
Liver Cells ◽  
Hepatocyte Proliferation ◽  
Enzyme Linked Immunosorbent Assay ◽  
Gene Fragment

Abstract Objective To construct the prokaryotic expression vector pET-32a(+)-C2orf69 and induce the expression of recombinant proteins in vitro. Then the possible effects of recombinant protein on cell proliferation was observed and rabbit-anti-C2orf69 protein polyclonal antibodies was obtained. Methods Gene fragment of C2orf69 was amplified by PCR and then prokaryotic expression plasmid pET-32a(+)-C2orf69 was constructed. Recombinant protein C2orf69 expression was identified by SDS-PAGE and Western blot. The white-ear rabbits were immunized with purified recombinant protein C2orf69, and the potency and specificity of polyclonal antibody were evaluated by enzyme-linked immunosorbent assay (ELISA) and Western blot. Also, different liver cells were incubated with recombinant protein C2orf69 in vitro. Results C2orf69 gene fragment was successfully amplified, results of gene sequencing were consistent with the sequence in GenBank. Recombinant protein of C2orf69 was successfully induced and expressed. The polyclonal antibody titer was up to 1︰1 280 000 through enzyme-linked immunosorbent assay. Results of cell proliferation showed that the recombinant protein could inhibit the proliferation of different liver cells. Conclusions The recombinant protein C2orf69 could inhibit the proliferation of different liver cells, and we speculated that it may be a widely roled inhibitor of hepatocyte proliferation. Our experiment showed that the proliferation inhibition of cells may be realized by G1 phase extending and S phase shortening.

scholarly journals COMPARISON OF A RAPID ENZYME-LINKED IMMUNOSORBANT ASSAY TEST WITH AN INDIRECT IMMUNOFLUORESCENT ANTIBODY TEST IN DIAGNOSING EHRLICHIA AND ANAPLASMA INFECTIONS IN DOGS

2019 ◽  
Vol 17 (4) ◽  
pp. 346-352
Author(s):  
Kr. Gospodinova ◽  
G. Zhelev ◽  
V. Petrov
Keyword(s):  
Serum Antibody ◽  
Rapid Test ◽  
Antibody Test ◽  
Diagnostic Value ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Immunofluorescent Antibody ◽  
Antibody Titers ◽  
Indirect Immunofluorescent ◽  
Immunofluorescent Antibody Test

PURPOSE: The objective of this study is to compare the diagnostic value of commercial enzyme-linked immunosorbent assay (ELISA) with indirect immunofluorescent antibody test (IFA) in detecting immunoglobulin-G (IgG) antibodies to Ehrlichia canis and Anaplasma phagocytophilum. METHODS: Seventy-four serum samples, obtained from dogs believed to be naturally infected with E. canis or A. phagocytophilum, were analyzed. RESULTS: By ELISA, 48 (64.9%) samples were found positive for IgG to E. canis, 10 (13.5%) to A. phagocytophilum, 12 (16.2%) to both E. canis and A. phagocytophilum, and in 4 (5.4%) samples no presence of antibodies was detected. The number of serologically positive dogs for IgG was 44 (59.5%) to E. canis, 10 (13.5%) to A. phagocytophilum, 16 (21.6%) to both E. canis and A. phagocytophilum, and 4 (5.4%) were determined negative by means of IFA. In most samples the antibody titer did not exceed 1:80 but in 5 it reached a level of 1:320, and in other 4 of even above 1:640. CONCLUSIONS: This study shows that IFA assay is more sensitive than commercial ELISA rapid test when serum antibody titers are low.

scholarly journals Comparative analysis of the allergenic characteristics and serodiagnostic potential of recombinant chitinase-like protein-5 and -12 from Sarcoptes scabiei

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Nengxing Shen ◽  
Yuhang Chen ◽  
Wenrui Wei ◽  
Lang Xiong ◽  
Yuanyuan Tao ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
High Sensitivity ◽  
Specific Ige ◽  
Sarcoptes Scabiei ◽  
Positive Control ◽  
Skin Tests ◽  
Enzyme Linked Immunosorbent Assay ◽  
Allergic Reactions ◽  
Diagnostic Potential ◽  
Recombinant Chitinase

Abstract Background Scabies is caused by burrowing of the mite Sarcoptes scabiei into the stratum corneum. Currently, diagnosis via routine skin scraping is very difficult, and information on the allergenic identification of S. scabiei remains limited. Methods We performed comparative analysis of the serological diagnostic potential of recombinant S. scabiei chitinase-like protein-5 (rSsCLP5) and recombinant S. scabiei chitinase-like protein-12 (rSsCLP12) by measuring the levels of serum-specific IgG and IgE antibodies (Abs) as diagnostic markers. In addition, the allergenic characteristics of rSsCLP5 and rSsCLP12 were evaluated using IgE-binding experiments and skin tests. Results The IgE Abs-based indirect enzyme-linked immunosorbent assay (ELISA) methods showed high sensitivity and specificity: the rSsCLP5-based assay had 93.5% sensitivity and 94.4% specificity; the rSsCLP12-based assay had 100% sensitivity and 98.1% specificity. The specific IgE Abs in infested mouse sera could bind rSsCLP5 and rSsCLP12. In skin tests, rabbits in the rSsCLP5 and rSsCLP12 groups and positive control (histamine) groups exhibited allergic reactions. Most test sites in the rSsCLP12 group had edema, bleeding spots, and even ulcers or scabs, but such allergy symptoms were rare in the rSsCLP5 group. Moreover, the allergic history rabbit group had more severe allergic reactions and lower levels of IgE Abs compared to the healthy rabbit group in the same protein group. Conclusions These findings validate the use of IgE Abs to rSsCLP5 and rSsCLP12 as potentially useful markers for diagnosing scabies. Moreover, both rSsCLP5 and rSsCLP12 have allergenic properties, and the potential allergen rSsCLP12 is a stronger allergen than rSsCLP5.

Diagnosis of schistosomiasis using recombinant fructose-1,6-bisphosphate aldolase from a Formosan strain ofSchistosoma japonicum

2009 ◽  
Vol 83 (3) ◽  
pp. 211-218 ◽  
Author(s):  
S.-Y. Peng ◽  
J.C. Tsaihong ◽  
P.-C. Fan ◽  
K.-M. Lee
Keyword(s):  
Amino Acid ◽  
Recombinant Protein ◽  
Clonorchis Sinensis ◽  
Diagnostic Value ◽  
Enzyme Linked Immunosorbent Assay ◽  
Small Animals ◽  
Amino Acid Residues ◽  
Faecal Examination ◽  
Human Schistosomiasis ◽  
Fructose 1,6 Bisphosphate

AbstractSchistosoma japonicumobtained from Taiwan is a zoophilic strain that only infects domestic and small animals. Recombinant fructose-1,6-bisphosphate aldolase (FBPA) derived from this strain was used as an antigen in an enzyme-linked immunosorbent assay (ELISA) for the diagnosis of human schistosomiasis. The full-length DNA sequence of FBPA was found to be 1092 bp, encoding a protein of 363 amino acid residues, with a molecular mass of 39.6 kDa. A total of 120 participants were recruited from China and Taiwan to evaluate the diagnostic value of this recombinant protein. In these participants, 34 were found to be infected withS. japonicum, 16 withAscaris lumbricoides, 15 with hookworm, 13 withParagonimus westermaniand 13 withClonorchis sinensis, whereas 29 had no ova on faecal examination. Western blot analysis showed that the recombinant FBPA reacts strongly with schistosome ova-positive sera. The sensitivity and specificity of ELISA with FBPA were found to be 85.3% and 93.0%, respectively. These results indicate that FBPA derived from the Formosan strain ofS. japonicumcan be used for the diagnosis of human schistosomiasis.

scholarly journals Circ_0000396 inhibits rheumatoid arthritis synovial fibroblast growth and inflammatory response via miR-203/HBP1 axis

2021 ◽  
Vol 28 (1) ◽  
Author(s):  
Laifang Wang ◽  
Qing Zhao ◽  
Na Wang ◽  
Yanjie Ding ◽  
Lingli Kong ◽  
...  
Keyword(s):  
Rheumatoid Arthritis ◽  
Western Blot ◽  
Synovial Fibroblasts ◽  
Diagnostic Value ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
Protective Effects ◽  
Tnf Α ◽  
Malignant Changes ◽  
Factor Α

Abstract Background Circ_0000396 was found to be down-regulated in the rheumatoid arthritis (RA) patients and had a high diagnostic value. However, the function and mechanisms underlying circ_0000396 in RA progression remain unclear. Methods The expression of circ_0000396, microRNA (miR)-203 and HMG-box transcription factor 1 (HBP1) was detected using qRT-PCR and western blot. The proliferative and apoptotic capabilities of rheumatoid arthritis synovial fibroblasts (RASFs) were measured by colony formation, CCK-8, flow cytometry and western blot assays, respectively. The levels of interleukins (IL)-6, IL-1β, IL-8 and tumor necrosis factor-α (TNF-α) were detected using enzyme-linked immunosorbent assay (ELISA). The target correlations between miR-203 and circ_0000396 or HBP1 were validated using pull-down and dual-luciferase reporter assay. Results Circ_0000396 was decreased in RA synovial tissues and RASFs, and overexpression of circ_0000396 suppressed cell proliferation, induced cell apoptosis and reduced the release of inflammatory cytokine IL-6, IL-1β, IL-8 and TNF-α in RASFs, while circ_0000396 deletion functioned oppositely. MiR-203 was confirmed to be a target of circ_0000396, and miR-203 reversed the protective effects of circ_0000396 on the dysfunction and inflammation of RASFs. HBP1 was a target of miR-203, and silencing miR-203 inhibited RASFs malignant changes by regulating HBP1. In addition, circ_0000396 could regulate HBP1 by sponging miR-203, and HBP1 decrease attenuated the effects of circ_0000396 on RASF growth and inflammation. Conclusion Circ_0000396 inhibited the growth and inflammation in RASFs by regulating miR-203/HBP1 axis, providing a potential therapeutic target for RA.

scholarly journals Evaluation of the Recombinant Protein TpF1 of Treponema pallidum for Serodiagnosis of Syphilis

2013 ◽  
Vol 20 (10) ◽  
pp. 1563-1568 ◽  
Author(s):  
Chuanhao Jiang ◽  
Feijun Zhao ◽  
Jinhong Xiao ◽  
Tiebing Zeng ◽  
Jian Yu ◽  
...  
Keyword(s):  
Recombinant Protein ◽  
Western Blot ◽  
Western Blotting ◽  
Congenital Syphilis ◽  
Treponema Pallidum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Western Blot Assay ◽  
Content Type ◽  
Human Sera ◽  
Serodiagnosis Of Syphilis

ABSTRACTSyphilis is a chronic infection caused byTreponema pallidumsubsp.pallidum, and diagnosis with sensitive and specific methods is a challenging process that is important for its prevention and treatment. In the present study, we established a recombinant protein TpF1-based indirect immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) and a Western blot assay for human and rabbit sera. The 20-kDa recombinant protein TpF1 was detected by Western blotting performed with sera from rabbits immunized with recombinant TpF1 and infected with theT. pallidumNichols strain andT. pallidumclinical isolates but was not detected by Western blotting with sera from uninfected rabbits. The sensitivity of the recombinant protein was determined by screening sera from individuals with primary, secondary, latent, and congenital syphilis (n= 82). The specificity of the recombinant protein was determined by screening sera from uninfected controls (n= 30) and individuals with potentially cross-reactive infections, including Lyme disease (n= 30) and leptospirosis (n= 5). The sensitivities of TpF1-based ELISAs were 93.3%, 100%, 100%, and 100% for primary, secondary, latent, and congenital syphilis, respectively, and the specificities were all 100% for sera from uninfected controls and individuals with potentially cross-reactive infections. In Western blot assays, the sensitivities and specificities of TpF1 for human sera were all 100%. The reactivities of TpF1 with syphilitic sera were proportional to the titers of theT. pallidumparticle agglutination (TPPA) assay. These data indicate that the recombinant protein TpF1 is a highly immunogenic protein in human and rabbit infections and a promising marker for the screening of syphilis.

scholarly journals Evaluation of salivary biomarkers for the diagnosis of periodontitis

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yong Zhang ◽  
Ni Kang ◽  
Fei Xue ◽  
Jing Qiao ◽  
Jinyu Duan ◽  
...  
Keyword(s):  
Periodontal Disease ◽  
Healthy Subjects ◽  
Type I Collagen ◽  
Area Under The Curve ◽  
Diagnostic Value ◽  
Enzyme Linked Immunosorbent Assay ◽  
Type I ◽  
Cross Sectional ◽  
Diagnostic Potential ◽  
Auc Value

Abstract Background Salivary interleukin (IL)-1β, matrix metalloproteinase (MMP)-8, pyridinoline cross-linked carboxyterminal telopeptide of type I collagen (ICTP) and Porphyromonas gingivalis (Pg) are related to periodontitis. This study aimed to investigate the diagnostic potential of these biomarkers and to build a prediction panel for diagnosing periodontal disease. Methods A total of 80 participants were enrolled in a cross-sectional study and divided into healthy (n = 25), gingivitis (n = 24), and periodontitis (n = 31) groups based on their periodontal exam results. A full mouth periodontal examination was performed and unstimulated saliva was collected. Salivary IL-1β, MMP-8, ICTP, and Pg were assessed using enzyme-linked immunosorbent assay (ELISA) and quantitative real time PCR (qPCR). Their potentials for diagnosing periodontal disease were analyzed and combined prediction panels of periodontal disease were evaluated. Results As a single marker, IL-1β showed the best diagnostic value of the four markers evaluated and exhibited an area under the curve (AUC) value of 0.88 with 90% sensitivity and 76% specificity for discriminating periodontitis subjects from healthy subjects, an AUC value of 0.80 with 83% sensitivity and 76% specificity for discriminating gingivitis subjects from healthy subjects and an AUC value of 0.66 with 68% sensitivity and 64% specificity for differentiating periodontitis subjects from gingivitis subjects. The combination of IL-1β, ICTP, and Pg exhibited the highest efficacy for discriminating periodontitis subjects from healthy subjects (AUC = 0.94) and gingivitis subjects (AUC = 0.77). The combination of IL-1β and MMP-8 exhibited the best ability to discriminate gingivitis from healthy subjects (AUC = 0.84). Conclusions Salivary IL-1β, MMP-8, ICTP, and Pg showed significant effectiveness for diagnosing periodontal disease. The combination of IL-1β, ICTP, and Pg can be used to discriminate periodontitis subjects from healthy subjects and gingivitis subjects, and the combination of IL-1β and MMP-8 can be used to discriminate gingivitis subjects from healthy subjects.

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