High Enzyme-linked Immunosorbent Assay Titers Research Articles

Anti-double stranded DNA antibodies: A rational diagnostic approach in limited-resource settings

ContextAnti-double-stranded deoxyribonucleic acid antibodies (dsDNA Abs) are highly specific markers of systemic lupus erythematosus (SLE). Multiple methods are employed for their detection in routine diagnostics. ObjectivesThe aim of this study was to evaluate a diagnostic approach for anti-dsDNA Abs using DNA-ELISA and Crithidia luciliae fluorescence test (CLIFT), in combination with antinuclear antibody (ANA) screening. MethodsWe enrolled 113 patients—53 with SLE, 50 with other systemic autoimmune rheumatic diseases (OSARD), and 10 with non-autoimmune clinical conditions (NAICC).Patients’ samples were tested for anti-dsDNA Abs using an enzyme-linked immunosorbent assay (ELISA) and CLIFT, combined to ANA screening by indirect immunofluorescence assay (ANA-IIFA). ResultsThe mean age of patients was 39.94 ± 15 years (ranges: 11–85 years). Overall, specimens from 77.3%, 11.7%, and 20% of patients with SLE, OSARD and NAICC respectively were ELISA-positive; and those from 54.7% to 4% of patients with SLE and OSARD, respectively, were CLIFT-positive. CLIFT positivity was significantly associated with high ELISA titers (p = 0.002) and homogeneous ANA-IIF pattern (p = 0.0002). ConclusionFor better clinical relevance of anti-dsDNA antibodies, we suggest a combined detection strategy based on ELISA, CLIFT and ANA-IIFA, considering the clinical criteria of SLE.

Human polyclonal immunoglobulin G from transchromosomic bovines inhibits MERS-CoV in vivo.

As of 13 November 2015, 1618 laboratory-confirmed human cases of Middle East respiratory syndrome coronavirus (MERS-CoV) infection, including 579 deaths, had been reported to the World Health Organization. No specific preventive or therapeutic agent of proven value against MERS-CoV is currently available. Public Health England and the International Severe Acute Respiratory and Emerging Infection Consortium identified passive immunotherapy with neutralizing antibodies as a treatment approach that warrants priority study. Two experimental MERS-CoV vaccines were used to vaccinate two groups of transchromosomic (Tc) bovines that were genetically modified to produce large quantities of fully human polyclonal immunoglobulin G (IgG) antibodies. Vaccination with a clade A γ-irradiated whole killed virion vaccine (Jordan strain) or a clade B spike protein nanoparticle vaccine (Al-Hasa strain) resulted in Tc bovine sera with high enzyme-linked immunosorbent assay (ELISA) and neutralizing antibody titers in vitro. Two purified Tc bovine human IgG immunoglobulins (Tc hIgG), SAB-300 (produced after Jordan strain vaccination) and SAB-301 (produced after Al-Hasa strain vaccination), also had high ELISA and neutralizing antibody titers without antibody-dependent enhancement in vitro. SAB-301 was selected for in vivo and preclinical studies. Administration of single doses of SAB-301 12 hours before or 24 and 48 hours after MERS-CoV infection (Erasmus Medical Center 2012 strain) of Ad5-hDPP4 receptor-transduced mice rapidly resulted in viral lung titers near or below the limit of detection. Tc bovines, combined with the ability to quickly produce Tc hIgG and develop in vitro assays and animal model(s), potentially offer a platform to rapidly produce a therapeutic to prevent and/or treat MERS-CoV infection and/or other emerging infectious diseases.

Seroprevalence of Aichi Virus in a Spanish Population from 2007 to 2008

Viruses are among the most common causes of acute gastroenteritis. In recent years, new viruses causing outbreaks of acute gastroenteritis have been described. Among these, Aichi virus was identified in Japan in 1989. Aichi virus belongs to the Kobuvirus genus in the family Picornaviridae. This virus has been detected in outbreaks of gastroenteritis associated with oyster consumption and in pediatric stool samples, but little is known about its epidemiology or pathogenesis. In the present study, the prevalence of antibodies to Aichi virus in a Spanish population was determined between 2007 and 2008 by using an enzyme-linked immunosorbent assay (ELISA). As in previous studies, a high seroprevalence of antibodies to Aichi virus (70%) was observed, with levels differing according to age. We observed significant differences in titers of antibody to Aichi virus among different age groups, grouped by decades. We report high ELISA and neutralizing antibody titers, and both titers fitted a sigmoid curve significantly. However, this virus is seldom detected; therefore, further studies are needed to gain a better understanding of its importance as a pathogenic agent.

Survey of Japanese Layer Farms for Salmonella Enteritidis with Vaccination- and Infection-Specific Antigens for Egg Yolk Antibodies

Japanese layer farms were surveyed for Salmonella Enteritidis vaccination and infection with specific antigens for egg yolk antibodies with the use of vaccination-specific antigen Salmonella Enteritidis FliC-specific 9-kDa polypeptide (SEP9) and infection-specific antigen deflagellated Salmonella Enteritidis whole cell (DEWC). The specific antibodies in eggs from 201 commercial layer farms throughout Japan were surveyed. The percentages of farm flocks with a mean enzyme-linked immunosorbent assay (ELISA) titer of over 0.1 were 56.2% (113 of 201) in DEWC-ELISA and 22.3% (45 of 201) in SEP9-ELISA. Flocks indicating high titers in SEP9-ELISA always showed high titers in DEWC-ELISA. Because both specific antibody titers of the vaccinated flocks monitored long term remained high throughout life, flocks with high titers of both ELISAs in this survey must be vaccinated. On the other hand, 34.3% (69 of 201) of flocks had high titers of DEWC-specific antibody alone. Because Salmonella Enteritidis infection induces the DEWC-specific antibody but not the SEP9-specific antibody, detecting only high ELISA titers of DEWC-specific antibody can be an effective monitoring tool for Salmonella Enteritidis exposure rather than vaccination. These results suggest that vaccination programs in Japanese layer farms would be insufficient to control Salmonella Enteritidis infection, and egg screening to detect specific antibodies would be valuable in obtaining the necessary information to control Salmonella Enteritidis infection.

Cloning and expression of the guinea pig cytomegalovirus glycoprotein B (gB) in a recombinant baculovirus: utility for vaccine studies for the prevention of experimental infection

The guinea pig cytomegalovirus (GPCMV) is unique among the cytomegaloviruses of small mammals, insofar as during pregnancy it crosses the placenta, causing infection of the fetus. Although the guinea pig model is well suited to vaccine studies, the lack of cloned, recombinant forms of immunogenic GPCMV proteins, such as envelope glycoproteins, has hindered experimental evaluations of subunit immunization for prevention of fetal disease. Since the glycoprotein B (gB) is a major target of neutralizing antibody responses, the GPCMV gB was cloned and expressed in a recombinant baculovirus. A recombinant was generated which expressed gB, truncated at codon 692, upstream of the putative transmembrane domain. Processing and expression of the recombinant protein, designated Bac-gB, was assessed, and the protein was characterized immunologically. Anti-gB antibodies were immunoreactive with Bac-gB by enzyme linked immunosorbent assay (ELISA) and immunoblot assay. Immunoprecipitation with polyclonal anti-GPCMV antisera identified protein species of 120, 80 and 30 kDa by reducing SDS-PAGE, suggesting that authentic cleavage and processing of Bac-gB occurred in insect cells. Sera from guinea pigs immunized with lectin-column purified native glycoproteins had high ELISA titers to Bac-gB. Recombinant GPCMV gB expressed in insect cells should prove useful in defining correlates of protective immunity in the GPCMV congenital infection model.

Efficiency of protection of guinea pigs against infection with Bacillus anthracis spores by passive immunization.

The efficacy of passive immunization as a postexposure prophylactic measure for treatment of guinea pigs intranasally infected with Bacillus anthracis spores was evaluated. Antisera directed either against the lethal toxin components (PA or LF) or against a toxinogenic strain (Sterne) were used for this evaluation. All antisera exhibited high enzyme-linked immunosorbent assay titers against the corresponding antigens, high titers of neutralization of cytotoxicity activity in an in vitro mouse macrophages cell line (J774A.1), as well as in vivo neutralization of toxicity when administered either directly to Fisher rats prior to challenge with the lethal toxin or after incubation with the lethal toxin. In these tests, anti-LF antiserum exhibited the highest neutralization efficiency, followed by anti-Sterne and anti-PA. The time dependence and antibody dose necessary for conferring postexposure protection by the various antibodies of guinea pigs infected with 25 50% lethal doses of Vollum spores was examined. Rabbit anti-PA serum was found to be the most effective. Intraperitoneal injections of anti-PA serum given 24 h postinfection protected 90% of the infected animals, whereas anti-Sterne and anti-LF were less effective. These results further emphasizes the importance of anti-PA antibodies in conferring protection against B. anthracis infection and demonstrated the ability of such antibodies to be effectively applied as an efficient postexposure treatment against anthrax disease.

Canine paracoccidioidomycosis: a seroepidemiologic study

Sera from 305 dogs were analyzed by enzyme-linked immunosorbent assay (ELISA) to determine presence of the antibody anti-gp43, which reacts to a specific antigen of Paracoccidioides brasiliensis. The dogs were divided into three groups according to their origin: urban dogs (animals with little or no contact with rural areas); suburban dogs (from the urban outskirts); and rural dogs. There was a significant difference between groups (P < 0.05). Rural dogs reacted positively in 89.5% of cases, followed by suburban (48.8%) and urban dogs (14.8%). There were no differences between male and female dogs. In an attempt to verify the feasibility of skin testing with gp43 to determine sensitization against P. brasiliensis in dogs, suburban (n = 61) and rural (n = 21) dogs were tested, showing positivity of 13.1 and 38.1%, respectively. Six dogs that had higher ELISA titers and also showed strong reactions in skin testing were killed in an attempt to isolate P. brasiliensis. The fungus was not detected by culture or histopathological analysis in these dogs, suggesting that dogs have a natural resistance or that they encounter an inoculum level that is insufficient to cause disease. These results indicate that ELISA and skin testing can be useful in the epidemiological study of paracoccidioidomycosis in dogs and that encounter with the fungus in nature is a frequent event.

Effect of Route of Vaccination on the Prevention of Infectious Laryngotracheitis in Commercial Egg-Laying Chickens

Commercial egg-laying chickens were vaccinated for infectious laryngotracheitis (ILT) with one of five commercially available vaccines (designated A, B, C, D, and E) on five separate farms by either eyedrop (e), spray (s), or double dose in the water (w) method. Groups were identified by the vaccine designation and the method of vaccination. Birds from the test groups were transferred to an isolation facility and challenged intratracheally 3 wk after vaccination. The remaining birds were given a second vaccination with the original chicken embryo origin vaccine by spray or a chicken embryo origin vaccine if the first vaccine was of tissue culture origin. After challenge, birds were monitored for clinical signs. Those surviving were euthanatized on day 6 postchallenge, and tissues and blood were collected for histopathology, virus isolation, and serology. On the basis of histopathology and enzyme-linked immunosorbent assay (ELISA) results, after one vaccination, all chickens given vaccines by eyedrop were provided better protection than nonvaccinated controls (CTLs). Birds in groups Bs and Ds had lower microscopic lesion scores whereas only birds given Bs had higher ELISA titers than CTLs. Birds in groups As and Cs and groups Bw birds taken from the rear of the barn (r) had microscopic lesion scores that were no different from those of CTLs. These same birds in addition to vaccine Ds had ELISA titers no different from those of CTLs. Of all vaccines, only A given by eyedrop or spray produced higher virus isolation titers than those of CTLs. The remainder of the vaccines produced virus isolation titers that were no different from those of CTLs. After two vaccinations, all groups had lower microscopic lesion scores than CTLs. Only Bw birds from the middle of the barn Bs, EeDs, and AsAs had virus isolation results that were higher than those of CTLs. Only groups BwrBs, CsCs, and DsDs had ELISA titers no different from those of controls. These results suggest that a priming vaccination followed by a booster dose offers better protection against ILT than a single vaccination alone. Vaccine application by eyedrop provides more uniform protection if only one vaccination is given, whereas spray vaccination may serve as an alternative method of vaccination for birds receiving two doses of vaccine.

Evaluation of culture enrichment procedures for use with Salmonella detection immunoassay

To design efficient culture strategies for use with immunoassays to detect Salmonella in food, the growth of these organisms was investigated according to the Bacteriological Analytical Manual (BAM) and enrichment-immunoassay (EI) culture procedures. The cultures were further evaluated using a commercial enzyme-linked immunosorbent assay (ELISA) kit . The BAM procedure includes pre-enrichment in nutrient broth (NB) for 16 h followed by selective enrichment in either Rappaport–Vassiliadis (RV) or tetrathionate brilliant green (TBG) broth for 16 h. The EI procedure includes pre-enrichment in NB for 4 h, selective enrichment in RV for 16 h and post-enrichment in NB for 4 h. The effects of different incubation times for pre- and post-enrichment, and different culture media for selective enrichment (TBG and RV) and post-enrichment in NB and Brain Heart Infusion broth (BHI) on the growth of the bacteria and ELISA titers in the EI procedure were also investigated. Salmonella enteritidis and S. typhimurium inoculated at different initial concentrations between 0.1 and 35 CFU/ml grew to similar concentrations of 10 7 to 10 8 colony forming unit (CFU)/ml in pure culture and generally 2 to 4 fold lower concentrations ( P<0.05) in mixed culture using spiked chicken rinse. In the BAM procedure, the concentration of Salmonella cultured in RV was higher ( P<0.01) than that in TBG. The cultures in TBG showed positive results for ELISA, but those in RV were generally negative. In the EI procedure, the ELISA titers from cultures post-enriched in NB or BHI were higher ( P<0.01) when TBG, as compared to RV, was used for selective enrichment. Post-enrichment in BHI yielded higher numbers of Salmonella and higher ELISA titers than those in NB ( P<0.05) for post-enrichment. This study demonstrated that in both culture procedures small numbers of Salmonella could be increased to at least 10 7 CFU/ml which is detectable by most ELISAs, and that the type of the culture media used may have a significant impact on ELISA results.

Fibrinogen cleavage by the Streptococcus pyogenes extracellular cysteine protease and generation of antibodies that inhibit enzyme proteolytic activity.

The extracellular cysteine protease from Streptococcus pyogenes is a virulence factor that plays a significant role in host-pathogen interaction. Streptococcal protease is expressed as an inactive 40-kDa precursor that is autocatalytically converted into a 28-kDa mature (active) enzyme. Replacement of the single cysteine residue involved in formation of the enzyme active site with serine (C192S mutation) abolished detectable proteolytic activity and eliminated autocatalytic processing of zymogen to the mature form. In the present study, we investigated activity of the wild-type (wt) streptococcal protease toward human fibrinogen and bovine casein. The former is involved in blood coagulation, wound healing, and other aspects of hemostasis. Treatment with streptococcal protease resulted in degradation of the COOH-terminal region of fibrinogen alpha chain, indicating that fibrinogen may serve as an important substrate for this enzyme during the course of human infection. Polyclonal antibodies generated against recombinant 40- and 28-kDa (r40- and r28-kDa) forms of the C192S streptococcal protease mutant exhibited high enzyme-linked immunosorbent assay titers but demonstrated different inhibition activities toward proteolytic action of the wt enzyme. Activity of the wt protease was readily inhibited when the reaction was carried out in the presence of antibodies generated against r28-kDa C192S mutant. Antibodies produced against r40-kDa C192S mutant had no significant effect on proteolysis. These data suggest that the presence of the NH(2)-terminal prosegment prevents generation of functionally active antibodies and indicate that inhibition activity of antibodies most likely depends on their ability to bind the active-site region epitope(s) of the protein.

Jararhagin ECD-Containing Disintegrin Domain: Expression in Escherichia coli and Inhibition of the Platelet–Collagen Interaction

Jararhagin, a hemorrhagin from Bothrops jararaca venom, is a soluble snake venom component comprising metalloproteinase and disintegrin cysteine-rich domains and, therefore, is structurally closely related to the membrane-bound ADisintegrin And Metalloproteinase (ADAMs) protein family. Its hemorrhagic activity is associated with the effects of both metalloproteinase and disintegrin domains; the metalloproteinase enzymatically damages the endothelium and the disintegrin domain inhibits platelet–collagen interactions. The expression of whole jararhagin or its disintegrin domain has never been attempted before. The aim of this study was to investigate whether we could express the disintegrin domain of jararhagin and to verify whether this domain displays an inhibitory effect on the platelet–collagen interaction. Therefore, the cDNA fragment coding for the disintegrin plus cysteine-rich domains of jararhagin was cloned into the pET32a vector, used to transform the Escherichia coli AD494(DE3)pLysS strain. The thioredoxin–disintegrin fusion protein was recovered from the soluble extract of the cells, yielding up to 50 mg/liter culture. The fusion protein was isolated using polyhistidine binding resin which resulted in a main band of 45 kDa recognized by anti-native jararhagin antibodies. Antibodies raised in rabbits against the fusion protein had high enzyme-linked immunosorbent assay titers against native jararhagin and detected a band of 52 kDa on Western blots of whole B. jararaca venom demonstrating that these antibodies recognize the parent jararhagin molecule. Treatment of the fusion protein with enterokinase, followed by further capture of the enzyme, resulted in a band of 30 kDa, the expected size for jararhagin-C. Further purification of the cleaved disintegrin using FPLC Mono-Q columns resulted in one fraction capable of efficiently inhibiting collagen-induced platelet aggregation in a dose-dependent manner (IC50 of 8.5 μg/ml).

Sexual Transmission of the Hepatitis C Virus and Efficacy of Prophylaxis With Intramuscular Immune Serum Globulin

To estimate the risk of sexual transmission of hepatitis C and to assess the value of prophylaxis with periodic intramuscular immune serum globulin administration. Of 1102 steady heterosexual partners of patients with antibodies to the hepatitis C virus (HCV), 899 were enrolled in a single-blind, randomized, controlled trial. All the partners tested negative for antibodies to HCV and had normal baseline serum aminotransferase concentrations. The partners were assigned to receive 4 mL of 16% polyvalent immune serum globulin prepared from unscreened donors every 2 months (n = 450) or a placebo (n = 449). Tests for HCV infection were performed every 4 months. Eight hundred eighty-four partners completed the study. Seven partners became infected with HCV: 6 in the control group (incidence density, 12.00 per 1000 person-years; 95% confidence interval, 3.0 21.61) and 1 in the immune serum globulin group (incidence density, 1.98 per 1000 person-years; 95% confidence interval, 0-5.86). The risk of infection was significantly higher for partners in the control group (P = .03): for each year approximately 1% of the partners became infected. Sequence homology studies strongly suggest the sexual transmission of HCV. All immune serum globulin lots used had high enzyme-linked immunosorbent assay titers of neutralizing antibodies to HCV envelope glycoproteins and high neutralization titers in the neutralization of binding assay. Hepatitis C can be sexually transmitted. Immune serum globulin prepared from unscreened donors significantly reduced the risk. The treatment was safe and well tolerated. Because only immune serum globulin from unscreened donors (and not from those screened for HCV) contain anti-HCV neutralizing antibodies, hyperimmune anti-HCV immune serum globulin should be prepared from blood testing positive for antibodies to HCV, which is currently discarded.

Serological survey of canine distemper virus infection using enzyme-linked immunosorbent assay.

For detection of antibodies against canine distemper virus (CDV), an enzyme-linked immunosorbent assay (ELISA) was developed using a crude extract from cells infected with the Onderstepoort strain of CDV as antigen. Twenty-six sera from dogs experimentally vaccinated with the Snyder-Hill strain of CDV were compared by ELISA and a standard virus neutralization (NV) test. Since a good correlation between the titers obtained by both tests was observed, ELISA was considered to be a rapid and reliable method for a serological survey of CDV infection. When a total of 167 sera from dogs suspected of canine distemper under natural conditions were examined by the ELISA, 29 of the sera (17%) were found to have low VN antibody titers and high ELISA titers. The reason for the discrepancy in the titers was discussed.

Characterization of monoclonal antibodies to bovine herpesvirus type I, Los Angeles strain

We produced monoclonal antibodies (mAbs) against bovine herpesvirus type 1 (BHV-1), Los Angeles strain, and then evaluated them as potential candidates for preparing diagnostic reagents. Of the 318 cell lines expressing antibodies to the virus, 60% (192)_secreted IgG and 40% (126) secreted IgM. Twenty-six mAbs were selected based on enzyme-linked immunosorbent assay (ELISA) and virus neutralization (VN) titres and characterized by immunoprecipitation, immunofluorescence and immunoblots. The selected mAbs were assigned to one of groups based on their immunoprecipitation patterns. Group A (4 mAbs) precipitated a complex of three glycoproteins with molecular weight (MW) 130 kDa, 72 kDa and 55 kDA, which presumably represented gI of BHV-1. Monoclonal antibodies of this group were highly reactive in ELISA but had low VN titers. Group B (4 mAbs) precipitated a glycoprotein with MW of 71 kDa (gIV). This group of mAbs had high VN titers. Group Cc (16 mAbs) precipitated a 97 kDa glycoprotein (gIII). Monoclonal antibodies of this group had high ELISA but low VN titers. Group D (2 mAbs) prepicitated a double band of non-glycolysated proteinss with MW of 37 32 kDa; these proteins could not be assigned to any of the antigens of BHV-1 previously described. ELISA and VN titers of this group of mAbs were low. To test the antigenic variability of the antigenic determinants which were recognized by these 4 groups of mAbs, we adapted Madin Darby bovine kidney cell-propagated BHV-1 Los Angeles strain to Crandell's feline kidney cell line. After the tenth passage in feline kidney cells, the epitopes on the 37 32 kDa peptide recognized by the mAbs group D were no longer detectable. Additional changes were noted in the electrophoretic mobility of the 130 kDa and 71 kDa glycoproteins (gI) identified by mAb of group A shifted downward. The 71 kDa glycoprotein (gIV) reactive with mAb group B and the 97 kDa (gIII) reactive with mAb group C remained stable. Since clone No. 191 of group B mAb was protein in ELISA, VN, immunoblots and immunofluorescence, and recognized an epitope which did not change under selective pressure, we feel that the mAb produced by this clone are a good candidate for the production of diagnostic reagents.

Polyneuropathy syndromes associated with serum antibodies to sulfatide and myelin‐associated glycoprotein

We studied a series of 64 patients with sensory +/- motor peripheral neuropathies by comparing clinical and physiologic features to serum antibody reactivity against compounds containing sulfated carbohydrate moieties. We determined antibody reactivity by an enzyme-linked immunosorbent assay (ELISA) using purified glycolipids and glycoproteins as antigens, and we used high-performance thin-layer chromatography and Western blotting to test the specificity of results. Twelve patients with high titers of IgM antibodies directed against the myelin-associated glycoprotein (MAG) had sensory-motor polyneuropathies with physiologic evidence of demyelination. IgM antibody reactivity to MAG was associated with an IgM serum M protein in five patients. Eight other patients, most with sensory greater than motor polyneuropathies, had high titers of antibody reactivity to sulfatide but not of IgM to MAG. Two had an associated IgM paraprotein. None of the patients with selective serum antisulfatide activity had predominantly demyelinating features on physiologic testing. We conclude that (1) high ELISA titers of antibodies to MAG may be more common than previously suspected in patients with chronic demyelinating sensory-motor neuropathies, and (2) the presence of high titers of antisulfatide antibodies in serum may provide clues to the pathogenesis of otherwise idiopathic, axonal, predominantly sensory neuropathies.

Serum immune response to Shigella protein antigens in rhesus monkeys and humans infected with Shigella spp.

The serum antibody response to proteins encoded by the virulence-associated plasmid of Shigella flexneri was determined in monkeys challenged with virulent S. flexneri serotype 2a. With water-extractable antigen in an enzyme-linked immunosorbent assay, a significant increase in antibody titer against proteins from a plasmid-carrying, virulent strain of S. flexneri serotype 5 could be demonstrated in convalescent sera. There were minimal antibody titers against proteins from an avirulent (plasmid-free) organism. Previously identified plasmid-coded polypeptides a, b, c, and d were predominant antigens recognized by a majority of the convalescent sera in immunoblots. An additional 140-megadalton plasmid-coded polypeptide was also recognized by half of the sera. Convalescent serum from an infected monkey recognized antigens on the bacterial surface in several different plasmid-containing Shigella species and in an enteroinvasive Escherichia coli strain. A survey of sera obtained from children 5 to 10 years of age who had been infected with S. flexneri or S. sonnei revealed high enzyme-linked immunosorbent assay titers in both acute and convalescent sera against a water extract from a virulent Shigella strain. In contrast, children under 3 years of age had no antibody titer in either acute or convalescent sera against the virulence-associated shigella proteins, while 3- to 4-year-old children mounted an immune response against these proteins only in convalescence.

A glycopolypeptide (gp 100) is the main antigen detected by HTLV-III antisera.

Sera from homosexuals and hemophiliacs in Germany were examined for antibodies to human T-lymphotropic retrovirus type III (HTLV-III) by an enzyme-linked immunosorbent assay (ELISA) against purified virus. ELISA positive sera were used to search by immunoprecipitation for HTLV-III related antigens in a persistently infected human T-cell line. A glycopolypeptide with a mol. wt. of 100 000 was regularly recognized by all positive sera. In analogy to glycosylation and high antigenicity of envelope polypeptides of other mammalian retroviruses, gp 100 seemed to be related to the env gen. Two polypeptides with mol. wts. of 24 000 and 22 000 probably representing viral core polypeptides were additionally detected by sera with high ELISA titers.

Evaluation of an enzyme-linked immunosorbent assay specific for human immunoglobulin G as a Screening test for detecting anti-toxoplasma antibodies.

An evaluation of the enzyme-linked immunosorbent assay (ELISA) as a screening test for immunoglobulin G class-specific anti-Toxoplasma antibodies was conducted. Results for 191 serum specimens assayed by the ELISA, indirect hemagglutination (IHA), and methylene blue dye tests indicated that good qualitative agreement exists among these tests. High ELISA titers relative to methylene blue dye titers may be attributed in part to the presence of rheumatoid factor. ELISA results were found to be reproducible. A parallel study of 1,760 sera assayed by ELISA and IHA was performed with 3.0-mm-diameter serum filter paper spots as a source of specimen. When a 1:64 titer and 40 IU were used as threshold values for IHA and the ELISA, respectively, 4.8% of the specimens tested were positive in both tests. An additional 2% of these specimens were ELISA positive and IHA negative, whereas 2% were ELISA negative and IHA positive. The total number of positive specimens by either test (7%) closely agreed with previously reported results. In view of these data, the simplicity of the test, and relative economic considerations, the ELISA may be considered an alternative method for screening for anti-Toxoplasma antibodies.