Gelatin Particle Agglutination Test Research Articles

Comparison of multiple detection methods of Mycoplasma pneumoniae antibody for the early diagnosis of pediatric mycoplasma pneumonia

Mycoplasma pneumoniae pneumonia (MPP) is a common disease in infants and young children. MPP can also be a potential cause of healthcare-associated pneumonia. However, it is challenging to make an accurate diagnosis in a timely fashion. Our goal is to determine the assay consistencies of acridinium ester chemiluminescence immunoassay (AECLIA) and gelatin particle agglutination (GPA) test for the detection of Mycoplasma pneumoniae (MP) antibody. In this study, a total of 1404 children with suspected MPP were enrolled. Among them, 130 were diagnosed as MPP positive by mycoplasma culture, and 186 were negative. MP antibodies were detected by AECLIA, enzyme-linked immunosorbent assay (ELISA) and GPA. Consistency rates, differences among assays, and diagnosis performance were compared for the three methods. The independent χ2 test results of AECLIA and ELISA for the detection of MP-IgG and MP-IgM antibodies were χ2 = 29.210, P < 0.001; χ2 = 9.081, P = 0.017, respectively, suggesting that the two detection methods are well correlated. Similar analyses were done for the comparison of AECLIA and GPA as well as the comparison of ELISA and GPA. The positive rates of these methods agree with epidemiology data and have good consistency. Thus, AECLIA with a much shorter assay time could be a better option for the screening of MPP.

Detection of Viruses and Mycoplasma pneumoniae in Hospitalized Patients with Severe Acute Respiratory Infection in Northern China, 2015-2016.

Severe acute respiratory infection (SARI) presents a huge disease and economic burden worldwide. The present study described the frequency and types of different infectious etiologies among hospitalized patients with SARI in Tianjin, China, during 2015 and 2016. Basic information, in addition to a throat or serum sample, was collected from SARI patients. Nine viruses were detected using reverse transcription polymerase chain reaction, and Mycoplasma pneumoniae was detected using the Serodia Myco II gelatin particle agglutination test. A total of 585 specimens from 2,290 SARI cases were collected. The most common infection (19.66%, 115/585) was M. pneumoniae, followed by influenza virus A/B (6.15%, 36/585), and respiratory syncytial virus (4.96%, 29/585). Identification of viral or M. pneumoniae infections was the highest in the pediatric medicine ward (74.84%, 119/159), followed by the intensive care unit (37.04%, 80/216) and respiratory medicine ward (34.29%, 72/210). M. pneumoniae was highest (38.71%, 24/62) in the 5-14-year age group. Influenza was the main infection in January 2015 and March 2016. The correlation coefficient for the proportion of hospitalized cases of SARI and the positive detection rate within the same week was 0.25. M. pneumoniae and influenza were the leading pathogens among hospitalized SARI patients. A continued surveillance of hospitalized cases of SARI can detect emerging diseases, such as avian influenza A (H7N9) virus and other respiratory disease outbreaks.

P265 Anti-NMDAR auto-immune encephalitis

for a week. MRI brain showed bilateral signal changes in thalami and in the brain stem suggestive of rhombencephalitis. Lumbar puncture (LP) showed 38 cells (98% lymphocytes). He received IV Cefatoxime and IV Acyclovir till CSF bacterial and viral cultures were negative. Blood Mp titre by gelatin particle agglutination test (GPAT) was 2560 (normal less than 40). He was treated with Clarithromycin and steroids for three weeks. Following extubation, he demonstrated mutism with extrapyramidal movements for 2 weeks which spontaneously resolved. At 3 months follow up, he is reported to be normal and back at school. Case 2: A sixteen-year old girl with retroviral infection on treatment presented with a five-day history of headache preceded by a cough of two weeks. She had bilateral papilloedema with no focal neurological signs. MRI brain was normal. LP revealed raised opening CSF pressure at 25 cm with normal cells, chemistry and cultures confirming the diagnosis of idiopathic intracranial hypertension. Blood Mp titre (GPAT) was 5120. She received a three week course of Clarithromycin. Follow up at 4 weeks showed normal fundoscopy with disappearance of headaches. Conclusion: Mp infection should be considered as a differential diagnosis in children with an unusual neurological presentation as the good outcome depends on early diagnosis and specific therapy.

An outbreak of mycoplasma pneumoniae pneumonia in a kindergarten

To study the epidemiological and clinical features of the mycoplasma pneumoniae pneumonia (MPP) that occurred in a single class of a kindergarten in Beijing in July 2006. The environment and the attendance record of the kindergarten from the beginning of August 2005 to the end of July 2006 were investigated, and the sick status of the children absent for illness were interviewed by face to face or telephone through their parents. The disease data of the in-patient children with MPP were collected through questionnaires and analyzed. Serological screening for MP was performed with the Serodia Myco II gelatin particle agglutination test (Fujirebio, Japan). In mid-July 2006, in a day-care kindergarten with 3 grade classes, 3 out of 25 six-year-old children in the top class were hospitalized within 4 days and diagnosed as MPP. A total of 8 children had the symptoms of fever and cough during late May and mid-July in 2006, 5 children conduct chest radiographs and all had pneumonia, all these five children showed antibody positive for MP, 3 of them showed a more than 4-fold increase in antibody titer to MP in serum. There were no pneumoniae cases in the other two classes during the same period, and no pneumoniae cases happened among the teachers in the top class and the parents of the 5 pneumoniae children. All the children were moved to this classroom temporarily with limited ventilation and sunshine in March 2006. After improvement of the ventilation in the classroom, no additional pneumoniae cases occurred in the top class till the early September 2006. The 5 MPP children showed neither sneeze and nasal obstruction, nor skin rash, earache and any other extrapulmonary complication, and their peripheral white blood cell count was in the normal range (3.9 - 7.7) x 10(9). The MPP outbreak in a kindergarten was caused by poor ventilation of the temporary classroom. MP infection in children is liable to cause pneumonia.

HLA profile in patients with AIDS and tuberculosis

Studies carried out in various populations have reported an association between some HLA specificities and susceptibility to tuberculosis. We investigated the class I and class II HLA profile in Brazilian patients of various ethnic backgrounds who had AIDS and tuberculosis. Twenty-two adult patients with AIDS and tuberculosis (Group I), 103 patients with AIDS without tuberculosis (Group II) and 423 healthy individuals not infected with HIV (Group III) were evaluated. Diagnosis of HIV infection was made by ELISA, confirmed by a gelatin particle agglutination test. Diagnosis of tuberculosis was made based on clinical/radiological presentation and direct bacilloscopy or clinical specimen cultures. Class I antigens were typed by microlymphotoxicity. Class II alleles were characterized by the polymerase chain reaction (PCR). Differences in frequency of HLA specificities between groups were found in the following antigens/alleles: Group I x Group II: HLA-A31 - p=0.026; HLA-B41 - p= 0.037; HLA-DRB1*10 - p=0.037; HLA-DQB1*5 - p=0.009. Group I x Group III (control): HLA-A31 - p = 0.000008; odds ratio (OR)=31.75; HLA-B41 - p=0.003; HLA-DQB1*5 - p=0.02. HLA-A31 and HLA-B41 antigens and the HLA-DRB1*10 and HLA-DQB1*05 alleles were over-represented in patients with AIDS and tuberculosis (Group I), suggesting that these HLA molecules are associated with susceptibility to tuberculosis in Brazilian patients with AIDS.

Comparative Assessment of the Gelatin Particle Agglutination Test and an Enzyme-Linked Immunosorbent Assay for Diagnosis of Strongyloidiasis

The performances of the gelatin particle agglutination test (GPAT) and enzyme-linked immunosorbent assay (ELISA) for the diagnosis of strongyloidiasis with reference to the results of the agar plate culture technique (APCT) were evaluated with samples from 459 individuals from communities in northeast Thailand where strongyloidiasis is endemic. The prevalence of strongyloidiasis in five sample groups determined by GPAT varied between 29.3 and 61.5% (mean, 38.8%). ELISA and APCT, employed concurrently, gave lower prevalence rates of 27.5% (range, 21.6 to 42.1%) and 22.7% (range, 12.7 to 53.8%), respectively. By using APCT as the standard method, the sensitivity of GPAT was generally higher than that of ELISA (81 versus 73%). The specificity of GPAT was slightly lower than that of ELISA (74 versus 86%). The resulting GPAT titers exhibited positive linear relationships with the ELISA values (optical density at 490 nm) (P < 0.05), which suggests that the GPAT titer also reflects the levels of specific antibody comparable to those reflected by the ELISA values. Based on the relative ease and simplicity of use of the technique as well as the acceptable rates of sensitivity and specificity of the test, GPAT is more practical for screening for strongyloidiasis than the conventional ELISA.

Plasma hydroxy metronidazole/metronidazole ratio in anti-HCV carriers with and without apparent liver disease

Aims To evaluate plasma hydroxy-metronidazole/metronidazole ratio as a dynamic liver function test in HCV-infected individuals with/without liver disease, in the absence of liver cirrhosis. Methods Metronidazole was administered intravenously in healthy volunteers, asymptomatic anti-HCV-positive blood donors, and in chronic hepatitis C patients. Serology to HCV was determined by a second generation assay and confirmed by gelatin particle agglutination test using recombinant antigens C22-3 and C200. Plasma concentration of metronidazole and hydroxy-metronidazole was measured by high performance liquid chromatography in samples collected 5, 10, 20 and 30 min following the end of metronidazole infusion. Results Chronic hepatitis C patients had abnormal liver enzymes, while healthy volunteers and anti-HCV-positive blood donors had normal liver biochemistry tests. Plasma metronidazole concentration was similar in all groups studied. Plasma hydroxy-metronidazole/metronidazole ratio was significantly reduced in HCV-infected subjects, an effect observed 10 min after the end of drug infusion. Conclusions Metronidazole clearance is impaired in anti-HCV-positive blood donors and chronic hepatitis C patients, indicating that HCV is capable of affecting liver function at early stages of the disease. The metronidazole clearance test can detect impaired liver function in HCV-infected individuals even in the absence of liver cirrhosis.

Markers of human T lymphotropic virus type I in patients with cancer of uterine cervix in Amazon, Brazil

Human T lymphotropic virus type I(HTLV-I) has been implicated in various human diseases. Serum samples of 390 Brazilian Amazonians with cancer of various types were tested for HTLV-I antibodies by Gelatin particle agglutination test, Enzyme linked immunosorbent assay and Western blotting. Of 134 sera from patients with cancer of uterine cervix, 4 were positive by all the methods. Three of these were from non-transfused patients. DNA was extracted from 2 of 4 seropositive sera that gave strong reactions and were analyzed by PCR-SSCP for HTLV-I sequences. One was positive for all HTLV-I genes tested while the other one was positive for LTR and tax and negative for gag. In view of a possible pathway of the virus by sexual contact, the involvement of HTLV-I in cervical cancer warrants further studies.

More reliable diagnosis of infection with human immunodeficiency virus type 1 (HIV-1) by detection of antibody IgGs topol andgag proteins of HIV-1 and p24 antigen of HIV-1 in urine, saliva, and/or serum with highly sensitive and specific enzyme immunoassay (immune complex transfer enzyme immunoassay): A review

Ultrasensitive enzyme immunoassays (immune complex transfer enzyme immunoassays) were developed for antibody IgGs to HIV-1 using recombinant reverse transcriptase (rRT), p17 (rp17), and p24 (rp24) as antigens. Antibody IgGs were reacted with 2,4-dinitrophenyl-recombinant antigens and recombinant antigen-beta-D-galactosidase conjugates, and the immune complexes formed, comprising the three components, were trapped onto polystyrene beads coated with (anti-2,4-dinitrophenyl group) IgG. After washing, the immune complexes were eluted from the polystyrene beads with excess of epsilon N-2,4-dinitrophenyl-L-lysine and were transferred to clean polystyrene beads coated with (antihuman IgG gamma-chain) IgG. beta-D-Galactosidase activity bound to the last polystyrene beads was assayed by fluorometry. By transfer of the immune complexes from one solid phase to another, the nonspecific binding of the beta-D-galactosidase conjugates was minimized and the sensitivity was markedly improved. The immune complex transfer enzyme immunoassays using rRT, rp17, and rp24 as antigens were 300-1,000-fold, 1,000-3,000-fold, and 30-100-fold, respectively, more sensitive than Western blotting for the corresponding antigens and 10-300-fold more sensitive than a conventional ELISA and a gelatin particle agglutination test. For urine (100 microliters), whole saliva (1 microliter), and serum (1 microliter) samples, the sensitivity and specificity of the immune complex transfer enzyme immunoassay using rRT as antigen were both 100%. However, for urine samples in which the specific activities of antibody IgG to RT, p17, and p24 were much lower than those in serum samples probably due to degradation by the kidney, a longer assay of bound beta-D-galactosidase activity or/and a concentration process for urine was required. The use of more than 1 microliter of whole saliva was recommended for reliable diagnosis of the infections, whereas 1 microliter of serum was sufficient for the purpose. The positivity with rRT as antigen could be confirmed by demonstration of antibody IgGs to p17 and p24 in most of the urine, whole saliva, and serum samples. In HIV-1 seroconversion serum panels, antibody IgG to p17 was detected as early as or even earlier than antibodies to HIV-1 by a conventional ELISA or/and a gelation particle agglutination test, whereas antibody IgGs to RT and p24 were detected as early as or later than antibody IgG to p17. Thus the uses of rRT and rp17 as antigens were advantageous over that of the other antigens for randomly collected serum samples probably long after the infection and serum samples at early stages of the infection, respectively. On the basis of these results and other reports, the immune complex transfer enzyme immunoassay was developed for simultaneous detection of p24 antigen and antibody IgGs to RT and p17 in a single assay tube, and the window period (8 weeks, although widely variable), during which diagnosis of HIV-1 infection is not possible due to the absence of detectable antibodies to HIV-1, was shortened by 2 weeks. As a result, the simultaneous detection made possible not only as early diagnosis as that by detection of p24 antigen, but also as reliable diagnosis as that by detection of antibodies to HIV-1. Finally, the immune complex transfer enzyme immunoassay has been recently improved so as to be performed within shorter periods of time (2-3 hr) with higher sensitivity, and testing many samples has become easy.

Detection of anti-phenolic glycolipid I antibodies by ELISA and gelatin particle agglutination test in leprosy in São Paulo (SP), Brazil

A total of 214 serum samples obtained from leprosy patients (96 with the lepromatous form, 36 with the tuberculoid form, 33 with the indeterminate form, and 19 with the borderline form) before and during chemotherapy, from household contacts ( n = 18) and from controls ( n = 12) were submitted to the M. leprae particle agglutination test (MLPA) and to enzyme-linked immunosorbent assay (ELISA) for the detection of anti-phenolic glycolipid (PGL I) antibodies. ELISA was more sensitive for all clinical forms of leprosy, with greater seropositivity for the lepromatous form (54.64%). For 88 patients with the lepromatous form, we noted that the shorter the time of treatment (≤ 3 years), the higher the percentage of seropositive results ( P ≤ 0.01). The present results suggest that both tests could be used to monitor leprosy treatment and in epidemiologic surveys.

Age Distribution Among Patients at High Risk for Human T-Cell Lymphotrophic Virus Type I Infection

Human T-celllymphotropic virus type I (HTL V-I) is a human retrovirus associated with adult T cell leukemia and tropical spastic paraparesis. We recently reported a high incidence of HTLV-I infection in Israeli Jews originating from the city of Mashhad in Iran [1, 2]. There is a characteristic age-dependent increase in the seroprevalence of HTLV-I infection in areas of endemicity. This increase typically occurs in adulthood and is higher in females than in males throughout their lifetimes [3, 4]. It is not fully understood whether age, immunity, and the CNS are distinctive factors or cofactors associated with morbid­ ity in elderly populations. We designed the present study to investigate whether there is an age-dependent increase in the incidence of HTLV-I infection in a population at high risk; the results of our investigation were designed to be used as a model for CNS infectivity in the elderly. The Jews of Mashhad were persecuted because of their religious beliefs since 1747, but they secretly continued to practice the Jewish religion and to intermarry, resulting in families that are closely related genetically. We included 321 Iranian Jews of Mash­ hadi origin (either the subject or at least one of his or her parents was born in Mashhad) in our study. All subjects underwent physi­ cal and neurological examination, and samples of peripheral blood were obtained from each subject after informed consent was ob­ tained. Sera were tested for antibodies to HTLV-I with use of the gelatin-particle agglutination test, and genomic DNA was ampli­ fied as previously reported [2]. Statistical analysis was performed with analysis of variance (ANOVA). Of the 321 subjects included in the study, 203 were female and 118 were male (mean age z SD, 45.7::':: 22.3 years). Fifty­ eight (18%) of the 321 subjects (38 females and 20 males; mean age z SD, 50::':: 17.3 years) were positive for HTLV-I antibody by serological testing and by peR analysis. Of the 58 HTLV-I infected subjects, 36 were asymptomatic carriers, whereas 22 (14 females and 8 men; 38%) had signs of spastic paraparesis. None of the 263 subjects who were not infected had abnormali­ ties of the pyramidal tract compatible with spastic paraparesis.

Use of polymerase chain reaction and quantitative antibody tests in children born to human immunodeficiency virus‐1‐infected mothers

The diagnosis of human immunodeficiency virus (HIV) infection in children born to HIV-infected mothers is complicated by the presence of passively acquired maternal antibodies, and exclusion of infection in these infants remains problematic. The use of genome detection by polymerase chain reaction (PCR) amplification and the quantification of anti-HIV-1 antibodies were examined as methods for early diagnosis. Blood samples were taken from 84 non-breast-fed infants of HIV-infected mothers in five Italian and Spanish centres, a subgroup of children enrolled in the European Collaborative Study (ECS) for whom clinical and immunological information has been documented from birth. Whole blood was added to glycigel cryopreservative, stored, and tested in the United Kingdom by a nested PCR method. Antibody to HIV-1 was detected and quantified by titration using a gelatin particle agglutination test. PCR sensitivity and specificity were assessed. Twenty-one of the 84 children tested were infected. The estimated PCR sensitivity ranged from 0% (95% CI 0-26%) on day 1, 57% (19-85) on day 7, to 63% (33-92) on day 30. The negative predictive value of PCR ranged from 85% (83-88) on day 0 to 98% (94-100) at 3 months of age. On average, the level of maternal antibody halved every 33 days (31-36.5) in uninfected children. Between 6 and 9 months of age, increases in antibody titres in infected children were not more informative than absolute levels. These findings suggest that antibody measurement may supplement genomic diagnosis and that this collection method provides an alternative to the use of dried blood spots.

Use of dried blood spots for the detection and confirmation of HTLV-I specific antibodies for epidemiological purposes.

AIMS--To modify and evaluate a gelatin particle agglutination test that could provide a sensitive, specific and inexpensive method for the detection of HTLV-I antibody in dried blood spot samples (DBS) collected on filter paper. METHODS--A set of 26 reference samples confirmed as HTLV-I antibody positive were assembled from patients with tropical spastic paraparesis or adult T cell leukaemia and blood donors. Serum samples and simulated antibody positive dried blood spot eluates were tested using the Serodia assay together with two confirmatory tests: HTLV BLOT 2.3, a western blot, and Select-HTLV, an enzyme immunoassay (EIA). Both confirmatory tests use synthetic peptides to differentiate between antibodies to HTLV-I and -II. The modified Serodia assay was then used to test anonymously 10,135 DBS collected from neonates from London. Samples reactive in the modified Serodia test producing a positive result were titrated to an end point and confirmed as before. RESULTS--All 26 eluates made from simulated DBS derived from positive reference samples were identified as positive by the modified Serodia HTLV-I test and were confirmed as anti-HTLV-I positive by EIA. Two eluates derived from relatively low titre reference samples gave indeterminate results on western blotting. Screening of the 10,135 neonatal DBS resulted in six repeat reactives, five of which were confirmed. The remaining reactive sample gave an indeterminate result on western blotting and there was insufficient eluate for testing by EIA. The overall seroprevalence of HTLV-I in this population was 0.05% (five of 10,135). CONCLUSION--The modified Serodia HTLV-I assay provides a sensitive, specific and inexpensive (10 pence/test) method for screening large numbers of DBS. The format of the assay makes it ideally suited for simultaneous screening of antibodies to HIV-1, HIV-2 and HTLV-I using semi-automated equipment.

Anti‐HTLV‐I IgG in urine detected by sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using a synthetic peptide, Cys‐env gp46(188‐224), as antigen

Antibody IgG to human T-cell leukemia virus type I (HTLV-I) in urine was detected by a sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) using a synthetic peptide, Cys-env gp46(188-224), as antigen, the sensitivity and specificity of which were 100 and 98.5%, respectively, using serum samples. Anti-HTLV-I IgG in urine was reacted simultaneously with 2,4-dinitrophenyl-bovine serum albumin-Cys-env gp46(188-224) conjugate and Cys-env gp46(188-224)-beta-D-galactosidase (Escherichia coli) conjugate. The complex formed, consisting of the three components, was trapped onto polystyrene balls coated with affinity-purified (anti-2,4-dinitrophenyl group) IgG, eluted with epsilon N-2,4-dinitrophenyl-L-lysine and transferred to polystyrene balls coated with affinity-purified (anti-human IgG gamma-chain) IgG. Finally, bound beta-D-galactosidase activity was assayed by fluorometry. Thirty-one urine samples from seropositive subjects and 100 urine samples from seronegative subjects were tested. The sensitivity and specificity were 87 and 100%, respectively, with unconcentrated urine samples and 94 and 100%, respectively, with approximately 10-fold concentrated urine samples. These results were superior to those by the conventional ELISA and gelatin particle agglutination test.

Diagnosis of infection with human T-lymphotropic virus type II (HTLV-II) in Norwegian HIV-infected individuals

Sera from 298 HIV-infected individuals from Southern Norway were examined for antibodies against HTLV. 30 sera (10.1%) were HTLV-II positive and 1(0.3%) HTLV-I positive. 25 of the HTLV-II infected subjects were intravenous drug abusers (IVDAs), giving a prevalence of HTLV-II infection of 24.5% in this group. Examination of blood samples by polymerase chain reaction followed by restriction enzyme analysis or sequencing confirmed the serological diagnosis. To evaluate current screening and verification HTLV tests, 44 sera were examined using a gelatin particle agglutination test, 5 different enzyme-linked immunoassays (ELISA) and 4 Western blots (WB). While earlier ELISAs and WBs were inadequate, a recent ELISA and WB including recombinant envelope glycoproteins from both viruses permitted serological diagnosis and distinction between HTLV-I and HTLV-II. Thus, HTLV-II now spreads among IVDAs in a North-European country. Health authorities in other countries should estimate the magnitude of the problem to decide upon measures to avoid transmission through blood transfusion.

USE OF CYTOMEGALOVIRUS-SERONEGATIVE BLOOD COMPONENTS FOR PREVENTION OF CYTOMEGALOVIRUS INFECTION AFTER ALLOGENEIC BONE MARROW TRANSPLANTATION

Cytomegalovirus (CMV) infection, especially CMV pneumonia, is a serious event in immunocompromised individuals such as recipients of bone marrow transplantation. Since there is no effective treatment, prevention is the most effective way to control CMV infection. Blood transfusion is one potentially dangerous source of the virus. Screening for CMV negative donors is mandatory in order to provide blood components for these patients. We have screened 1, 827 donors for CMV antibody by using the gelatin particle agglutination test and ELISA. As it has been shown that the positivity rate is closely related to donor age, we tested mainly younger individuals. The results showed that the CMV antibody negative rate was 18.3%.

Is there a wastage of resources due to non-specificity of anti-HIV ELISAs?

A library of anti-HIV ELISA 'grey-area' and repeatably reactive samples sent for confirmatory testing, were retested using a second technique, the modified (Fujirebio Gelatin Particle) agglutination test (MAT). On testing 224 grey-area reactive samples, only four were found to be reactive with this second test. On retesting a further 259 ELISA repeatably reactive samples, of which only 33 were confirmed to be anti-HIV-1, only 45 were reactive by MAT; these included the 33 confirmed as positive samples, thereby reducing the false-positive from 226 to 12. The introduction of this second technique supported our decision to cease the referral of grey-area samples. It also demonstrated the high prevalence of non-specificity of repeatable reactivity associated with some of the most specific ELISA kits currently available.

Improvement of Gelatin Particle Agglutination Test for Detection of Anti‐HTLV‐I Antibody

Partial modifications of antigen components were made to improve the gelatin particle agglutination (PA) test for the detection of antibodies against human T cell leukemia virus type‐I. Envelope glycoproteins prepared by lentil lectin affinity chromatography were further added to the purified viral antigens to be coated on the gelatin particles. Comparative studies with a conventional PA test kit (Serodia ATLA) and indirect immunofluorescence assay showed that the specificity and sensitivity of the new PA test were increased and that abnormal agglutination such as the prozone phenomenon was abolished by this improvement.

Simple and inexpensive detection and confirmation of anti-HIV-1 in sera from Africa

250 sera from Ndola Zambia collected from consecutive patients with clinical evidence of HIV infection were screened by an algorithm utilizing the Serodia-HIV gelatin particle agglutination test (Fujirebio Inc Tokyo) and if positive with the HIVCHEK (DuPont de Nemours Deutschland Bad Homburg Germany). As a validation of the algorithm the sera were also divided in 2 and retested both locally and after being carried frozen to Gottingen Germany with 5 HIV tests as follows: 1) ENV-80 ELISA for a biosynthetic polypeptide from gp41; 2) DPZ-ELISA for a viral extract from Jurkat/HTLV-IIIB cultures; 3) Serodia-HIV; 4) HIVCHEK 1+2; and 5) radio-immuno-precipitation assay (RIPA) for confirmation. Sera were classified as negative if they failed to react with all 4 screening tests; positive if they reacted with 1 virus core protein and 1 virus envelope glycoprotein screening test; and indeterminant if they reacted to only 1 core or 1 envelope screening test. 53 of the 250 sera were unreactive to all 4 screening tests. There were 151 sera positive on RIPA 4 with weak bands and 42 unreactive on RIPA. The Serodia-HIV was highly sensitive detecting all positives but 9 false positives. The HIVCHEK was more specific with only 3 false positives but 15 false negatives. By following the algorithm 151 positive sera and 94 negative sera were classified correctly. 4 sera appearing indeterminant on RIPA had weak reactions on Serodia-HIV and negative findings on HIVCHEK: thus the algorithm identified the indeterminant sera correctly. One serum was clearly positive on Serodia-HIV and negative on RIPA then on retesting showed a weak band for p24 on RIPA indicating a patient in early stages of seroconversion. Using the algorithm with RIPA done at a tertiary center instead of Western Blot would lower the cost of screening with confirmation of indeterminant sera from US$25 to $7.80 per average result. Neither of the 2 screening tests requires laboratory apparatus and could be done in primary and secondary health centers in a rural African setting. Furthermore HIVCHEK 1+2 can be used where HIV-2 as well as HIV-1 viruses are epidemic.

Evaluation of gelatin particle agglutination assay for the detection of anti-PGLI antibodies. Comparison with ELISA method and applicability on a large scale study using blood collected on filter paper

Given the technical difficulties of the ELISA method, a gelatin particle agglutination test (MLPA) has been developed recently for the detection of anti-PGLI antibodies. The purpose of this study was to compare these 2 tests. MLPA was found to be less specific than ELISA (91% versus 98%, chi 2 = 66.8, p less than 0.001). The sensitivity of both tests was of 95% for the diagnosis of multibacillary patients. In the case of paucibacillary patients. MLPA was found to be less sensitive than ELISA (21% versus 35%, chi 2 = 6.98, p greater than 0.01). The agreement between the 2 tests for a positive or a negative result was satisfying (85% to 100%), except for the weakly seropositive individuals (71%). The correlation between OD obtained with ELISA and antibody titre obtained with MLPA was statistically significant (r = 0.70, p less than 0.001). Conversely to ELISA, MLPA was not applicable on blood samples absorbed on filter paper without a serious loss of sensitivity. In conclusion, this study demonstrated that the MLPA test can only reliably detect anti-PGLI antibodies in multibacillary cases.