Double-antigen Sandwich Enzyme-linked Immunosorbent Assay Research Articles

Epidemiological assessment of hepatitis E virus infection among 1565 pregnant women in Siem Reap, Cambodia using an in-house double antigen sandwich ELISA.

This study investigated hepatitis E virus (HEV) prevalence among pregnant women in Siem Reap, Cambodia, by developing a cost-effective, user-friendly in-house enzyme-linked immunosorbent assay (ELISA) for detecting total anti-HEV immunoglobulins (Ig). The in-house ELISA was designed for large-scale screening in resource-limited settings. Its performance was benchmarked against two commercial tests: the Anti-HEV IgG EIA (Institute of Immunology, Co. Ltd) and the Anti-HEV IgG RecomLine LIA (Mikrogen). The in-house ELISA demonstrated a sensitivity of 76% and 71.4%, and a specificity of 94.1% and 98.6%, against the two commercial tests, respectively, with overall agreement rates of 92.4% and 94.3%. Among 1565 tested pregnant women, 11.6% were anti-HEV positive. Prevalence increased with age, particularly in women aged 35-40years and over 40years. No significant associations were found with education, number of children, family size, or history of blood transfusion and surgery, except for the occupation of the family head as a public officer. Of the total anti-HEV positive women, 22.7% had anti-HEV IgM, indicating recent or ongoing infection. The study concluded that the in-house ELISA is a viable option for HEV screening in regions with limited resources due to its high accuracy and cost-effectiveness. It is particularly suitable for large-scale studies and public health interventions in areas where HEV is endemic and poses a significant risk to pregnant women.

WITHDRAWN: Development of a double-antigen sandwich ELISA for African swine fever virus antibody detection based on K205R protein

African swine fever (ASF) is a highly fatal infectious disease that affects swine caused by the African swine fever virus (ASFV) and poses a significant threat to the swine industry, and its effective diagnosis is crucial due to the current lack of commercially available vaccine for disease prevention and control. In this regard, the K205R protein is a suitable diagnostic antigen for enzyme-linked immunosorbent assay (ELISA) owing to its continuous expression throughout all ASFV replication stages, high immunogenicity, and sequence conservation. The double-antigen sandwich ELISA (DAgS-ELISA) is capable of detecting all immunoglobulins present in the serum, and outperforms alternative methods by enabling earlier antibody detection. Therefore, in this study, the K205R protein was prokaryotically expressed, and the K205R protein digested His-tag was labeled with horseradish peroxidase (HRP) and named K205R-HRP. The DAgS-ELISA was conducted using a combination of digested His-tag proteins and K205R-HRP. This method exhibits exceptional properties, including high sensitivity and no cross-reactivity with other porcine antibodies, with both the intra- and inter-assay coefficients of variation (CV) below 10 %. Antibody kinetics experiments have demonstrated that this method is capable of detecting antibodies as early as 7 to 9 days post-infection, which is a significant improvement compared to the commercial kit that can only detect antibodies from 11 to 13 days post-infection. In 120 clinical serum tests, the Kappa value between the DAgS-ELISA and the commercial kit was 0.96 (almost perfect). Owing to its high sensitivity, impressive repeatability, and strong specificity, this method enables earlier antibody detection and is suitable for ASFV antibody testing.

Development of an effective one-step double-antigen sandwich ELISA based on p72 to detect antibodies against African swine fever virus.

African swine fever (ASF), caused by ASF virus (ASFV), is a highly contagious and lethal disease of domestic pigs leading to tremendous economic losses. As there are no vaccines and drugs available. An effective diagnosis to eliminate ASFV-infected pigs is a crucial strategy to prevent and control ASF. To this end, ASFV capsid protein p72 was expressed using Chinese hamster ovary (CHO) cells and subsequently conjugated with horseradish peroxidase (HRP) to develop a one-step double-antigen sandwich enzyme-linked immunosorbent assay (one-step DAgS-ELISA). The performance of this ELISA for detecting ASFV antibodies was evaluated. Overall, a diagnostic sensitivity of 97.96% and specificity of 98.96% was achieved when the cutoff value was set to 0.25. No cross-reaction with healthy pig serum and other swine viruses was observed. The coefficients of variation of the intra-assay and inter-assay were both <10%. Importantly, this ELISA could detect antibodies in standard serum with 12,800-fold dilution, and seroconversion started from the 7th day post-inoculation (dpi), showing excellent analytical sensitivity and great utility. Furthermore, compared to the commercial kit, this ELISA had a good agreement and significantly shorter operation time. Collectively, a novel one-step DAgS-ELISA for detecting antibodies against ASFV is developed, which will be reliable and convenient to monitor ASFV infection.

Development and evaluation of recombinant E2 protein based IgM capture enzyme-linked immunosorbent assay (ELISA) and double antigen sandwich ELISA for detection of antibodies to Chikungunya virus

Chikungunya virus (CHIKV) reemerged and caused millions of human infections since 2004. The disease could be established, when the virus has been introduced to areas where the appropriate vectors are endemic. The differential diagnosis of CHIKV infection varies based on place of residence, travel history, and exposures. Serological tests are commonly used to diagnose CHIKV infection, but their availability and assessments of the performance of the diagnostics have been limited. To develop and evaluate antibodies detection methods for chikungunya diagnosis and serological investigation. Recombinant E2 protein based IgM capture enzyme-linked immunosorbent assay (Mac-ELISA) and double antigen sandwich ELISA (Das-ELISA) for detection of antibodies to Chikungunya virus were developed and evaluated. The repeatability was evaluated by testing of three reference sera at single dilutions in triplicated for 5 times. The sensitivity, specificity, accuracy, and agreement of the MAC-ELISA and Das-ELISA were obtained by comparing the detection results of 225 serum samples (45 positive; 180 negative) with a real-time RT-PCR assay and an IFA commercial tests manufactured by Euroimmun. The established ELISA assays were standardized by determining the optimal concentrations of the key reagents. The coefficient values of repeat testing were within 10% and 20% for intraassay and interassay precision, respectively. A sensitivity of 60.0% and 52.5%, a specificity of 96.2% and 96.8%, and an accuracy of 89.8% and 88.9% were obtained for the Mac-ELISA and Das-ELISA, respectively, when compared to a CHIKV qRT-PCR method. And a sensitivity of 100%, a specificity of 97.5% and 99.5%, and an accuracy of 97.8% and 99.6% were yielded respectively when using the IIFT as a reference method, which showed a highly consistence to the commercial IIFT assay with a Kappa value greater than 0.90. The Mac-ELISA and Das-ELISA based on recombinant E2 protein of CHIKV were developed and standardized, which could detect IgM or total antibodies against CHIKV in 2-3 hours with acceptable sensitivities and specificities. These assays can be used for laboratory diagnosis and serological investigation of CHIKV infections to evaluate the risk of CHIKV transmission.

Seroprevalence of Crimean-Congo Hemorrhagic Fever Among Small Ruminants from Southern Romania.

Crimean-Congo hemorrhagic fever (CCHF) is a tick-borne disease that can be contracted by direct contact with viremic animals or humans. Domestic animals are accidental hosts and contribute to the spread and amplification of the virus. The main objective of this study was to provide updated information related to CCHF virus (CCHFV) infection in Southern Romania by assessing the seroprevalence of CCHF in small ruminants (sheep and goats) using a double-antigen sandwich enzyme-linked immunosorbent assay and by detection of CCHFV in engorged ticks and serum samples using real-time RT-PCR. The overall seroprevalence of CCHF in small ruminants was 37.7% (95% CI 31.7 to 43.7). No statistical seroprevalence difference was observed between the two species of ruminants (p = 0.76), but a significant difference was established between the locations (p < 0.01). No CCHFV RNA was detected in tick pools and small ruminant's sera tested by real-time RT-PCR, although the high seroprevalence to CCHFV among ruminants indicates that CCHV or a closely related virus circulates in Southern Romania.

Seroprevalence of Severe Fever with Thrombocytopenia Syndrome in Southeastern Korea, 2015.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne disease characterized by fever, thrombocytopenia and diarrhea. SFTS was firstly reported in Korea in 2013 but its seroprevalence in the country has yet to be investigated. Here, we investigate the seroprevalence of SFTS in a Korean population. A cross-sectional study was conducted on patients who had their sera tested for various reasons at a tertiary university hospital on particular days in May 2015. This study was conducted in a tertiary hospital in southeastern Korea. Total antibodies including immunoglobulin G (IgG) and immunoglobulin M (IgM), specific to SFTS virus (SFTSV) in serum samples were detected by a double-antigen sandwich enzyme-linked immunosorbent assay (ELISA). A total of 1,069 serum samples were tested. Median age was 59 years (range 12–96 years), and 51.5% were male. Overall, 22 patients (2.1%) were tested positive for anti-SFTSV antibodies. The SFTS seroprevalence increased significantly with age (P = 0.034). The seropositive rate of rural area was higher than that of urban area (7.7% vs. 1.9%, P = 0.040). Seropositive rates were not significantly different among underlying diseases. None of the antibody-positive patients showed typical symptoms or laboratory findings of SFTS at the time of sample collection. Results of real-time reverse transcription polymerase chain reaction (RT-PCR) were negative for all the seropositive patients. Our study shows 2.1% SFTS seroprevalence among the patients visiting a tertiary hospital in Korea. Seroprevalence is higher in older and rural population.

Seroprevalence of Leptospira spp. in Horses of Distinct Climatic Regions of Punjab, Pakistan

Leptospirosis is zoonotic disease which is globally distributed among many domestic and wild animals. In equines, it causes uveitis, jaundice, and reproductive disorders. A cross-sectional epidemiological study was conducted for the first time in Pakistan to unveil the seroprevalence and to identify associated risk factors of leptospirosis in distinct climatic regions. A total of 384 horse sera (n = 128 from each of three study areas) and data on biologically plausible risk factors along with spatial locations were collected from horses reared in Bahawalpur, Lahore, and Rawalpindi districts of Punjab, Pakistan. Sera were subjected to double-antigen sandwich enzyme-linked immunosorbent assay to analyze the existence or not of Horse LS-IgG in horses. Chi-square, univariate analysis, and multivariate logistic regression were used to analyze the data. Overall, seroprevalence in Punjab was found to be 33.85% (confidence interval [CI]: 29.13–38.83). Significant statistical difference (P < .05) was observed among prevalence in study areas with highest prevalence in Rawalpindi (40.62%; CI: 32.04–49.66), followed by Lahore (38.28%; CI: 29.83–47.28), and lowest in Bahawalpur (22.65%; CI: 15.73–30.89). Age, gender, living area, herd size, water source, rodents, flooding, feeding management and practices, and usage of animals were the significant risk factors associated with the occurrence. This study is first serological evidence of equine Leptospira and its associated risk factors in climatically distinct regions of Punjab, Pakistan.

P08.11 Chlamydia trachomatisincidence from self-reports and serology by age-period, sex and partner numbers in a birth cohort

Background Better understanding of the epidemiology of Chlamydia trachomatis (CT) would assist in prevention and control, but is hindered by asymptomatic infections and analyses based on people tested for clinical reasons that could differ by age and gender. If improved serological detection of CT infection were available, epidemiological studies could more confidently estimate past exposure. We have explored CT incidence by age period in a cohort study, using a combination of a recently characterised serological assay (with higher sensitivity and high persistence) and self-reports. Methods Sexual health and behaviour information was collected from a cohort of initially 1,037 participants born in Dunedin, New Zealand in 1972/3, at regular intervals up to age 38. Sera drawn at ages 26, 32 and 38 were tested for antibodies to CT-specific Pgp3 antigen using a double-antigen sandwich enzyme-linked immunosorbent assay. CT incidence was examined by gender, age and number of partners. Results By age 38, 31.5% (146/464) women and 21.8% (102/469) men had been seropositive and/or self-reported CT infection. More occurred before age 26 than in the 12-year period 26–38 years, the difference being more marked in women than men. In all age periods the risk of acquiring CT increased with number of partners. Once the age-period specific incidence rates were adjusted for the number of partners there was no relationship between CT risk and age period. Overall the partner number adjusted risk was lower in men, although this may reflect that men are less likely to seroconvert than women. Conclusions CT infection was very common amongst this cohort by age 38. Adjusted analyses showed a major risk factor was number of partners, with no interaction by age-period. The increased risk in men must be interpreted cautiously due to the known difference in serological responses between men and women. Disclosure of interest statement This study was funded by Health Research Council of New Zealand. No pharmaceutical grants were received in the development of this study.

Seroprevalence of human T-cell lymphoma/leukemia virus type-1 (HTLV-1) antibodies among blood donors at Enugu, Nigeria

BackgroundHuman T-cell lymphotrophic/leukemia virus (HTLV-1) is a retrovirus implicated in transfusion-transmitted infection.ObjectiveThe objective of this study was to determine the seroprevalence of HTLV-1 antibodies among blood donors at the University of Nigeria Teaching Hospital, Enugu, Eastern Nigeria.MethodsA cross-sectional study was carried out on consented participants over 4 months. A total of 300 blood donors were recruited consecutively from the blood bank. The serum of the collected 5 mL of blood obtained from each participant was stored at −20°C until required for analysis. The serum samples were then analyzed for antibodies to HTLV-1 using a one-step incubation double-antigen sandwich ELISA (enzyme-linked immunosorbent assay) kit. Participants’ demographic characteristics and degree of exposure to the risk factors associated with HTLV-1 infection were captured using a questionnaire. Statistical analysis of results was done using SPSS version 17.ResultsOf the 300 blood donors, 288 (96%) were male, while 12 (4%) were female. The average age of the blood donors was 26.85±8.52 years. The age group with the highest representation among the blood donors were those aged between 21 and 25 years. Only 22.3% of the blood donors were above 30 years. None of the 300 screened blood donors tested positive to HTLV-1 antibodies. Hence, the seroprevalence of HTLV-1 infection among blood donors was 0%. Of the blood donors, 5% had history of previous sexually transmitted disease, while 34.7% used condoms during sexual intercourse.ConclusionThe seroprevalence obtained in this study cannot statistically support the justification of routine screening of blood donors for HTLV-1 infection. More prospective and multicentered studies are required to determine the infectivity of HTLV-1 in blood donors in Nigeria.

Seroprevalence and correlates of human T-cell lymphoma/leukemia virus type 1 antibodies among pregnant women at the University of Nigeria Teaching Hospital, Enugu, Nigeria.

BackgroundHuman T-cell lymphoma/leukemia virus (HTLV)-1 is a retrovirus transmitted vertically from mother to child parenterally and sexually by infected lymphocytes.ObjectiveThe objective of this study was to determine the seroprevalence of HTLV-1 antibodies and associated risk factors for HTLV-1 infection among pregnant women in University of Nigeria Teaching Hospital, Enugu, southeast Nigeria.Materials and methodsA cross-sectional study was carried out from July to October 2010. Two hundred pregnant women were recruited consecutively from the antenatal clinic. Five milliliters of blood was collected from each of the participants into a plain sterile bottle and allowed to clot. The serum obtained was stored at −20°C until required for analysis. The serum samples were then analyzed for antibodies to HTLV-1 using a one-step incubation double-antigen sandwich enzyme-linked immunosorbent assay kit. Participants’ demographic characteristics and degree of exposure to the risk factors associated with HTLV-1 infection were captured using a questionnaire. Statistical analysis of results was done using SPSS version 17.ResultsThe average age of the pregnant women was 28.94 years (standard deviation 4.17). The age-group with the highest representation was those between the ages of 26 and 30 years. Thirty-six percent of the population was above 30 years old. The result of the tests showed that only one respondent, a 31-year-old pregnant woman tested positive for HTLV-1 antibodies. Therefore, the seroprevalence of HTLV-1 antibodies among pregnant women attending the antenatal clinic at University of Nigeria Teaching Hospital was 0.5%, with a 95% confidence interval of 0%–2.8%. Some of the sociodemographic risk factors of HTLV-1 infection found to be applicable to the 31-year-old woman who tested positive included positive history of previous sexually transmitted diseases, high parity, low socioeconomic status, female sex, and age above 30 years. The pregnant women that participated in this study were exposed to risk factors and behaviors associated with HTLV-1 infection. Some of the pregnant women (17.5%) had contracted sexually transmitted diseases, and 80.5% did not use condoms during coitus.ConclusionThe seroprevalence obtained in this study was low, though it is 100% for anyone infected. More prospective and multicenter studies are required to determine the infectivity of HTLV-1 among pregnant women in Nigeria.

A Cross-Sectional Survey of Severe Fever with Thrombocytopenia Syndrome Virus Infection of Domestic Animals in Laizhou City, Shandong Province, China

A serosurvey of severe fever with thrombocytopenia syndrome virus (SFTSV) infection in domestic animals was conducted in the rural areas of Laizhou City, Shandong Province, China to determine strategies for control and prevention of SFTS. Serum samples were collected from cattle, goats, dogs, pigs, and chickens and antibodies against SFTSV were detected by double-antigen sandwich enzyme-linked immunosorbent assay (ELISA). Of 641 serum samples, the SFTSV seropositive rate was 41.8% (268/641): 74.8%, 57.1%, 52.1%, 35.9%, and 0%, for goats, cattle, dogs, chickens, and pigs, respectively. We also found that the SFTSV seropositive rates were high among the aged cattle, goats, dogs, and chickens. SFTSV infections existed among cattle, goats, dogs, and chickens in Laizhou City, and goats had the highest seroprevalence. SFTSV seroprevalence increased with an increase in age among animals. To control of animal infestation with ticks may prevent human SFTSV infections.

Elevated serum interleukin-33 levels in patients with Henoch-Schönlein purpura

Henoch-Schönlein purpura (HSP) is the most common systemic vasculitis and is known as an immunoglobulin (Ig) A related immune complex-mediated disease. However, the molecular mechanisms in the development of HSP are not yet fully understood. Herein, we investigated the serum levels of Interleukin (IL)-33 and soluble ST2 (sST2) in HSP patients and their association with disease severity and IgA autoantibodies production. The serum levels of IL-33 and sST2 were measured by double antibody sandwich enzyme-linked immunosorbent assay (ELISA) in the serum of 33 patients with HSP and 22 controls. Serum levels of IgA anti-endothelial cell antibodies (AECA) and IgA anticardiolipin antibodies (ACA) in HSP patients were detected by double antigen sandwich ELISA. Our results indicated that serum levels of IL-33 but not sST2 were significantly elevated in patients with HSP in acute stage and restored to normal levels in convalescent stage. Moreover, serum IL-33 levels were correlated with the severity of HSP and serum concentrations of AECA-IgA and ACA-IgA. Taken together, we show firstly that serum IL-33 is abnormally elevated in HSP patients. IL-33 might be associated with the IgA autoantibodies production in the pathogenesis of HSP.

Development and Application of a Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay for Detection of Antibodies to Porcine Circovirus 2

A double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) is described for detection of porcine circovirus 2 (PCV2) antibodies using the well-characterized recombinant PCV2 capsid protein. In a comparative test of 394 pig sera against an indirect immunofluorescence (IIF) test and a commercial ELISA kit (also based on the recombinant PCV2 capsid protein), the results showed that the diagnostic sensitivity, specificity, and accuracy of the assay were, respectively, 90.61, 94.02, and 91.62% compared with IIF and 94.38, 95.28, and 94.67% compared with the commercial ELISA kit. Assay of 12 PCV-free pigs over a 5-week period produced only PCV2-negative titers by all 3 methods. These results and the seroprofiles of 4 pig farms obtained by both the commercial ELISA kit and the double-antigen sandwich ELISA indicate that the sandwich ELISA is a reliable method for detection of antibodies to PCV2. Additionally, the method described here permits the use of undiluted test serum samples simultaneously loaded with horseradish peroxidase (HRP)-conjugated antigen into the test well, and the complete test procedure can be performed in less than 90 min. This double-antigen sandwich ELISA should be a useful tool to aid swine industry professionals in deciding the intervention strategies for the control of PCV2-associated diseases.

Preparation and Evaluation of Recombinant Severe Fever with Thrombocytopenia Syndrome Virus Nucleocapsid Protein for Detection of Total Antibodies in Human and Animal Sera by Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay

The recent emergence of the human infection confirmed to be caused by severe fever with thrombocytopenia syndrome virus (SFTSV) in China is of global concern. Safe diagnostic immunoreagents for determination of human and animal seroprevalence in epidemiological investigations are urgently needed. This paper describes the cloning and expression of the nucleocapsid (N) protein of SFTSV. An N-protein-based double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) system was set up to detect the total antibodies in human and animal sera. We reasoned that as the double-antigen sandwich ELISA detected total antibodies with a higher sensitivity than traditional indirect ELISA, it could be used to detect SFTSV-specific antibodies from different animal species. The serum neutralization test was used to validate the performance of this ELISA system. All human and animal sera that tested positive in the neutralization test were also positive in the sandwich ELISA, and there was a high correlation between serum neutralizing titers and ELISA readings. Cross-reactivity was evaluated, and the system was found to be highly specific to SFTSV; all hantavirus- and dengue virus-confirmed patient samples were negative. SFTSV-confirmed human and animal sera from both Anhui and Hubei Provinces in China reacted with N protein in this ELISA, suggesting no major antigenic variation between geographically disparate virus isolates and the suitability of this assay in nationwide application. ELISA results showed that 3.6% of the human serum samples and 47.7% of the animal field serum samples were positive for SFTSV antibodies, indicating that SFTSV has circulated widely in China. This assay, which is simple to operate, poses no biohazard risk, does not require sophisticated equipment, and can be used in disease surveillance programs, particularly in the screening of large numbers of samples from various animal species.

Identification of Natural Bispecific Antibodies against Cyclic Citrullinated Peptide and Immunoglobulin G in Rheumatoid Arthritis

BackgroundPrevious studies indicate that natural bispecific antibodies can be readily produced in vivo when the body is simultaneously stimulated with 2 distinct antigens. Patients with rheumatoid arthritis (RA) usually exhibit persistent immune responses to various autoantigens, raising the possibility that natural bispecific antibodies against 2 distinct autoantigens might exist.Methodology/Principal FindingsWe identified the presence of natural bispecific antibodies against cyclic citrullinated peptide (CCP) and immunoglobulin G (IgG) in RA patients' sera by means of a double-antigen sandwich enzyme-linked immunosorbent assay (ELISA). The spontaneous emergence of bispecific antibodies was confirmed by mixing different proportions of 1 anti-CCP-positive serum and 1 rheumatoid factor (RF)-positive serum in vitro. Among the tested samples, positive correlations were found between the presence of bispecific antibodies and both IgG4 anti-CCP antibodies and IgG4 RF (r = 0.507, p<0.001 and r = 0.249, p = 0.044, respectively), suggesting that the IgG4 subclass is associated with this phenomenon. Furthermore, bispecific antibodies were selectively generated when several anti-CCP- and RF-positive sera were mixed pairwise, indicating that factors other than the monospecific antibody titers may also contribute to the production of the natural bispecific antibodies.Conclusions/SignificanceWe successfully identified the presence of natural bispecific antibodies. Our results suggest that these antibodies originate from anti-CCP and RF in the sera of RA patients. The natural occurrence of bispecific antibodies in human diseases may provide new insights for a better understanding of the diseases. Further investigations are needed to elucidate their precise generation mechanisms and explore their clinical significance in disease development and progression in a larger study population.

A Novel Double-Antigen Sandwich Enzyme-Linked Immunosorbent Assay for Measurement of Antibodies against Rabies Virus

A novel double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) was developed to measure rabies antibodies in dogs. In contrast to the 4 days required for detecting rabies antibody with conventional rabies antibody virus neutralization assays, this ELISA can be completed in hours, without using live virus, in routine laboratories.

Seroepidemiological Study On SARS-CoV IgG Antibodies Of Different Populations From Several Areas

Objective In order to investigate the clinical and epidemiological rules of severe acute respiratory syndrome(SARS), rates and levels SARS coronavirus( SARS-CoV) IgG antibodies of the patients and community populations from several areas were detected . Methods Indirect immunofluorescent assay(IFA) and double-antigen sandwich enzyme-linked immunosorbent assay (ELISA) were used to detect the SARS coronavirus-specific IgG antibodies in sera of 1700, including 1453 general populations from Hongkong, Marco, Guangzhou and Peking and 257 SARS patients from Guangzhou and Peking. The dynamics of the serum antibodies of SARS patients were observed from 3 to 360 days after onset of symptoms. Results 90% of 257 patient serum specimens after 20 days of disease onset showed positive SARS-CoV IgG either using ELISA or IFA . 257 SARS patients, antibodies titers increased steadily in early 4 to 6 months after onset of SARS. The titers of most cases came to the peak in the 6th month. then antibodies titers declined rapidly in some cases. However, all specimens still were positive for SARS-CoV IgG in the 48th month . Conclusions This study suggest that few inapparent infectious patients exist during SARS epidemic. Serum IgG antibodies has diagnostic value for SARS in the late course of disease and the antibodies present more than 48 months.