Rapid, Simple, Quantitative, and Highly Sensitive Antibody Detection for Lyme Disease

ABSTRACT There is currently a need for improved serological tests for the diagnosis and monitoring of Lyme disease, an infection caused by Borrelia burgdorferi. In the present study, we evaluated luciferase immunoprecipitation systems (LIPSs) for use for profiling of the antibody responses to a panel of B. burgdorferi proteins for the diagnosis of Lyme disease. Initially, serum samples from a cohort of patients and controls (n = 46) were used for training and were profiled by the use of 15 different B. burgdorferi antigen constructs. For the patient sera, the antibody responses to several B. burgdorferi antigens, including VlsE, flagellin (FlaB), BmpA, DbpA, and DbpB, indicated that the antigens had high levels of immunoreactivity. However, the best diagnostic performance was achieved with a synthetic protein, designated VOVO, consisting of a repeated antigenic peptide sequence, VlsE-OspC-VlsE-OspC, Analysis of an independent set of serum samples (n = 139) used for validation showed that the VOVO LIPS test had 98% sensitivity (95% confidence interval [CI], 93% to 100%; P < 0.0001) and 100% specificity (95% CI, 94% to 100%; P < 0.0001). Similarly, the C6 peptide enzyme-linked immunosorbent assay (ELISA) also had 98% sensitivity (95% CI, 93% to 100%; P < 0.0001) and 98% specificity (95% CI, 90% to 100%; P < 0.0001). Receiver operating characteristic analysis revealed that the rates of detection of Lyme disease by the LIPS test and the C6 ELISA were not statistically different. However, the VOVO LIPS test displayed a wide dynamic range of antibody detection spanning over 10,000-fold without the need for serum dilution. These results suggest that screening by the LIPS test with VOVO and other B. burgdorferi antigens offers an efficient quantitative approach for evaluation of the antibody responses in patients with Lyme disease.

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Borrelia burgdorferi Spirochetes That Harbor Only a Portion of the lp28-1 Plasmid Elicit Antibody Responses Detectable with the C6 Test for Lyme Disease

Clinical and Vaccine Immunology ◽

10.1128/cvi.00266-06 ◽

2006 ◽

Vol 14 (1) ◽

pp. 90-93 ◽

Cited By ~ 9

Author(s):

Monica E. Embers ◽

Gary P. Wormser ◽

Ira Schwartz ◽

Dale S. Martin ◽

Mario T. Philipp

Keyword(s):

Lyme Disease ◽

Borrelia Burgdorferi ◽

Antibody Responses ◽

Antibody Detection ◽

23S Rrna ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Altered Expression ◽

Genetic Region ◽

Length Polymorphisms

ABSTRACT Detection of antibody to C6, a peptide that reproduces the sequence of the sixth invariable region within the central domain of the VlsE protein of Borrelia burgdorferi, is used currently for the serologic diagnosis of Lyme disease in humans. B. burgdorferi isolates taken from infected humans can be categorized into specific genetic subtypes (designated RST1, -2, and -3) by restriction fragment length polymorphisms in the 16S to 23S rRNA spacer sequence. Many of these, usually categorized as RST2, retain only segments of the linear plasmid lp28-1, which encodes VlsE. The VlsE genetic region is retained, but altered expression of this molecule could affect diagnosis by the C6 enzyme-linked immunosorbent assay (ELISA). Serum samples from patients infected with each of the three genotypes and from mice infected with three RST2 isolates were tested with the C6 ELISA. Such isolates elicited marked C6 responses in infected mice. The sensitivity of C6 antibody detection in patients infected with RST2 spirochetes was statistically indistinguishable from detection of RST1 and RST3 infections. These findings demonstrate that diagnosis by C6 ELISA remains effective for infection with all B. burgdorferi genotypes, including those with incomplete lp28-1 plasmids.

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B-Cell-Specific Peptides of Leptospira interrogans LigA for Diagnosis of Patients with Acute Leptospirosis

Clinical and Vaccine Immunology ◽

10.1128/cvi.00456-13 ◽

2014 ◽

Vol 21 (3) ◽

pp. 354-359 ◽

Cited By ~ 11

Author(s):

Murugesan Kanagavel ◽

Santhanam Shanmughapriya ◽

Kumarasamy Anbarasu ◽

Kalimuthusamy Natarajaseenivasan

Keyword(s):

Early Diagnosis ◽

B Cell ◽

Predictive Value ◽

Acute Stage ◽

Antibody Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Serological Tests ◽

Content Type

ABSTRACTLeptospirosis is a reemerging infectious disease that is underdiagnosed and under-recognized due to low-sensitivity and cumbersome serological tests. Rapid reliable alternative tests are needed for early diagnosis of the disease. Considering the importance of the pathogenesis-associated leptospiral LigA protein expressedin vivo, we have evaluated its application in the diagnosis of the acute form of leptospirosis. The C-terminal coding sequence ofligA(ligA-C) was cloned into pET15b and expressed inEscherichia coli. Furthermore, the B-cell-specific epitopes were predicted and were synthesized as peptides for evaluation along with recombinant LigA-C. Epitope 1 (VVIENTPGK), with a VaxiJen score of 1.3782, and epitope 2 (TALSVGSSK), with a score of 1.2767, were utilized. A total of 140 serum samples collected from leptospirosis cases during the acute stage of the disease and 138 serum samples collected from normal healthy controls were utilized for evaluation. The sensitivity, specificity, positive predictive value, and negative predictive value were calculated for the recombinant LigA-C-specific IgM enzyme-linked immunosorbent assay (ELISA) and were found to be 92.1%, 97.7%, 92.8%, and 97.5%, respectively. Epitopes 1 and 2 used in the study showed 5.1 to 5.8% increased sensitivity over recombinant LigA-C in single and combination assays for IgM antibody detection. These findings suggest that these peptides may be potential candidates for the early diagnosis of leptospirosis.

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Prevalence and clinical correlation of hepatitis E virus antibody in the patients’ serum samples from a tertiary care hospital in Thailand during 2015–2018

Virology Journal ◽

10.1186/s12985-021-01616-x ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Atiporn Boonyai ◽

Anchalee Thongput ◽

Thidarat Sisaeng ◽

Parisut Phumchan ◽

Navin Horthongkham ◽

...

Keyword(s):

Pregnant Women ◽

Tertiary Care ◽

Laboratory Data ◽

Organ Transplant ◽

Hospital Setting ◽

Antibody Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Care Hospital ◽

Serum Samples ◽

Igm Antibodies

Abstract Background Prevalence and incidence of hepatitis caused by HEV infection are usually higher in developing countries. This study demonstrated the HEV seroprevalence and incidence of HEV infection in patients with clinical hepatitis in a tertiary hospital in Thailand. Methods A laboratory-based cross-sectional study was conducted using 1106 serum samples from patients suspected of HEV infection sent to the Serology laboratory, Siriraj Hospital, for detecting HEV antibodies during 2015–2018. Prevalence of anti-HEV IgG and IgM antibodies in general patients, including organ transplant recipients and pregnant women in a hospital setting, were determined using indirect enzyme-linked immunosorbent assay (ELISA) kits. Comparison of laboratory data between groups with different HEV serological statuses was performed. Results HEV IgG antibodies were detected in 40.82% of 904 serum samples, while HEV IgM antibodies were detected in 11.75% of 1081 serum samples. Similar IgG and IgM antibody detection rates were found in pregnant women. Interestingly, anti-HEV IgM antibodies were detected in 38.5% of patients who underwent organ transplantation. Patients who tested positive for anti-HEV IgM antibodies had higher alanine aminotransferase levels than those who had not. In contrast, patients who tested positive for anti-HEV IgG had more elevated levels of total bilirubin than those who tested negative. Conclusions HEV seroprevalence and incidence in patients with clinical hepatitis were relatively high in the Thai population, including the pregnancy and organ transplant subgroups. The results potentially benefit the clinicians in decision-making to investigate HEV antibodies and facilitating proper management for patients.

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

Infection and Immunity ◽

10.1128/iai.69.5.3224-3231.2001 ◽

2001 ◽

Vol 69 (5) ◽

pp. 3224-3231 ◽

Cited By ~ 17

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.

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Diagnosis of Dengue Virus Infection by Detection of Specific Immunoglobulin M (IgM) and IgA Antibodies in Serum and Saliva

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.10.2.317-322.2003 ◽

2003 ◽

Vol 10 (2) ◽

pp. 317-322 ◽

Cited By ~ 83

Author(s):

Angel Balmaseda ◽

María G. Guzmán ◽

Samantha Hammond ◽

Guillermo Robleto ◽

Carolina Flores ◽

...

Keyword(s):

Dengue Virus ◽

Immunoglobulin M ◽

Antibody Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Control Group ◽

Serum Samples ◽

Diagnostic Modality ◽

Iga Antibodies ◽

Specific Immunoglobulin ◽

Feasible Alternative

ABSTRACT To evaluate alternative approaches to the serological diagnosis of dengue virus (DEN) infection, the detection of DEN-specific immunoglobulin M (IgM) and IgA antibodies in serum and saliva specimens was assessed in 147 patients with symptoms of DEN infection seen at the Ministry of Health in Nicaragua. Seventy-two serum samples were determined to be positive for anti-DEN antibodies by IgM capture enzyme-linked immunosorbent assay, the routine diagnostic procedure. Serum and saliva specimens were obtained from 50 healthy adults as additional controls. IgM was detected in the saliva of 65 of the 72 serum IgM-positive cases, 6 of the 75 serum IgM-negative cases, and none of the control group, resulting in a sensitivity of 90.3% and a specificity of 92.0% and demonstrating that salivary IgM is a useful diagnostic marker for DEN infection. Detection of IgA in serum may be another feasible alternative for the diagnosis of DEN infection, with serum IgA found in 68 (94.4%) of the IgM-positive cases. In contrast, detection of IgA in saliva was not found to be a useful tool for DEN diagnosis in the present study. Further studies of the kinetics of antibody detection in another set of 151 paired acute- and convalescent-phase serum samples showed that DEN-specific IgA antibodies were detected in more acute-phase samples than were IgM antibodies. Thus, we conclude that DEN-specific IgA in serum is a potential diagnostic target. Furthermore, given that saliva is a readily obtainable, noninvasive specimen, detection of DEN-specific salivary IgM should be considered a useful, cheaper diagnostic modality with similar sensitivity and specificity to IgM detection in serum.

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Serological Evaluation of Specific-Antibody Levels in Patients Treated for Chronic Chagas' Disease

Clinical and Vaccine Immunology ◽

10.1128/cvi.00106-07 ◽

2007 ◽

Vol 15 (2) ◽

pp. 297-302 ◽

Cited By ~ 38

Author(s):

Olga Sánchez Negrette ◽

Fernando J. Sánchez Valdéz ◽

Carlos D. Lacunza ◽

María Fernanda García Bustos ◽

María Celia Mora ◽

...

Keyword(s):

Chagas Disease ◽

Treatment Effectiveness ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Serological Tests ◽

Recombinant Antigens ◽

Antibody Titers ◽

Laboratory Procedures ◽

The Individual ◽

Antibody Levels

ABSTRACT Serological tests are the main laboratory procedures used for diagnosis during the indeterminate and chronic stages of Chagas' disease. A serological regression to negativity is the main criterion used to define parasitological cure in treated patients. The aim of this work was to monitor the individual specificities of antibody levels for 3 years posttreatment in 18 adult patients. Conventional serological techniques (hemagglutination assays and enzyme-linked immunosorbent assay [ELISA]) were modified by using recombinant antigens to detect early markers of treatment effectiveness. For this purpose, serum samples were taken before and during treatment and every 6 months after treatment for at least 3 years. When hemagglutination assays were used, a decrease in antibody levels was observed in only one patient. When ELISA with serum dilutions was used, antibody clearance became much more apparent: in 77.7% (14/18) of the patients, antibody titers became negative with time. This was observed at serum dilutions of 1/320 and occurred between the 6th and the 30th months posttreatment. The immune response and the interval for a serological regression to negativity were different for each patient. For some of the recombinant antigens, only 50% (9/18) of the patients reached the serological regression to negativity. Recombinant antigen 13 might be a good marker of treatment effectiveness, since 66.6% (six of nine) of the patients presented with an early regression to negativity for specific antibodies to this antigen (P = 0.002).

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Quantitative Approach for the Serodiagnosis of Canine Lyme Disease by the Immunoblot Procedure

Journal of Clinical Microbiology ◽

10.1128/jcm.38.7.2628-2632.2000 ◽

2000 ◽

Vol 38 (7) ◽

pp. 2628-2632 ◽

Cited By ~ 11

Author(s):

Marta A. Guerra ◽

Edward D. Walker ◽

Uriel Kitron

Keyword(s):

Logistic Regression ◽

Lyme Disease ◽

Natural Infection ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Immunoblot Assay ◽

Serological Status ◽

Low Probability ◽

Positive Results ◽

Logistic Regression Equation

Serum samples obtained from healthy, asymptomatic dogs in areas of Wisconsin and northern Illinois where Lyme disease is endemic or nonendemic were assayed for antibodies to Borrelia burgdorferi by enzyme-linked immunosorbent assay (ELISA), and positive results were confirmed by immunoblot assay. We found that 56.9% (562 of 1,077) of the samples were positive by ELISA and 82.0% (461 of 562) were positive by immunoblotting. A logistic regression model was developed to distinguish between nonvaccinated dogs naturally infected with B. burgdorferi from areas where the disease is endemic and dogs from areas where the disease is nonendemic that were vaccinated against Lyme disease. Of the 18 protein bands analyzed, 8 were found to be significantly different (P < 0.05) between the two groups. p93, p34, p31, and p28 occurred with increased frequency in vaccinated dogs, while p58, p37, p35, and p30 occurred more frequently in naturally infected dogs. The logistic regression equation obtained was used to determine the probability of natural infection among vaccinated dogs residing in areas where the disease is endemic. Of 125 samples, 87.2% had a very low probability of natural infection and only 2.4% were highly likely to be infected. Logistic regression is a useful method for distinguishing between vaccinated and naturally infected dogs and predicting the serological status of vaccinated dogs from areas where Lyme disease is endemic.

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Serodiagnosis of Louse-Borne Relapsing Fever with Glycerophosphodiester Phosphodiesterase (GlpQ) fromBorrelia recurrentis

Journal of Clinical Microbiology ◽

10.1128/jcm.38.10.3561-3571.2000 ◽

2000 ◽

Vol 38 (10) ◽

pp. 3561-3571 ◽

Cited By ~ 54

Author(s):

Stephen F. Porcella ◽

Sandra J. Raffel ◽

Merry E. Schrumpf ◽

Martin E. Schriefer ◽

David T. Dennis ◽

...

Keyword(s):

Serological Test ◽

Geometric Mean ◽

Eastern Africa ◽

Enzyme Linked Immunosorbent Assay ◽

Relapsing Fever ◽

Serum Samples ◽

Serological Tests ◽

Whole Cell ◽

Convalescent Phase ◽

Glycerophosphodiester Phosphodiesterase

Human louse-borne relapsing fever occurs in sporadic outbreaks in central and eastern Africa that are characterized by significant morbidity and mortality. Isolates of the causative agent,Borrelia recurrentis, were obtained from the blood of four patients during a recent epidemic of the disease in southern Sudan. TheglpQ gene, encoding glycerophosphodiester phosphodiesterase, from these isolates was sequenced and compared with the glpQ sequences obtained from other relapsing-fever spirochetes. Previously we showed that GlpQ of Borrelia hermsii is an immunogenic protein with utility as a serological test antigen for discriminating tick-borne relapsing fever from Lyme disease. In the present work, we cloned and expressed theglpQ gene from B. recurrentis and used recombinant GlpQ in serological tests. Acute- and convalescent-phase serum samples obtained from 42 patients with louse-borne relapsing fever were tested with an indirect immunofluorescence assay (IFA) and an enzyme-linked immunosorbent assay (ELISA) that used whole cells ofB. recurrentis and with immunoblotting to whole-cell lysates of the spirochete and Escherichia coli producing recombinant GlpQ. The geometric mean titers of the acute- and convalescent-phase serum samples measured by IFA were 1:83 and 1:575, respectively. The immunoblot analysis identified a high level of reactivity and seroconversion to GlpQ, and the assay was more sensitive than the whole-cell IFA and ELISA using purified, recombinant histidine-tagged GlpQ. Serum antibodies to GlpQ and other antigens persisted for 27 years in one patient. We conclude that assessment of anti-GlpQ antibodies will allow serological confirmation of louse-borne relapsing fever and determination of disease prevalence.

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Immunoglobulin A Antibody Responses in Dengue Patients: a Useful Marker for Serodiagnosis of Dengue Virus Infection

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.12.10.1235-1237.2005 ◽

2005 ◽

Vol 12 (10) ◽

pp. 1235-1237 ◽

Cited By ~ 14

Author(s):

M. Nawa ◽

T. Takasaki ◽

M. Ito ◽

S. Inoue ◽

K. Morita ◽

...

Keyword(s):

Virus Infection ◽

Dengue Virus ◽

Immunoglobulin A ◽

Antibody Responses ◽

Enzyme Linked Immunosorbent Assay ◽

Virus Infections ◽

Serum Samples ◽

Dengue Virus Infection ◽

Iga Antibody ◽

Antibody Capture

ABSTRACT We determined the usefulness of an immunoglobulin A (IgA) antibody-capture enzyme-linked immunosorbent assay for serodiagnosis of dengue virus infections. The results indicate that the presence of IgA and IgM in serum samples assures recent primary dengue virus infection even with a single serum sample.

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The latest FAD – Faecal antibody detection in cattle. Protocol and results from three UK beef farms naturally infected with gastrointestinal nematodes

Parasitology ◽

10.1017/s0031182018000902 ◽

2018 ◽

Vol 146 (1) ◽

pp. 89-96 ◽

Cited By ~ 1

Author(s):

A. S. Cooke ◽

K. A. Watt ◽

E. R. Morgan ◽

J. A. J. Dungait

Keyword(s):

Animal Health ◽

Nematode Species ◽

Gastrointestinal Nematodes ◽

Vital Role ◽

Antibody Detection ◽

Enzyme Linked Immunosorbent Assay ◽

Serum Samples ◽

Fecal Samples ◽

Gastrointestinal Health ◽

Immunological Protection

AbstractAntibodies at gastrointestinal mucosal membranes play a vital role in immunological protection against a range of pathogens, including helminths. Gastrointestinal health is central to efficient livestock production, and such infections cause significant losses. Fecal samples were taken from 114 cattle, across three beef farms, with matched blood samples taken from 22 of those animals. To achieve fecal antibody detection, a novel fecal supernatant was extracted. Fecal supernatant and serum samples were then analysed, using adapted enzyme-linked immunosorbent assay protocols, for levels of total immunoglobulin (Ig)A, IgG, IgM, andTeladorsagia circumcincta-specific IgA, IgG, IgM and IgE (in the absence of reagents for cattle-specific nematode species). Fecal nematode egg counts were conducted on all fecal samples. Assays performed successfully and showed that IgA was the predominant antibody in fecal samples, whereas IgG was predominant in serum. Total IgA in feces and serum correlated within individuals (0.581,P= 0.005), but other Ig types did not. Results support the hypothesis that the tested protocols are an effective method for the non-invasive assessment of cattle immunology. The method could be used as part of animal health assessments, although further work is required to interpret the relationship between results and levels of infection and immunity.

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