scholarly journals Identification of Campylobacter jejuni Proteins Recognized by Maternal Antibodies of Chickens

2008 ◽  
Vol 74 (22) ◽  
pp. 6867-6875 ◽  
Author(s):  
Kari D. Shoaf-Sweeney ◽  
Charles L. Larson ◽  
Xiaoting Tang ◽  
Michael E. Konkel
Keyword(s):  
Campylobacter Jejuni ◽  
Immunoblot Analysis ◽  
Protective Role ◽  
Maternal Antibodies ◽  
In Vitro Assays ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Homologous Strain ◽  
Protective Antibodies

ABSTRACT Campylobacter jejuni is one of the leading bacterial causes of food-borne gastroenteritis. Infection with C. jejuni is frequently acquired through the consumption of undercooked poultry or foods cross-contaminated with raw poultry. Given the importance of poultry as a reservoir for Campylobacter organisms, investigators have performed studies to understand the protective role of maternal antibodies in the ecology of Campylobacter colonization of poultry. In a previous study, chicks with maternal antibodies generated against the S3B strain of C. jejuni provided protection against Campylobacter colonization (O. Sahin, N. Luo, S. Huang, and Q. Zhang, Appl. Environ. Microbiol. 69:5372-5379, 2003). We obtained serum samples, collectively referred to as the C. jejuni S3B-SPF sera, from the previous study. These sera were determined to contain maternal antibodies that reacted against C. jejuni whole-cell lysates as judged by enzyme-linked immunosorbent assay. The antigens recognized by the C. jejuni S3B-SPF antibodies were identified by immunoblot analysis, coupled with mass spectrometry, of C. jejuni outer membrane protein extracts. This approach led to the identification of C. jejuni proteins recognized by the maternal antibodies, including the flagellin proteins and CadF adhesin. In vitro assays revealed that the C. jejuni S3B-SPF sera retarded the motility of the C. jejuni S3B homologous strain but did not retard the motility of a heterologous strain of C. jejuni (81-176). This finding provides a possible mechanism explaining why maternal antibodies confer enhanced protection against challenge with a homologous strain compared to a heterologous strain. Collectively, this study provides a list of C. jejuni proteins against which protective antibodies are generated in hens and passed to chicks.

scholarly journals Development of a flow cytometric bead immunoassay and its assessment as a possible aid to potency evaluation of enterotoxaemia vaccines

2014 ◽  
Vol 85 (1) ◽  
Author(s):  
Angela Buys ◽  
Raynard Macdonald ◽  
Jannie Crafford ◽  
Jacques Theron
Keyword(s):  
Cost Effective ◽  
In Vitro Assays ◽  
Enzyme Linked Immunosorbent Assay ◽  
In Vitro Tests ◽  
Serum Samples ◽  
Type D ◽  
Epsilon Toxin ◽  
Bead Immunoassay

Enterotoxaemia, an economically important disease of sheep, goats and calves, is caused by systemic effects of the epsilon toxin produced by the anaerobic bacterium Clostridium perfringens type D. The only practical means of controlling the occurrence of enterotoxaemia is to immunise animals by vaccination. The vaccine is prepared by deriving a toxoid from the bacterial culture filtrate and the potency of the vaccine is tested with the in vivo mouse neutralisation test (MNT). Due to ethical, economic and technical reasons, alternative in vitro assays are needed. In this study an indirect cytometric bead immunoassay (I-CBA) was developed for use in vaccine potency testing and the results were compared with those obtained using an indirect enzyme-linked immunosorbent assay (I-ELISA) and the MNT. Sera were collected from guinea pigs immunised with three different production batches of enterotoxaemia vaccine and the levels of anti-epsilon toxin antibodies were determined. Although the intra- and inter-assay variability was satisfactory, epsilon antitoxin levels determined by both the I-ELISA and indirect cytometric bead immunoassay (I-CBA) tests were higher than those of the MNT assay. In contrast to the MNT, all of the serum samples were identified as having antitoxin levels above the required minimum (not less than 5 U/mL). These results indicate that the respective in vitro tests in their current formats are not yet suitable alternatives to the in vivo MNT. The growing demand for a more humane, cost-effective and efficient method for testing the potency of enterotoxaemia vaccines, however, provides a strong impetus for further optimisation and standardisation of the I-CBA assay but further analytical research is required.

scholarly journals Prevalence, Antigenic Specificity, and Bactericidal Activity of Poultry Anti-Campylobacter Maternal Antibodies

2001 ◽  
Vol 67 (9) ◽  
pp. 3951-3957 ◽  
Author(s):  
Orhan Sahin ◽  
Qijing Zhang ◽  
Jerrel C. Meitzler ◽  
Brian S. Harr ◽  
Teresa Y. Morishita ◽  
...  
Keyword(s):  
Broiler Chickens ◽  
Potential Role ◽  
Maternal Antibodies ◽  
Antigenic Specificity ◽  
Enzyme Linked Immunosorbent Assay ◽  
Poultry Production ◽  
Serum Samples ◽  
Animal Reservoir

ABSTRACT Poultry are considered the major reservoir for Campylobacter jejuni, a leading bacterial cause of human food-borne diarrhea. To understand the ecology of C. jejuni and develop strategies to control C. jejuni infection in the animal reservoir, we initiated studies to examine the potential role of anti-Campylobacter maternal antibodies in protecting young broiler chickens from infection by C. jejuni. Using an enzyme-linked immunosorbent assay (ELISA), the prevalence of anti-C. jejuni antibodies in breeder chickens, egg yolks, and broilers from multiple flocks of different farms were examined. High levels of antibodies to the organism were detected in serum samples of breeder chickens and in egg yolk contents. To determine the dynamics of anti-Campylobacter maternal antibody transferred from yolks to hatchlings, serum samples collected from five broiler flocks at weekly intervals from 1 to 28 or 42 days of age were also examined by ELISA. Sera from the 1-day and 7-day-old chicks showed high titers of antibodies to C. jejuni. Thereafter, antibody titers decreased substantially and were not detected during the third and fourth weeks of age. The disappearance of anti-Campylobacter maternal antibodies during 3 to 4 weeks of age coincides with the appearance of C. jejuniinfections observed in many broiler chicken flocks. As shown by immunoblotting, the maternally derived antibodies recognized multiple membrane proteins of C. jejuni ranging from 19 to 107 kDa. Moreover, in vitro serum bactericidal assays showed that anti-Campylobacter maternal antibodies were active in antibody-dependent complement-mediated killing of C. jejuni. Together, these results highlight the widespread presence of functional anti-Campylobacter antibodies in the poultry production system and provide a strong rationale for further investigation of the potential role of anti-C. jejunimaternal antibodies in protecting young chickens from infection byC. jejuni.

P–314 Serum Stem Cell Factor as a predictor of top quality blasotcysts formation in women with endometriosis

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
J Liss ◽  
M Kuczynska ◽  
A Knight ◽  
K Lukaszuk
Keyword(s):  
Stem Cell ◽  
Ovarian Stimulation ◽  
Stem Cell Factor ◽  
Enzyme Linked Immunosorbent Assay ◽  
Tyrosine Kinase Receptor ◽  
Target Cells ◽  
Serum Samples ◽  
The Mean ◽  
Ovary Reserve

Abstract Study question To evaluate the correlation between the serum level of stem cell factor (s-SCF) during the stimulation and results of embryo culture. Summary answer The serum SCF concentration at the stimulation stage may be a potential predictor of IVF outcome in endometriosis patients. What is known already Stem cell factor (SCF) is a pleiotropic cytokine that affects the target cells via the c-kit receptor, a tyrosine kinase receptor. Recent evidence indicates that SCF and c-kit may play a role in regulation and growth of ovarian follicular function. It is unclear whether endometriosis primarily affects in vitro fertilization outcomes via oocyte quality. SCF is produced during the human follicular phase, immediately before the ovulatory phase, and may play an important role in folliculogenesis and in the mechanism of ovulation. It may reflect a successful stimulation with ample follicle maturation. Study design, size, duration This was a prospective case-control study and consisted four group of patients: 10 with endometriosis, 24 PCOs, 20 with normal (AMH 1.2–4.0 ng/ml) and 11 with lower (AMH<1.2 ng/ml) ovary reserve who were undergoing IVF treatment with the assessment of serum SCF concentration between August 2019 and March 2020 at INVICTA Fertility Centre, Poland. The age of the patients ranged from 22 to 42 years (median 34 years). Participants/materials, setting, methods s-SCF was measured in duplicate by enzyme-linked immunosorbent assay (ELISA) kit in 195 serum samples collected during ovarian stimulation on days 1 and 8 and on the day of oocyte retrieval. We analysed correlation between s-SCF level and formation of top quality (TQ) blastocysts on day 5 formation in the study groups. Main results and the role of chance We have compared mean level of s-SCF within each group dividing the patients into two subgroups – those with at least one TQ blastocyst (TQ) on day 5 vs. those with no TQ blastocysts (no-TQ). There were no significant differences in mean s-SCF level on day 1 of stimulation between no-TQ and TQ patients in PCOs, normal and lower ovary reserve groups (41.1 pg/ml vs. 40.9 pg/ml; 34.8 pg/ml vs. 38.9 pg/ml and 32.3pg/ml vs. 28.7 pg/ml respectively). The mean level of s-SCF in endometriosis patients was higher in case of no-TQ compared to the TQ subgroup and were 41.1 pg/ml and 29.1 pg/ml respectively. Also no significant differences were also observed in the mean level of s-SCF in the no-TQ and TQ subgroups on the 8 day of stimulation and pick-up in PCOs, normal and lower ovary reserve patients. However, again in the case of endometriosis patients, the mean level of s-SCF was significantly lower on the 8 day of stimulation (28.1 pg/ml vs. 49.1 pg/ml; p < 0.05) and pick-up day (33.4 pg/ml vs. 50.4 pg/ml; p < 0.005) in samples from patients who had at least one TQ blastocysts on day 5 of culture. Limitations, reasons for caution More data are required to confirm the corelation of s-SCF level and presence of top quality blastocysts in patients with endometriosis. Wider implications of the findings: Our study suggests that the level of serum SCF during ovarian stimulation in patients with endometriosis of less 30 pg/ml may potentially be a predictor for the chance of obtaining at least one top quality blastocyst on day 5 and thus a chance to successful treatment. Trial registration number Not applicable

scholarly journals Expression of the Campylobacter jejuni FliD protein and its reaction to chicken sera

2020 ◽  
Vol 367 (14) ◽  
Author(s):  
Xiao-Yan Zhang ◽  
Qian Zhou ◽  
Meng-Jun Tang ◽  
Jun-Hua Pu ◽  
Yan-Feng Fan ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Broiler Chickens ◽  
Human Infection ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Capping Protein ◽  
Cultural Status ◽  
Bacterial Gastroenteritis ◽  
Antibody Levels ◽  
Culture Negative

ABSTRACT Campylobacter is a leading causative pathogen of acute bacterial gastroenteritis among humans. Contaminated chicken products are regarded as major sources of human infection. The flagellar capping protein (FliD), which plays important roles in colonization and adhesion to the mucosal surface of chicken ceca, is conserved among Campylobacter jejuni strains. In this study, the recombinant C. jejuni FliD protein was expressed, purified and used as a coated protein to examine the prevalence of C. jejuni antibodies in chickens. The anti-FliD antibody was prevalent among chicken serum samples taken from different farms in the diverse regions of Jiangsu province by using enzyme-linked immunosorbent assay. The Campylobacter antibody was present in culture-negative chickens. No strong dose–response relationships were observed between serum FliD antibody levels and Campylobacter cultural status. These results provide a basis for further evaluating FliD as a vaccine candidate for broiler chickens or for examining host–C. jejuni interactions, with implications for improving food safety.

scholarly journals Serodiagnosis of Human Granulocytic Ehrlichiosis by a Recombinant HGE-44-Based Enzyme-Linked Immunosorbent Assay

1999 ◽  
Vol 37 (11) ◽  
pp. 3540-3544 ◽  
Author(s):  
Jacob W. Ijdo ◽  
Caiyun Wu ◽  
Louis A. Magnarelli ◽  
Erol Fikrig
Keyword(s):  
Immunosorbent Assay ◽  
Fluorescent Antibody ◽  
Immunoblot Analysis ◽  
Normal Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antibody Testing ◽  
Serum Samples ◽  
Granulocytic Ehrlichiosis ◽  
Healthy Humans ◽  
Human Granulocytic Ehrlichiosis

Current antibody testing for human granulocytic ehrlichiosis relies predominantly on indirect fluorescent-antibody assays and immunoblot analysis. Shortcomings of these techniques include high cost and variability of test results associated with the use of different strains of antigens derived from either horses or cultured HL-60 cells. We used recombinant protein HGE-44, expressed and purified as a maltose-binding protein (MBP) fusion peptide, as an antigen in a polyvalent enzyme-linked immunosorbent assay (ELISA). Fifty-five normal serum samples from healthy humans served as a reference to establish cutoff levels. Thirty-three of 38 HGE patient serum samples (87%), previously confirmed by positive whole-cell immunoblotting, reacted positively in the recombinant ELISA. In specificity analyses, serum samples from patients with Lyme disease, syphilis, rheumatoid arthritis, and human monocytic ehrlichiosis (HME) did not react with HGE-44–MBP antigen, except for one sample (specificity, 98%). We conclude that recombinant HGE-44 antigen is a suitable antigen in an ELISA for the laboratory diagnosis and epidemiological study of HGE.

IL-38 exerts anti-inflammatory and anti-fibrotic effects in thyroid-associated ophthalmopathy

2021 ◽  
Author(s):  
Lu Shi ◽  
Huijing Ye ◽  
Jun Huang ◽  
Yanbing Li ◽  
Xing Wang ◽  
...  
Keyword(s):  
Disease Activity ◽  
Protective Role ◽  
Enzyme Linked Immunosorbent Assay ◽  
Clinical Activity ◽  
Connective Tissues ◽  
Thyroid Associated Ophthalmopathy ◽  
Organ Specific ◽  
Anti Inflammatory ◽  
Star Molecules

Abstract Context Thyroid-associated ophthalmopathy (TAO) is an organ-specific autoimmune disease closely associated with Graves’ disease (GD). IL-38, a novel cytokine in the IL-1 superfamily, has been reported to be involved in the pathogenesis of various autoimmune diseases. Objectives To evaluate the relationship between IL-38 and TAO disease activity and its role in inflammation and fibrosis in TAO. Design/Setting/Participants Blood samples and orbital connective tissues were collected from TAO patients and controls. Orbital fibroblasts (OFs) were isolated from patients with TAO. Main Outcome Measures Enzyme-linked immunosorbent assay (ELISA), immunohistochemistry (IHC), flow cytometry, immunofluorescence, quantitative real-time PCR and western blot analysis were performed. Results Here, we demonstrated that IL-38 levels decreased in the circulation and orbital connective tissues of patients with TAO compared to the controls, and were negatively correlated with the clinical activity score (CAS). In vitro, potent anti-inflammatory and anti-fibrotic effects of IL-38 were observed. Furthermore, we revealed that IL-38 can counteract the phosphorylation of star molecules in multiple classical pathways. Conclusions IL-38 plays a protective role in TAO and is associated with its pathogenesis. Our data suggest that IL-38 may be a promising marker of TAO disease activity and a potential target for TAO therapy.

scholarly journals Expression of a dominant allele of human ARF1 inhibits membrane traffic in vivo

1994 ◽  
Vol 124 (3) ◽  
pp. 289-300 ◽  
Author(s):  
CJ Zhang ◽  
AG Rosenwald ◽  
MC Willingham ◽  
S Skuntz ◽  
J Clark ◽  
...  
Keyword(s):  
Golgi Apparatus ◽  
Rat Kidney ◽  
Brefeldin A ◽  
Membrane Traffic ◽  
Immunoblot Analysis ◽  
In Vitro Assays ◽  
Wild Type ◽  
Discernible Effect

ADP-ribosylation factor (ARF) proteins and inhibitory peptides derived from ARFs have demonstrated activities in a number of in vitro assays that measure ER-to-Golgi and intra-Golgi transport and endosome fusion. To better understand the roles of ARF proteins in vivo, stable cell lines were obtained from normal rat kidney (NRK) cells transfected with either wild-type or a dominant activating allele ([Q71L]) of the human ARF1 gene under the control of the interferon-inducible mouse Mx1 promoter. Upon addition of interferon, expression of ARF1 proteins increased with a half-time of 7-8 h, as determined by immunoblot analysis. Induction of mutant ARF1, but not wild-type ARF1, led to an inhibition of protein secretion with kinetics similar to that observed for induction of protein expression. Examination of the Golgi apparatus and the ER by indirect immunofluorescence or transmission electron microscopy revealed that expression of low levels of mutant ARF1 protein correlated with a dramatic increase in vesiculation of the Golgi apparatus and expansion of the ER lumen, while expression of substantially higher levels of wild-type ARF1 had no discernible effect. Endocytosis was also inhibited by expression of mutant ARF1, but not by the wild-type protein. Finally, the expression of [Q71L]ARF1, but not wild-type ARF1, antagonized the actions of brefeldin A, as determined by the delayed loss of ARF and beta-COP from Golgi membranes and disruption of the Golgi apparatus. General models for the actions of ARF1 in membrane traffic events are discussed.

The effects of BAFF and BAFF-R-Fc fusion protein in immune thrombocytopenia

Blood ◽  
2009 ◽  
Vol 114 (26) ◽  
pp. 5362-5367 ◽  
Author(s):  
Xiao-juan Zhu ◽  
Yan Shi ◽  
Jun Peng ◽  
Cheng-shan Guo ◽  
Ning-ning Shan ◽  
...  
Keyword(s):  
Fusion Protein ◽  
Immune Thrombocytopenia ◽  
Interleukin 4 ◽  
In Vitro Assays ◽  
Enzyme Linked Immunosorbent Assay ◽  
Cd8 Cells ◽  
Promising Therapeutic Approach ◽  
Fc Fusion Protein ◽  
Ifn Γ

Abstract Elevated level of B-cell activating factor (BAFF) has been implicated in the pathogenesis of some autoimmune diseases. Blockade of receptor and ligand binding by decoy receptor has demonstrated a clinical benefit in both oncologic and immunologic diseases. In this report, we have detected plasma BAFF and BAFF mRNA expression in immune thrombocytopenia (ITP) patients by enzyme-linked immunosorbent assay (ELISA) and real-time quantitative reverse-transcription polymerase chain reaction (RT-PCR). The effects of recombinant human BAFF (rhBAFF) and BAFF-R-Fc fusion protein (BR3-Fc) on B cells, T cells, platelets, secretion of interferon γ (IFNγ), and interleukin-4 (IL-4) were measured by flow cytometry and ELISA. Patients with active disease had higher levels of plasma BAFF and BAFF mRNA than patients in remission and controls. In in vitro assays, rhBAFF promoted the survival of CD19+ and CD8+ cells, and increased the apoptosis of platelets and the secretion of IFN-γ. BR3-Fc successfully corrected the effects of rhBAFF on lymphocytes, platelets, and cytokines. These findings suggest that BAFF may play a pathogenic role in ITP by promoting the survival of CD19+ and CD8+ cells, and increasing the apoptosis of platelets and the secretion of IFN-γ. Blockade of BAFF by BR3-Fc might be a promising therapeutic approach for ITP.

scholarly journals Novel NFκB Inhibitor SC75741 Mitigates Chondrocyte Degradation and Prevents Activated Fibroblast Transformation by Modulating miR-21/GDF-5/SOX5 Signaling

2021 ◽  
Vol 22 (20) ◽  
pp. 11082
Author(s):  
Pei-Wei Weng ◽  
Vijesh Kumar Yadav ◽  
Narpati Wesa Pikatan ◽  
Iat-Hang Fong ◽  
I-Hsin Lin ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Protective Role ◽  
Enzyme Linked Immunosorbent Assay ◽  
Molecule Inhibitor ◽  
Tnf Α ◽  
Inflammatory Profile ◽  
Fibroblast Transformation ◽  
Tgf Β1

Osteoarthritis (OA) is a common articular disease manifested by the destruction of cartilage and compromised chondrogenesis in the aging population, with chronic inflammation of synovium, which drives OA progression. Importantly, the activated synovial fibroblast (AF) within the synovium facilitates OA through modulating key molecules, including regulatory microRNAs (miR’s). To understand OA associated pathways, in vitro co-culture system, and in vivo papain-induced OA model were applied for this study. The expression of key inflammatory markers both in tissue and blood plasma were examined by qRT-PCR, western blot, immunohistochemistry, enzyme-linked immunosorbent assay (ELISA) and immunofluorescence assays. Herein, our result demonstrated, AF-activated human chondrocytes (AC) exhibit elevated NFκB, TNF-α, IL-6, and miR-21 expression as compared to healthy chondrocytes (HC). Importantly, AC induced the apoptosis of HC and inhibited the expression of chondrogenesis inducers, SOX5, TGF-β1, and GDF-5. NFκB is a key inflammatory transcription factor elevated in OA. Therefore, SC75741 (an NFκB inhibitor) therapeutic effect was explored. SC75741 inhibits inflammatory profile, protects AC-educated HC from apoptosis, and inhibits miR-21 expression, which results in the induced expression of GDF-5, SOX5, TGF-β1, BMPR2, and COL4A1. Moreover, ectopic miR-21 expression in fibroblast-like activated chondrocytes promoted osteoblast-mediated differentiation of osteoclasts in RW264.7 cells. Interestingly, in vivo study demonstrated SC75741 protective role, in controlling the destruction of the articular joint, through NFκB, TNF-α, IL-6, and miR-21 inhibition, and inducing GDF-5, SOX5, TGF-β1, BMPR2, and COL4A1 expression. Our study demonstrated the role of NFκB/miR-21 axis in OA progression, and SC75741′s therapeutic potential as a small-molecule inhibitor of miR-21/NFκB-driven OA progression.

scholarly journals Characterization of a Borrelia burgdorferi VlsE Invariable Region Useful in Canine Lyme Disease Serodiagnosis by Enzyme-Linked Immunosorbent Assay

2000 ◽  
Vol 38 (11) ◽  
pp. 4160-4166 ◽  
Author(s):  
Fang Ting Liang ◽  
Richard H. Jacobson ◽  
Reinhard K. Straubinger ◽  
Amy Grooters ◽  
Mario T. Philipp
Keyword(s):  
Lyme Disease ◽  
Borrelia Burgdorferi ◽  
Antibody Response ◽  
Immunosorbent Assay ◽  
Protein A ◽  
Immunoblot Analysis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Entire Study ◽  
Variable Surface

Sera collected from dogs experimentally infected withBorrelia burgdorferi by tick inoculation were analyzed for an antibody response to each of the six invariable regions (IRs; i.e., IR1 to IR6) of VlsE, the variable surface antigen of B. burgdorferi. Six synthetic peptides (C1 to C6), which reproduced the six IR sequences were used as peptide-based, enzyme-linked immunosorbent assay (ELISA) antigens. Two IRs, IR2 and IR6, were found to be immunodominant. Studies with serially collected serum samples from experimentally infected dogs revealed that the antibody response to IR6 appears earlier and is stronger than that to IR2. Thus, the IR6 sequence alone appeared to be sufficient for serodiagnosis. When C6 alone was used as antigen, the peptide-based ELISA was positive in 7 of 23 dogs (30%) as early as 3 weeks postinfection. All dogs (n = 33) became strongly positive 1 or 2 weeks later, and this response persisted for the entire study, which lasted for 69 weeks. Of 55 sera submitted by veterinarians from dogs suspected of having Lyme disease, 19 were also positive by the C6 ELISA, compared to 20 positives detected by immunoblot analysis using cultured B. burgdorferi lysates as antigen. The sensitivity of using C2 and C6 together for detecting specific antibody in both experimentally infected and clinically diagnosed dogs was not better than sensitivity with C6 alone, confirming that C6 suffices as a diagnostic probe. Moreover, the C6 ELISA yielded 100% specificity with serum samples collected from 70 healthy dogs, 14 dogs with infections other than B. burgdorferi, and 15 animals vaccinated with either outer surface protein A, whole-spirochete vaccines, or the common puppy-vaccines. Therefore, this C6 ELISA was both sensitive and specific for the serodiagnosis of canine Lyme disease and could be used with vaccinated dogs.

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