scholarly journals Affimer-Based Europium Chelates Allow Sensitive Optical Biosensing in a Range of Human Disease Biomarkers

Sensors ◽  
2021 ◽  
Vol 21 (3) ◽  
pp. 831
Author(s):  
Eiman Al-Enezi ◽  
Alexandre Vakurov ◽  
Amy Eades ◽  
Mingyu Ding ◽  
Gin Jose ◽  
...  
Keyword(s):  
Human Serum ◽  
Cardiac Injury ◽  
Visible Region ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Optical Biosensing ◽  
Rapid Measurement ◽  
Fluorescence Characteristics ◽  
Human Myoglobin ◽  
Diagnostic Applications

The protein biomarker measurement has been well-established using ELISA (enzyme-linked immunosorbent assay), which offers good sensitivity and specificity, but remains slow and expensive. Certain clinical conditions, where rapid measurement or immediate confirmation of a biomarker is paramount for treatment, necessitate more rapid analysis. Biosensors offer the prospect of reagent-less, processing-free measurements at the patient’s bedside. Here, we report a platform for biosensing based on chelated Eu3+ against a range of proteins including biomarkers of cardiac injury (human myoglobin), stroke (glial fibrillary acidic protein (GFAP)), inflammation (C-reactive protein (CRP)) and colorectal cancer (carcinoembryonic antigen (CEA)). The Eu3+ ions are chelated by modified synthetic binding proteins (Affimers), which offer an alternative targeting strategy to existing antibodies. The fluorescence characteristics of the Eu3+ complex with modified Affimers against human myoglobin, GFAP, CRP and CEA were measured in human serum using λex = 395 nm, λem = 590 and 615 nm. The Eu3+-Affimer based complex allowed sensitive detection of human myoglobin, GFAP, CRP and CEA proteins as low as 100 fM in (100-fold) diluted human serum samples. The unique dependence on Eu3+ fluorescence in the visible region (590 and 615 nm) was exploited in this study to allow rapid measurement of the analyte concentration, with measurements in 2 to 3 min. These data demonstrate that the Affimer based Eu3+ complexes can function as nanobiosensors with potential analytical and diagnostic applications.

scholarly journals C-Terminal Invariable Domain of VlsE Is Immunodominant but Its Antigenicity Is Scarcely Conserved among Strains of Lyme Disease Spirochetes

2001 ◽  
Vol 69 (5) ◽  
pp. 3224-3231 ◽  
Author(s):  
Fang Ting Liang ◽  
Lisa C. Bowers ◽  
Mario T. Philipp
Keyword(s):  
Lyme Disease ◽  
Antibody Response ◽  
Human Serum ◽  
Synthetic Peptide ◽  
Species Complex ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serum Samples ◽  
Variable Surface ◽  
Entire Sequence ◽  
Carboxyl Terminal

ABSTRACT VlsE, the variable surface antigen of Borrelia burgdorferi, contains two invariable domains located at the amino and carboxyl terminal ends, respectively, and a central variable domain. In this study, both immunogenicity and antigenic conservation of the C-terminal invariable domain were assessed. Mouse antiserum to a 51-mer synthetic peptide (Ct) which reproduced the entire sequence of the C-terminal invariable domain of VlsE from B. burgdorferi strain B31 was reacted on immunoblots with whole-cell lysates extracted from spirochetes of 12 strains from the B. burgdorferi sensu lato species complex. The antiserum recognized only VlsE from strain B31, indicating that epitopes of this domain differed among these strains. When Ct was used as enzyme-linked immunosorbent assay (ELISA) antigen, all of the seven monkeys and six mice that were infected with B31 spirochetes produced a strong antibody response to this peptide, indicating that the C-terminal invariable domain is immunodominant. None of 12 monkeys and only 11 of 26 mice that were infected with strains other than B31 produced a detectable anti-Ct response, indicating a limited antigenic conservation of this domain among these strains. Twenty-six of 33 dogs that were experimentally infected by tick inoculation were positive by the Ct ELISA, while only 5 of 18 serum samples from dogs clinically diagnosed with Lyme disease contained detectable anti-Ct antibody. Fifty-seven of 64 serum specimens that were collected from American patients with Lyme disease were positive by the Ct ELISA, while only 12 of 21 European samples contained detectable anti-Ct antibody. In contrast, antibody to the more conserved invariable region IR6 of VlsE was present in all of these dog and human serum samples.

scholarly journals Seroprevalence of Jamestown Canyon virus in the Japanese general population

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Hirofumi Kato ◽  
Masaaki Satoh ◽  
Madoka Kawahara ◽  
Satoshi Kitaura ◽  
Tomoki Yoshikawa ◽  
...  
Keyword(s):  
General Population ◽  
Human Serum ◽  
Health Agency ◽  
Febrile Illness ◽  
Central Nervous System Disease ◽  
Enzyme Linked Immunosorbent Assay ◽  
Antibody Assay ◽  
Serum Samples ◽  
Human Serum Samples ◽  
Jamestown Canyon Virus

Abstract Background Jamestown Canyon virus (JCV) is a mosquito-borne orthobunyavirus that causes acute febrile illness, meningitis, and meningoencephalitis, mainly among adults. JCV is widely distributed in North America and the number of JCV cases in the U.S. has increased in recent years. Therefore, the central nervous system disease caused by JCV can be considered a potentially re-emerging viral disease. However, the seroprevalence of JCV is unknown in Japan. The purpose of this study is to evaluate the seroprevalence of JCV in the Japanese population. Methods We used an IgG enzyme-linked immunosorbent assay (IgG-ELISA) with JCV-infected cell-lysates and/or a neutralizing (NT) antibody assay. The cut-off value of IgG-ELISA was determined using IgG-ELISA to analyze serum specimens from 37 healthy Japanese donors. IgG-ELISA was validated by assessing its sensitivity and specificity, using 38 human serum samples previously tested for the presence or absence of antibodies against JCV and snowshoe hare virus (SSHV), in an in-house NT antibody assay conducted by the Public Health Agency of Canada. The seroepidemiological study was performed using IgG-ELISA and NT antibody assay to analyze 246 human serum samples from the serum bank of the National Institute of Infectious Diseases (NIID) in Japan. Results The cut-off value of IgG-ELISA was determined at 0.20, based on the mean (− 0.075) and standard deviation (0.092) values using Japanese donors’ sera. The sensitivity and the specificity of IgG-ELISA determined using 25 JCV-positive and 4 JCV-negative serum samples were 96 and 100%, respectively. Analysis of the 246 Japanese serum samples revealed that no specimen showed a higher value than the cut-off value of IgG-ELISA, and no sample tested positive by the NT antibody assay. Conclusions Our results showed that JCV is not circulating significantly in Japan. To the best of our knowledge, this is the first report to demonstrate the seroprevalence of JCV in the general population in Japan.

scholarly journals Development of the ELISA Test System for Quantitative Determination of IgG to the Varizella zoster virus in Human Serum and Assessment of its Diagnostic Efficiency

2022 ◽  
Vol 20 (6) ◽  
pp. 72-80
Author(s):  
L. N. Lukhverchyk ◽  
G. L. Alatortseva ◽  
L. N. Nesterenko ◽  
V. Y. Kabargina ◽  
V. V. Dotsenko ◽  
...  
Keyword(s):  
Herpes Zoster ◽  
Blood Serum ◽  
Human Serum ◽  
Immunosorbent Assay ◽  
Test System ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diagnostic Efficiency ◽  
Functional Characteristics ◽  
Serum Samples ◽  
Population Immunity

Relevance. The introduction of Varicella vaccine prophylaxis explains the need to develop a methodology for monitoring the vaccination effectiveness and the intensity of population immunity. This problem can be solved using quantitative immunoassay methods. Aim. Development of an enzyme-linked immunosorbent assay for the concentration of class G immunoglobulins (AB) to Varicella zoster virus (VZV) determining and assessing its functional characteristics and diagnostic efficiency. Materials and methods. Recombinant antigen GE VZV. WHO International Standard for Antibodies to VZV W1044. Blood serum samples from healthy people and patients with Chickenpox and Herpes zoster, blood serum samples containing IgG antibodies to herpes simplex viruses of the first and second types, cytomegalovirus, Epstein-Barr virus. Anti-VZV ELISA (IgG) reagent kit (Euroimmun, Germany). Indirect enzyme-linked immunosorbent assay. Immunization of animals with recombinant antigen GE, isolation, and purification of specific antibodies. Conjugation of monoclonal antibodies to human IgG with antibodies to antigen GE and with horseradish peroxidase. Results. An enzyme-linked immunosorbent assay in «an indirect» format has been developed to determine the specific antibodies to VZV concentration (IU/ml) in human serum/plasma. An artificial calibrator for determining the concentration of AB-VZV had been synthesized and standardized according to the International WHO-standard W1044. The main functional characteristics of the developed enzyme-linked immunosorbent assay are determined in accordance with GOST 51352-2013. The diagnostic kit was tested on blood serum samples from children with chickenpox (n = 43), adults with Herpes zoster (n = 158), healthy individuals (n = 781). The diagnostic sensitivity of the test system was 85%, the diagnostic specificity was 87% according to the ROC analysis. The absence of cross-reactivity of the test system was shown on samples with serological markers of other herpesvirus infections (n = 94). Comparative trials of the developed test system and its commercial analog, the Anti-VZV ELISA (IgG) reagent kit, did not reveal statistically significant differences between their functional characteristics. Conclusions. The developed test system for determining of the AB-VZV concentration in human serum/plasma in terms of its functional characteristics meets the GOST requirements, is characterized by high diagnostic efficiency, can be used to monitor the effectiveness of vaccine prophylaxis and strength of population immunity, as well as to assess the immune response in chickenpox and Herpes zoster.

Analysis of UK Sera for Aflatoxin by Enzyme-linked Immunosorbent Assay

1988 ◽  
Vol 7 (4) ◽  
pp. 353-356 ◽  
Author(s):  
A.P. Wilkinson ◽  
D.W. Denning ◽  
M.R.A. Morgan
Keyword(s):  
Human Serum ◽  
Direct Measurement ◽  
Immunosorbent Assay ◽  
Human Exposure ◽  
Blood Donors ◽  
Enzyme Linked Immunosorbent Assay ◽  
Fungal Metabolites ◽  
Serum Samples ◽  
Imported Food ◽  
The Uk

1 Aflatoxins are toxic, carcinogenic secondary fungal metabolites produced by certain moulds that commonly infest foods. Measurement of aflatoxins in human serum would give a direct measurement of exposure. 2 Twenty-seven serum samples from UK blood donors were found to contain aflatoxin levels not greater than 64 pmol/1 (20 pg/ml) by an enzyme-linked immunosorbent assay. 3 These findings may indicate that present UK guideline tolerances for aflatoxin in imported food are effective in limiting human exposure to toxic aflatoxins in the UK diet, though further work would be needed to confirm this. In particular, sub-populations suspected of being at higher risk may need special considerations.

scholarly journals Antibody Responses to Escherichia coli O157 and Other Lipopolysaccharides in Healthy Children and Adults

2003 ◽  
Vol 10 (5) ◽  
pp. 797-801 ◽  
Author(s):  
Armando Navarro ◽  
Carlos Eslava ◽  
Ulises Hernandez ◽  
Jose Luis Navarro-Henze ◽  
Magali Aviles ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Human Serum ◽  
Western Blotting ◽  
Rabbit Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Children ◽  
Serum Samples ◽  
E Coli ◽  
Human Serum Samples ◽  
Bactericidal Activities

ABSTRACT In Mexico, diarrheal disease due to different serotypes of Escherichia coli is highly prevalent, with only sporadic isolation of O157 non-H7 strains. This could be due to exposure to the O157 or related E. coli lipopolysaccharide (LPS), such as O7 or O116, at an early age. By using enzyme-linked immunosorbent assay (ELISA) and Western blotting, the present study analyzed 605 serum samples from Mexican adults and infants without clinical symptoms of disease for the presence of antibodies to these three E. coli LPSs. The bactericidal activities of homologous and heterologous rabbit and human serum samples against O7, O116, and O157 E. coli LPSs were also determined. By using a cutoff point of 0.7, it was found by the ELISAs that 28 of 562 (5%) of the serum samples from adolescents and adults and 2 of 43 (5%) of the serum samples from infants less than 1 year of age reacted with the O157 LPS. By using cutoff points between 0.4 and 0.699, the proportion of serum samples from both age groups that reacted with the O157 LPS increased to 20%. Western blotting analysis of selected serum samples that showed an intermediate response against the O157 LPS by the ELISAs showed that 61 of 88 (69%) reacted with the same LPS. A similar result was observed for maternal milk samples. The bactericidal activities of rabbit serum samples against the O7, O116, and O157 LPSs showed that they were positive for both homologous and heterologous antigens. Similar results were observed with the human serum samples. O157 non-H7 strains were identified in only 10% of the E. coli strains isolated from 263 Mexican children with and without diarrhea over the past 15 years. This absence of O157:H7 strains in Mexico may be associated with the presence of antibodies against O157 or related E. coli LPSs.

High positive frequency of antibodies to metallothionein and heat shock protein 70 in sera of patients with metal allergy

2020 ◽  
Author(s):  
Lungwani Muungo
Keyword(s):  
Heat Shock ◽  
Human Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Strong Positive Correlation ◽  
Metal Allergy ◽  
Serum Samples ◽  
Group Iii ◽  
Group I ◽  
Group Ii ◽  
Positive Frequency

Two principal types of stress protein, heat shock proteins (hsps) and metallothionein (MT), are inducedin cells responding to a variety of stresses. They play an important role in protecting cells from thesestresses. However, many reports indicate that antibodies to hsps are present in human serum and areassociated with several autoimmunity diseases. Metals, which are commonly allergenic to humans,induce both MT and hsp70 (one of the hsps family). Until now, there has been no report of any antibodyto MT in human serum. In the present study, serum samples from healthy controls (Group I), andpatients suffering from atopic dermatitis without (Group II) or with (Group III) metal allergy, weremeasured for antibodies to MT and hsp70, using an enzyme-linked immunosorbent assay (ELISA).Metal allergy was confirmed by patch testing. We first found that antibody to MT exists in human serum.We also found a high positive frequency of antibody to MT (51·3%) and to hsp70 (43·6%) in the seraof Group III, compared to those of Group I (3·8% and 5·1%) or Group II (6·4% and 5·1%). Furthermore,there was a strong positive correlation between antibody to MT and antibody to hsp70 in GroupIII (P = 0·0013), but not in Group I and Group II. Our results indicate that antibody to MT exists inhuman serum, as do antibodies to hsps, and suggest

scholarly journals Use of Moraxella catarrhalis Lipooligosaccharide Mutants To Identify Specific Oligosaccharide Epitopes Recognized by Human Serum Antibodies

2009 ◽  
Vol 77 (10) ◽  
pp. 4548-4558 ◽  
Author(s):  
Johanna M. Schwingel ◽  
Katie J. Edwards ◽  
Andrew D. Cox ◽  
Hussein Masoud ◽  
James C. Richards ◽  
...  
Keyword(s):  
Immune Response ◽  
Antibody Response ◽  
Human Serum ◽  
Moraxella Catarrhalis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Human Antibody ◽  
Serum Antibodies ◽  
Serum Samples ◽  
Carbohydrate Epitopes ◽  
Tract Infections

ABSTRACT Moraxella catarrhalis is a causative agent of otitis media in children and lower respiratory tract infections in adults suffering from chronic obstructive pulmonary disease (COPD). This strict human pathogen continues to be a significant cause of disease in this broad spectrum of patients because there is no available vaccine. Although numerous putative vaccine antigens have been described, little is known about the human immune response to M. catarrhalis infection in vivo. Human serum antibodies are directed at a number of surface proteins, and lipooligosaccharides (LOS) and detoxified LOS may be an effective immunogen in mice. In this study, we used a specific LOS-based enzyme-linked immunosorbent assay (ELISA), containing the three major M. catarrhalis serotypes together with a complete series of truncated LOS mutants, to detect the development of new antibodies to specific regions of the oligosaccharide molecule. We compared serum samples from COPD patients who had recently cleared an M. catarrhalis infection to serum samples collected prior to their infection. Variability in the antibody response to LOS was observed, as some patients developed serotype-specific antibodies, others developed antibodies to the LOS of each serotype, others developed broadly cross-reactive antibodies, and some did not develop new antibodies. These newly developed human antibodies are directed at both side chains and core structures in the LOS molecule. This LOS-based ELISA can be used to dissect the human antibody response to both internal and external carbohydrate epitopes, thus providing a better understanding of the humoral immune response to M. catarrhalis LOS epitopes developed during natural infection.

scholarly journals Evaluation of an Enzyme Immunoassay for Detection of Dengue Virus NS1 Antigen in Human Serum

2006 ◽  
Vol 13 (11) ◽  
pp. 1185-1189 ◽  
Author(s):  
Philippe Dussart ◽  
Bhety Labeau ◽  
Gisèle Lagathu ◽  
Philippe Louis ◽  
Marcio R. T. Nunes ◽  
...  
Keyword(s):  
Dengue Virus ◽  
Human Serum ◽  
Enzyme Immunoassay ◽  
Immunoglobulin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Diagnostic Strategy ◽  
Serum Samples ◽  
Dengue Disease ◽  
Reverse Transcription Pcr ◽  
Ns1 Antigen

ABSTRACT We evaluated a one-step sandwich-format microplate enzyme immunoassay for detecting dengue virus NS1 antigen (Ag) in human serum by use of Platelia Dengue NS1 Ag kits (Bio-Rad Laboratories, Marnes La Coquette, France). We collected 299 serum samples from patients with dengue disease and 50 serum samples from patients not infected with dengue virus. For the 239 serum samples from patients with acute infections testing positive by reverse transcription-PCR and/or virus isolation for one of the four dengue virus serotypes, the sensitivity of the Platelia Dengue NS1 Ag kit was 88.7% (95% confidence interval, 84.0% to 92.4%). None of the serum samples from patients not infected with dengue virus tested positive with the Platelia Dengue NS1 Ag kit. A diagnostic strategy combining the Platelia Dengue NS1 Ag test for acute-phase sera and immunoglobulin M capture enzyme-linked immunosorbent assay for early-convalescent-phase sera increased sensitivity only from 88.7% to 91.9%. Thus, NS1 antigen detection with the Platelia Dengue NS1 Ag kit could be used for first-line testing for acute dengue virus infection in clinical diagnostic laboratories.

Modeling of the desialylated human serum N-glycome for molecular diagnostic applications in inflammatory and malignant lung diseases

2020 ◽  
Vol 20 ◽  
Author(s):  
Anna Farkas ◽  
Brigitta Mészáros ◽  
Máté Szarka ◽  
Márton Szigeti ◽  
János Kappelmayer ◽  
...  
Keyword(s):  
Human Serum ◽  
Lung Diseases ◽  
Molecular Diagnostic ◽  
Control Group ◽  
Serum Samples ◽  
Protein Mixture ◽  
The Core ◽  
Diagnostic Potential ◽  
Alpha 1 Antitrypsin ◽  
Diagnostic Applications

Background: Immunoglobulin G and A, transferrin, haptoglobin and alpha-1-antitrypsin are representing approximately 85% of the human serum glycoproteome and their N-glycosylation analysis may lead to discover important molecular disease markers. However, due to the labile nature of the sialic acid residues, the desialylated subset of the serum N-glycoproteome has been traditionally utilized for diagnostic applications. Objective: Creating a five-protein model to deconstruct the overall N-glycosylation fingerprints in inflammatory and malignant lung diseases. Methods: The N-glycan pool of human serum and the five high abundant serum glycoproteins were analyzed. Simultaneous endoglycosidase/sialidase digestion was followed by fluorophore labeling and separation by CE-LIF to establish the model. Pooled serum samples from patients with COPD, lung cancer (LC) and their comorbidity were all analyzed. Results: Nine significant (>1%) asialo-N-glycan structures were identified both in human serum and the standard protein mixture. The core-fucosylated-agalacto-biantennary glycan differentiated COPD and LC and both from the control and the comorbidity groups. Decrease in the core-fucosylated-agalacto-biantennary-bisecting, monogalacto and bigalacto structures differentiated all disease groups from the control. The significant increase of the fucosylated- galactosylated-triantennary structure was highly specific for LC, in medium extent for COPD and in lesser extent for comorbidity. Also, some increase of the afucosylated-galactosylatedbiantennary structure in all three disease types and afucosylated-galactosylated-triantennary structures in COPD and LC were observed in comparison to the control group. Conclusion: Our results suggested that changes in the desialylated human serum N-glycome holds glycoprotein specific molecular diagnostic potential for malignant and inflammatory lung diseases, which can be modeled with the five-protein mixture.

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