Evaluation of a malaria antibody ELISA and its value in reducing potential wastage of red cell donations from blood donors exposed to malaria, with a note on a case of transfusion-transmitted malaria.

BACKGROUND AND OBJECTIVES: Blood donations are often wasted for lack of a satisfactory procedure to evaluate donors potentially exposed to malaria. MATERIALS AND METHODS: We evaluated a commercial ELISA for the detection of antibodies to malaria and compared it with an immunofluorescent antibody test (IFAT). RESULTS: When 5,311 sera from routine non-exposed donors were tested, 24 (0.45%) were positive by the ELISA, using a Plasmodium falciparum antigen. Seventeen were subjected to confirmatory testing but none were positive by IFAT. Of 1,000 donors potentially exposed in endemic areas 15 (1.5%) were repeatably reactive by ELISA. 10 of these were tested by IFAT and 2 were positive. When 150 patients attending the Hospital for Tropical Diseases in London with acute malaria were tested, 73% of those infected with P. falciparum were repeatably reactive for malarial antibodies by ELISA and 56% with Plasmodium vivax. Of 88 stored clinical sera tested by both IFAT and ELISA 56 were positive by IFAT and of these 52 (93 degrees/0) were positive by ELISA. CONCLUSION: The ELISA is sufficiently sensitive and specific to screen at-risk donors. Its use could safely retrieve 40,000 red cell units currently discarded each year in Great Britain.

Diagnosis of human toxocariasis by antigen capture enzyme linked immunosorbent assay.

AIMS: To evaluate an antigen capture enzyme linked immunosorbent assay (ELISA) which detects a carbohydrate epitope on the excretory-secretory (ES) antigens of Toxocara canis in clinical practice. METHODS: Serum specimens from healthy adults, patients with acute visceral larva migrans, ocular and inactive toxocariasis, and with other helminth infections were examined by two site antigen capture ELISA. RESULTS: Over half of the patients (19/28) with acute toxocariasis had a positive result in contrast to a small proportion of those with inactive disease (1/10) or ocular infection (2/7). False positive reactions, however, were found in 25% of the patients with serologically confirmed schistosomiasis and filariasis. CONCLUSIONS: This assay is useful in confirming the diagnosis of acute visceral larva migrans but could not be used alone in diagnosis because of false positive reactions in patients with other helminth infections.

Progress toward a simplified polymerase chain reaction and its application to diagnosis of tuberculosis.

The complexity, expense, and susceptibility to contamination of the polymerase chain reaction (PCR) are all issues which need to be overcome if PCR is to be used outside of research laboratories. We addressed these problems with respect to the diagnosis of tuberculosis. First, we simplified the procedure for extracting Mycobacterium tuberculosis DNA from sputum samples. Two methods of sample preparation were compared: the chaotrope-silica method and a novel, more simple chloroform method. Second, we developed a colorimetric method for product detection. This method was as sensitive and specific as agarose gel electrophoresis for detection of PCR product. By using a one-tube nested protocol, 5 to 50 genome equivalents of M. tuberculosis DNA were detected. The simplified colorimetric PCR was compared with microscopy and culture for detection of M. tuberculosis in clinical specimens of sputum. A total of 171 sputum samples were investigated from 108 patients, 12 of whom were subsequently found to have tuberculosis by culture and/or microscopy. PCR of samples prepared by the chaotrope-silica method had a sensitivity of 75% and a specificity of 100% whereas PCR of samples prepared by the chloroform method had a sensitivity of 92% and a specificity of 99% when compared with the sensitivities and specificities of the combined classical microbiological methods for the diagnosis of tuberculosis. The simplified colorimetric PCR in combination with the chloroform sample preparation method was at least as sensitive as microscopy but had a greater specificity because samples with atypical mycobacteria were not detected by PCR. The sensitivity of the method for detection of smear-negative and extrapulmonary tuberculosis remains to be investigated.

The development and validation of a simple antigen detection ELISA for Plasmodium falciparum malaria.

A double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) is described for the detection of Plasmodium falciparum antigen. The test is based on an immunoglobulin (Ig) M capture monoclonal antibody on the solid phase and an IgG monoclonal antibody conjugated to peroxidase. The simple test takes about 2.5 h to complete and, because it uses whole blood with no prior treatment, it is possible to process batches of 50-100 samples simultaneously. The test is specific to P.falciparum and has a sensitivity close to that usually achieved with Giemsa-stained blood films. The reagents employed are stable at refrigerator temperatures for over 6 months, and as the test is compatible with human immunodeficiency virus and hepatitis B surface antigen ELISAs it could be suitable for blood transfusion screening.

The spectrum of ocular toxocariasis.

Ocular toxocariasis is rare and therefore the spectrum of clinical disease is difficult to establish. We present a review of the clinical features and laboratory findings in a group of patients with positive Toxocara serology and ocular toxocariasis. The clinical spectrum was diverse and milder disease was commoner than might be supposed from reviews of the literature. Eosinophilia was unusual, but featured in two cases of unilateral pars planitis.

Comparison of an EBV transformed cell line and an EBV hybridoma cell line producing the same human anti-HBs monoclonal antibody.

A stable human-mouse heterohybridoma secreting human anti-HBs monoclonal antibody in continuous culture for 12 months was generated. It grew faster than the parent EBV transformed lymphoblastoid cell line (LCL) but produced the same level of specific antibody. The LCL was positive for the Epstein-Barr Virus Nuclear Antigen (EBNA), human CD 23 and contained a diploid number of human chromosomes. The heterohybridoma was negative for EBNA, CD 23 and mouse Ly-1 mouse, despite retaining a full complement of diploid mouse chromosomes and a limited number of human chromosomes.

Development and characterization of human anti-HBs antibodies.

Peripheral blood mononuclear cells (PBMC) from 13 healthy hepatitis B vaccines were transformed with the Epstein-Barr virus (EBV) and lymphoblastoid cell lines (LCL) producing antibodies to hepatitis B surface antigen (anti-HBs antibodies). Seven LCL and two clones secreting human anti-HBs monoclonal antibody were generated and their antibodies purified. One clone was fused with a mouse myeloma and the antibody from a cloned anti-HBs secreting heterohybridoma purified. One of the 10 purified human anti-HBs antibodies was characterized as IgG4, the remainder were IgG1. The antibodies had either kappa or lambda light chains. Five of the antibodies which were conjugated to horseradish peroxidase recognised the "a" group determinants.

Epitope mapping of HBsAg using a panel of human anti-HBs antibodies.

Mapping of B cell epitopes on HBsAg was performed using a panel of human anti-HBs antibodies. Synthetic peptides representing different regions of HBsAg failed to inhibit the binding of two antibodies which recognized non-conformational HBsAg determinants in dot-blot ELISA and HBsAg polypeptide bands in immunoblot analysis. Cross-inhibition studies using five of the antibodies conjugated to horseradish peroxidase suggested that at least three different epitopes are recognised by the panel of antibodies, two of which are within the 'a' group determinant.

Studies of a common idiotype PR4 in autoimmune rheumatic disease.

A new common idiotype, designated PR4, is described. This idiotype was originally identified on a human hybridoma-derived monoclonal antibody from a patient with leprosy, which binds the major Mycobacterium leprae-derived antigen, phenolic glycolipid-1, poly(ADP)-ribose, DNA, and poly(dT). The PR4 idiotype was found in patients with systemic lupus erythematosus (SLE) (70%), rheumatoid arthritis (40%), and Sjögren's syndrome (15%). It was not, however, found in the spouses of the SLE patients or (unlike other lupus idiotypes) in their healthy first-degree relatives. Although no correlation between PR4 idiotype levels and disease activity in SLE was found, a subset of rheumatoid arthritis patients with high levels of the idiotype was identified.

ELISA--a possible alternative to establish a therapeutic drug monitoring system in severe and complicated falciparum malaria.

A modification of an enzyme-linked immunosorbent assay (ELISA) for the quantitative determination of plasma quinine levels is described. The test is rapid, sensitive and reproducible.

An enzyme-linked immunosorbent assay for the detection of Entamoeba histolytica antigens in faecal material.

This paper describes a method for the detection of Entamoeba histolytica antigens in stool samples using a multi-layer ELISA. The method is sensitive and specific, showing no interference with other intestinal parasites, e.g. E. coli, E. hartmanni, Endolimax nana, Iodamoeba buetschlii, Hymenolepis nana, Giardia lamblia, Trichomonas and Ascaris. The method provides a rapid and simple screening assay for E. histolytica infections and should assist in diagnosis and epidemiological studies of the disease.

Comparison of an ELISA with a RIA method for serum alpha-fetoprotein determination in screening for fetal neural tube defects.

An enzyme-linked immunosorbent assay (ELISA) was evaluated for serum alpha-fetoprotein determination in the antenatal screening for fetal open neural tube defects. The ELISA was used concurrently with an existing radioimmunoassay (RIA) method until serum specimens from 5000 pregnant women, between 15 and 20 weeks gestation, had been tested. The accuracy of the ELISA was similar to that of the RIA; the median AFP values by gestational week obtained with the ELISA were, on average, 2 KIU/L higher than the corresponding RIA values; the 10th and 90th percentiles, in terms of multiples of the median (MoM), were very similar. The precision of the two methods was also similar. The ELISA method yielded 1.8% results from unaffected pregnancies above 2.5 MoM compared with 1.4% by RIA, a small but statistically significant difference (P = 0.03). Both methods detected the same affected pregnancies identified during this period; five open neural tube defects, three with exomphalos and three intra-uterine deaths. The ELISA method was simple, required about one quarter less operator time than the RIA and enabled results to be generated in one day rather than the two days required by RIA. The ELISA method is a suitable alternative to RIA for routine use in screening for fetal neural tube defects.

A critical reappraisal of the use of enzyme-linked immunosorbent assays in the study of snake bite.

The enzyme-linked immunosorbent assay (ELISA) has been the most widely used serological test in snake bite immunodiagnosis and epidemiology. The technique has been applied, however, without due consideration of the many factors which would affect an inherently sensitive test system, especially in tropical rural areas where large scale snake bite studies are usually carried out. This review discusses the effects of non-specific reactivity, cross reactivity and the quality of reagents on both the sensitivity and specificity of venom antigen and antibody detection assays. Simple laboratory modifications to optimize the assays are described. The importance of using the predictive value to assess the validity of applying the same test system in different circumstances is stressed. To fulfil its potential as the most versatile immunoassay technique in snake bite research, the test conditions of the ELISA will have to be much more stringently controlled in future.

Malaria chemoprophylaxis with chloroquine in young Nigerian children. I. Its effect on mortality, morbidity and the prevalence of malaria.

One hundred and ninety-eight Nigerian children who received weekly chemoprophylaxis with chloroquine from shortly after birth until the age of one year or two years and 185 age-matched controls were studied. Chemoprophylaxis with chloroquine was partially, but not completely, effective in controlling malaria. Clinical malaria was documented significantly less frequently in protected children than in control children, and only 9% of random blood films obtained from protected children were positive for Plasmodium falciparum while 41% of random blood films from control children were positive for this parasite. Mean malaria antibody levels were lower in protected than in control children; for ELISA and precipitin antibodies the difference between the two groups was less marked at two years than at one year. Mortality was similar among protected and among control children. No rebound mortality or morbidity was observed after chemoprophylaxis was stopped.

Malaria chemoprophylaxis with chloroquine in young Nigerian children. II. Effect on the immune response to vaccination.

The immune response of 198 young Nigerian children protected against malaria by chemoprophylaxis with chloroquine to immunization with triple, poliomyelitis, measles, typhoid, meningococcal and BCG vaccines was compared with the immune response to vaccination of 185 control children. Good responses to triple, measles and BCG vaccines were shown by children in both groups; poorer responses were obtained to poliomyelitis, typhoid and meningococcal vaccines. The response to immunization of protected children was similar to that observed among control children for all the vaccines tested except for meningococcal polysaccharide vaccine. Protected children showed a significantly greater antibody response to both group A and group C meningococcal polysaccharides than control children. This finding supports the results of previous studies which have shown that the immune response to meningococcal polysaccharide vaccines is adversely affected both by acute malaria and by asymptomatic malaria parasitaemia.

A comparison of chloroquine and pyrimethamine as malaria chemoprophylactics in young Nigerian children.

The efficacy of chloroquine and pyrimethamine as malaria chemoprophylactics was investigated in young Nigerian children. Chloroquine resistance had not been documented in the study area; pyrimethamine resistance was probably present but uncommon. Children who received weekly chemoprophylaxis with pyrimethamine had a lower prevalence of malaria parasitaemia and malaria antibodies than children who received weekly chemoprophylaxis with chloroquine. Pyrimethamine given monthly gave a comparable degree of protection to chloroquine given weekly. Chloroquine frequently induced vomiting in young children and this may have impaired its efficacy as a prophylactic. We conclude that, in an area where neither chloroquine nor pyrimethamine resistance is prevalent, pyrimethamine is a better chemoprophylactic for young children than chloroquine.