A field applicable diagnostic technique, the dipstick assay, was evaluated for its sensitivity and specificity in diagnosing human Schistosoma mansoni infection. A monoclonal antibody (mAb) against S. mansoni adult worm tegumental antigen (AWTA) was employed in dipstick and sandwich ELISA for detection of circulating schistosome antigen (CSA) in both serum and urine samples. Based on clinical and parasitological examinations, 60 S. mansoni-infected patients, 30 patients infected with parasites other than schistosomiasis, and 30 uninfected healthy individuals were selected. The sensitivity and specificity of dipstick assay in urine samples were 86.7% and 90.0%, respectively, compared to 90.0% sensitivity and 91.7% specificity of sandwich ELISA. In serum samples, the sensitivity and specificity were 88.3% and 91.7% for dipstick assay vs. 91.7% and 95.0% for sandwich ELISA, respectively. The diagnostic efficacy of dipstick assay in urine and serum samples was 88.3% and 90.0%, while it was 90.8% and 93.3% for sandwich ELISA, respectively. The diagnostic indices of dipstick assay and ELISA either in serum or in urine were statistically comparable (P>0.05). In conclusion, the dipstick assay offers an alternative simple, rapid, non-invasive technique in detecting CSA or complement to stool examinations especially in field studies.
Animals ; Antibodies, Helminth/diagnostic use/isolation & purification ; Antibodies, Monoclonal/diagnostic use/isolation & purification ; Antigens, Helminth/*blood/*urine ; Diagnostic Tests, Routine/*methods ; Humans ; Immunoassay/methods ; Parasitology/*methods ; *Point-of-Care Systems ; Schistosoma mansoni/immunology/*isolation & purification ; Schistosomiasis mansoni/*diagnosis ; Sensitivity and Specificity

OBJECTIVETo prepare and characterize monoclonal antibodies (mAb) and polyclonal antibodies against nucleocapsid (N) protein of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) and to establish antibodies-based sandwich ELISA for detecting N protein of SARS-CoV, which might apply to early diagnosis of patients with SARS-CoV infection.

METHODSBALB/c mice were immunized with purified recombinant N protein of SARS-CoV for producing mAbs, and New Zealand white rabbits were immunized for producing polyclonal antibodies. The identification of antibodies was performed using indirect enzyme-linked immunosorbent assay (ELISA), indirect fluorescent-antibody assay (IFA), and Western immunoblotting. Capturing and detecting antibodies were selected by pairing the mAbs and polyclonal antibodies one by one and an antibodies-based sandwich antigen capture ELISA was used for detecting N antigen of SARS-CoV.

RESULTSNine mAbs and hyperimmune rabbit polyclonal antibodies, specifically against SARS-CoV nucleocapsid protein were obtained. Using paired ELISA assay, three mAbs N1E8, N8E1 and N10E4 were selected as capturing antibody and rabbit polyclonal antibodies as detecting antibody then triple antibodies-based sandwich ELISA was established following horseradish peroxidase (HRP)-conjugated goat anti-rabbit immunoglobulin G. The recombinant N protein was used as a standard to establish a detection sensitivity of approximated 50 pg/ml with this assay. When tested with 420 serum specimens from serologically confirmed SARS patients, the positive rates of serum N protein were 90.1%, 23% and 0%, in which sera collected from 1 to 10 days, 11 to 20 days and beyond 21 days respectively after the onset of symptoms. The specificity of the assay was 99.86% in 715 control serum specimens. There was no cross-reaction with other respiratory viruses and coronaviruses.

CONCLUSIONSpecific and high affinity mAbs and rabbit polyclonal antibodies were obtained. By paired and optimized sandwich ELISA, a sensitive and specific antigen capture ELISA was established for detecting N antigen of SARS-CoV, which might apply to early diagnosis, source tracing and epidemiological studies of SARS.

Animals ; Antibodies, Monoclonal ; biosynthesis ; Antibodies, Viral ; blood ; Enzyme-Linked Immunosorbent Assay ; Humans ; Mice ; Mice, Inbred BALB C ; Nucleocapsid ; immunology ; Rabbits ; SARS Virus ; immunology ; isolation & purification ; Sensitivity and Specificity ; Severe Acute Respiratory Syndrome ; virology

Feline leukemia virus (FeLV) causes a range of neoplastic and degenerative diseases in cats. To obtain a more sensitive and convenient diagnosis of the disease, we prepared monoclonal antibodies specific for the FeLV p27 to develop a rapid diagnostic test with enhanced sensitivity and specificity. Among these antibodies, we identified two clones (hybridomas 8F8B5 and 8G7D1) that specifically bound to FeLV and were very suitable for a diagnostic kit. The affinity constants for 8F8B5 and 8G7D1 were 0.35 x 10(9) and 0.86 x 10(9), respectively. To investigate the diagnostic abilities of the rapid kit using these antibodies, we performed several clinical studies. Assessment of analytical sensitivity revealed that the detection threshold of the rapid diagnostic test was 2 ng/mL for recombinant p27 and 12.5 x 10(4) IU/mL for FeLV. When evaluating 252 cat sera samples, the kit was found to have a kappa value of 0.88 compared to polymerase chain reaction (PCR), indicating a significant correlation between data from the rapid diagnostic test and PCR. Sensitivity and specificity of the kit were 95.2% (20/21) and 98.5% (257/261), respectively. Our results demonstrated that the rapid diagnostic test would be a suitable diagnostic tool for the rapid detection of FeLV infection in cats.
Animals ; Antibodies, Monoclonal/blood ; Cats ; Diagnostic Tests, Routine/*veterinary ; Female ; Gene Products, gag/*blood ; Leukemia Virus, Feline/immunology/*isolation & purification ; Leukemia, Feline/*diagnosis ; Mice, Inbred BALB C ; Sensitivity and Specificity

OBJECTIVE: To observe the clinical effect of plasma exchange and tocilizumab in treatment of patients with severe coronavirus disease 2019 (COVID-19). METHODS: Six patients with severe COVID-19 admitted in First Affiliated Hospital of Bengbu Medical College from January 25 to February 25, 2020. Three patients were treated with plasma exchange and three patients were treated with tocilizumab. The effect on excessive inflammatory reaction of plasma exchange and tocilizumab was observed. RESULTS: The C-reactive protein (CRP) and IL-6 levels were significantly decreased and the lymphocyte and prothrombin time were improved in 3 patients after treatment with plasma exchange; while inflammation level was not significantly decreased, and lymphocyte and prothrombin time did not improve in 3 patients treated with tocilizumab. CONCLUSIONS For severe COVID-19 patients with strong inflammatory reaction, plasma exchange may be preferred.
Antibodies, Monoclonal, Humanized ; administration & dosage ; Betacoronavirus ; isolation & purification ; Coronavirus Infections ; blood ; immunology ; therapy ; Cytokine Release Syndrome ; therapy ; Humans ; Pandemics ; Plasma Exchange ; standards ; Pneumonia, Viral ; blood ; immunology ; therapy ; Prothrombin Time ; Treatment Outcome

OBJECTIVETo construct and express anti-human RBC and HIVgp160 fusion protein for rapid detection of antibody to HIV.

METHODSThe gene of the anti human RBC ScFv and HIV antigen were constructed together into expression vector. The fusion protein was expressed in E. coli.

RESULTSThe fusion protein was proved to be able to bind both anti-RBC and HIVgp160. It could cause agglutination of human RBC when HIVgp160 was present.

CONCLUSIONThe fusion protein has the potential in rapid detection of HIV.

Antibodies, Monoclonal ; immunology ; isolation & purification ; Autoantibodies ; immunology ; isolation & purification ; Cloning, Molecular ; Enzyme-Linked Immunosorbent Assay ; Erythrocytes ; immunology ; Gene Expression ; Genetic Vectors ; genetics ; HIV Antibodies ; blood ; immunology ; HIV Envelope Protein gp160 ; genetics ; immunology ; metabolism ; HIV Seropositivity ; blood ; Hemagglutination Tests ; methods ; Humans ; Recombinant Fusion Proteins ; genetics ; immunology ; metabolism

OBJECTIVETo prepare a single chain antibody (ScFv) of mAb SZ-21 against platelet GPIIIa for its future clinical application.

METHODSThe expression vector pET20b-SZ-21ScFv was constructed and the fusion protein was expressed in E. coli BL21 (DE3) PlysS. The activated fusion protein was obtained after a series of purification steps, including cell breakage, inclusion body solubilization, His-bind resin affinity chromatography and protein refolding.

RESULTSThe fusion protein yields were up to 21% of the total amount of bacteria protein. The ScFv fragment could inhibit ADP-induced platelets aggregation in a dose-dependent manner in vitro and the maximal inhibition rate was obtained at a concentration of 20 micro g/ml. It also reacted with endothelial cells as detected by flow cytometry. Moreover, the ScFv fragment was able to inhibit the binding of fibrinogen to platelet.

CONCLUSIONThe SZ-21ScFv fragment had the activity to inhibit platelets aggregation and the binding of fibrinogen to platelet, being potentially useful for the treatment of thrombotic diseases.

Antibodies, Monoclonal ; pharmacology ; Blood Platelets ; metabolism ; Endothelium, Vascular ; cytology ; Fibrinogen ; metabolism ; Humans ; Immunoglobulin Fragments ; pharmacology ; Platelet Aggregation ; Platelet Glycoprotein GPIIb-IIIa Complex ; immunology ; Recombinant Fusion Proteins ; isolation & purification ; pharmacology

This study was purposed to prepare eukaryotic expression vector of recombinant human platelet CD36 gene. The total RNA was extracted from human liver tissue and the cDNA encoding human platelet CD36 antigen extracellular region (Gly30-Asn439) was amplified by RT-PCR. The cDNA was cloned into the prokaryotic expression vector pMD18 and the recombinant vector was transformed into E. coli DH5α. The positive recombinant pMD18-CD36 plasmid was screened. After sequencing, this combinant vector was inserted into the transient eukaryotic expression vector pTE2, the pTE2-s-CD36-10 His transient eukaryotic expression vector was constructed. The recombinant CD36 Gly30-Asn439 expressed by HEK-293 cells was purified with Ni(2+) 2NTA chromatography. The results showed that 1.4 kb cDNA was amplified by RT-PCR, sequencing of the cDNA indicated the sequence was exactly the same to that in Genbank NM_001001547.2. The HEK293 cells with the plasmid were transfected, and SDS-PAGE confirmed that the transfect HEK293 cells expressed the human CD36 antigen extracellular protein fragments. Western-blot showed that the monoclonal antibody could recognize the recombinant CD36 with the sensitivity of 8 ng. It is concluded that the CD36 Gly30-Asn439 can be highly expressed by human embryonic kidney cells (HEK293), and the monoclonal antibody with biological activity has been obtained, which provide the basis for further study on platelet transfusion refractoriness.
Animals ; Antibodies, Monoclonal ; genetics ; isolation & purification ; Blood Platelets ; immunology ; Blotting, Western ; CD36 Antigens ; genetics ; immunology ; DNA, Complementary ; Genetic Vectors ; Humans ; Mice ; Plasmids ; Platelet Transfusion ; Reverse Transcriptase Polymerase Chain Reaction

Thirty-nine healthy pigs (28-32 days old) were purchased from a commercial swine farm and housed at swine pens of the College. The animals were vaccinated intramuscularly (1 ml) with an attenuated live hog cholera virus (HCV, LOM strain) and then boostered at 5 weeks after the first vaccination. The animals were divided into 4 experimental groups: 0.05% (w/w) PowerFeel-supplemented diet (T-1, n = 10); 3% (w/w) SuperFeed-supplemented diet (T-2, n = 10); diluted PowerFeel solution (1 : 500, v/v) as drinking water (T-3, n=9); control (n=10). PowerFeel is an original form of ionized alkali mineral complex (IAMC) and SuperFeed is a commercial product of IAMC. The subpopulation of lymphocyte in blood was assayed by a flow cytometry and HCV-specific antibody was determined by an indirect immunofluorescence assay. In IMAC-treated groups, the proportions of subpopulation expressing MHC-class II, CD2+, CD4+, CD8+, and surface IgM+ B lymphocytes were significantly decreased at 5-weeks after the first vaccination. Significant decreases were also observed in the proportions of MHC-class II, CD2+ and CD8+ lymphocyte at 3-weeks after the booster injection. The humoral immune responses in T-1 and T-2 groups were greater than those in T-3 or control group. These results suggest that IAMC-supplemented diets may have an HCV-specific immunostimulatory effect in pigs.
Animal Feed ; Animals ; Antibodies, Monoclonal/*blood/isolation & purification ; Antigens, CD2/blood ; B-Lymphocytes/immunology ; CD4-Positive T-Lymphocytes/immunology ; CD8-Positive T-Lymphocytes/immunology ; Classical Swine Fever/*immunology ; Classical swine fever virus/*immunology ; Dietary Supplements ; Ions ; Lymphocyte Subsets/*immunology ; *Minerals ; Swine ; Vaccines, Attenuated/*administration & dosage ; Viral Vaccines/*administration & dosage

This study measured the clinical prevalence of peste des petits ruminants (PPR) among sheep and goats in India between 2003 and 2009 by analyzing clinical samples from suspected cases of PPR that were submitted to the Rinderpest and Allied Disease Laboratory, Division of Virology, IVRI, Mukteswar for PPR diagnosis. PPR outbreaks were confirmed by detecting PPR virus (PPRV)-specific antigen in the clinical samples. Clinical samples (blood, nasal swabs, spleen, lymph node, kidney, liver, intestine, and pooled tissue materials) were taken from a total of 592 sheep and 912 goats in different states of India and screened for the presence of PPRV antigen using a monoclonal antibody-based sandwich ELISA kit. A total of 20, 38, and 11 laboratory-confirmed PPR outbreaks occurred among sheep, goat, and combined sheep and goat populations, respectively. Our findings provide evidence of widespread PPR endemicity in India. The underlying reasons could be variations in husbandry practices in different geographical regions, agro-climatic conditions, and livestock migration. Furthermore, decrease in the number of PPR outbreaks over time might be due to the effectiveness of current live PPR vaccines and timely vaccination of target species. Vaccination against PPR has been practiced in India since 2002 to control this disease.
Animals ; Antibodies, Monoclonal/immunology ; Antigens, Viral/*blood ; Disease Outbreaks/*veterinary ; Enzyme-Linked Immunosorbent Assay/veterinary ; Goat Diseases/*epidemiology/immunology/prevention & control ; Goats ; India/epidemiology ; Nucleocapsid Proteins/immunology ; Peste-des-Petits-Ruminants/epidemiology/immunology/prevention & control/*veterinary ; Peste-des-petits-ruminants virus/*immunology/isolation & purification ; Prevalence ; Risk Factors ; Seasons ; Sheep ; Sheep Diseases/*epidemiology/immunology/prevention & control ; Vaccination/veterinary ; Viral Vaccines/*immunology/therapeutic use

BACKGROUND/AIMS: The relationship between patient survival and biopsy-proven acute rejection (BPAR) in liver transplant recipients with hepatitis C remains unclear. The aims of this study were to compare the characteristics of patients with and without BPAR and to identify risk factors for BPAR. METHODS: We retrospectively reviewed the records of 169 HCV-RNA-positive patients who underwent LT at three centers. RESULTS: BPAR occurred in 39 (23.1%) of the HCV-RNA-positive recipients after LT. The 1-, 3-, and 5-year survival rates were 92.1%, 90.3%, and 88.5%, respectively, in patients without BPAR, and 75.7%, 63.4%, and 58.9% in patients with BPAR (P<0.001). Multivariate analyses showed that BPAR was associated with the non-use of basiliximab and tacrolimus and the use of cyclosporin in LT recipients with HCV RNA-positive. CONCLUSION: The results of the present study suggest that the immunosuppression status of HCV-RNA-positive LT recipients should be carefully determined in order to prevent BPAR and to improve patient survival.
Antibodies, Monoclonal/therapeutic use ; Biopsy ; Cyclosporine/therapeutic use ; Drug Therapy, Combination ; Genotype ; Graft Rejection/mortality/*prevention & control ; Hepacivirus/genetics/isolation & purification ; Hepatitis C/drug therapy/*virology ; Humans ; Immunosuppressive Agents/*therapeutic use ; *Liver Transplantation/adverse effects ; Polymerase Chain Reaction ; RNA, Viral/blood ; Recombinant Fusion Proteins/therapeutic use ; Recurrence ; Retrospective Studies ; Survival Rate ; Tacrolimus/therapeutic use