Cervix cancer is about twice as common in Asia as in the Western world and its incidence varies among different Asian ethnic groups. A study based in Singapore, the population of which comprises Chinese, Indians and Malaysians, offers the opportunity to evaluate whether the same risk factors are important in this part of the world as in the West. A total of 135 cases and an equal number of controls were interviewed and details concerning reproductive and sexual history, smoking, hygiene, socio-economic status and education were collected. Seventy-three cases had invasive cancer while 62 had micro-invasive disease or CIN III. The most important risk factors were parity and number of sexual partners. Smoking was rare in cases and controls and did not appear to be an important determinant of risk. Of the socio-economic factors, education appeared most predictive and lowered the risk. Age at first intercourse was strongly correlated with education (positively) and parity (negatively), but not with number of sexual partners. Biopsies were available for HPV DNA analysis in 38 cases and 37% were positive, mostly for HPV type 16. All these factors gave similar risks in invasive and preinvasive disease.
Normal human sera (NHS), heat-inactivated at 56 degrees C for 30 min, demonstrated positive ELISA reactions for anti-cardiolipin (aCL) antibodies. The heat-induced reactivity in ELISA was inhibitable by the cardiolipin antigen and was abolished by prior IgG depletion of the heated NHS with a protein A preparation. The heat-potentiated aCL also cross-reacted selectively with phosphatidic acid and phosphatidylserine, but not with phosphatidylcholine or phosphatidylethanolamine.
Non-reactive SLE sera in an ELISA for anticardiolipin antibody (aCL) retested positive in the immunoassay when the sera were first heat-inactivated at 56 degrees C for 30 minutes. This was not a false positive phenomenon since the positive ELISA reactivity of the heated SLE sera was markedly reduced by inhibition with the cardiolipin antigen. Furthermore, the heat-potentiated ELISA reaction was abolished by prior IgG depletion of the SLE sera with Protein A preparation. The unmasked aCL in the heat-treated SLE sera also exhibited selective binding in ELISA to other negatively-charged phospholipids, namely phosphatidylserine and phosphatidic acid but not against either phosphatidylcholine or phosphatidyl-ethanolamine. The data strongly indicate an interaction between antiphospholipid antibodies and heat-sensitive serum component(s), a reduction of the latter resulting in the ELISA detection of the autoantibody.
Heat treatment of sera at 56 degrees C for 30 min results in positive ELISA reactions for anti-cardiolipin antibody (aCL) in sera that had undetectable or low levels of aCL before heat inactivation. The positive, potentiated reactivity of the heated sera in the aCL ELISA could be inhibited with the cardiolipin antigen and was abolished by prior IgG depletion using staphylococcal protein A. The heat-potentiating effect of aCL binding in ELISA was evident in both normal human sera and clinical sera including sera from patients with systemic lupus erythematosus and syphilis.
Serum antibodies against Epstein-Barr virus (EBV)-determined antigens have traditionally been titrated by the indirect immunofluorescence (IIF) technique. The avidin-biotin complex (ABC) immunocytochemical technique was used to determine the serum levels of IgA against EBV viral capsid antigen (IgA/VCA) and IgA against EBV early antigen (IgA/EA) in sera of 106 nasopharyngeal carcinoma (NPC) patients prior to treatment and 100 normal individuals. The sensitivity of the ABC technique is enhanced by an amplification of the antigen-antibody reaction, which involves the binding of the enzyme-linked ABC to the second biotinylated antibody. There was a good correlation (r = 0.9988) between ABC and IIF-determined IgA/VCA-positive titres, with the ABC technique being more sensitive than IIF in the detection of IgA/VCA in NPC sera: 94% (99/106) and 76% (80/106), respectively. The frequency of IgA/EA reactivity in NPC sera was also markedly increased by immunodetection with the ABC technique as compared with IIF technique: 63% (69/106) and 28% (30/106) respectively. Both the immunocytochemical techniques were equally specific in discriminating between elevated serum titres of IgA/VCA and IgA/EA in NPC sera from normal human sera.
Heat-sensitive serum masking cofactor(s) of antiphospholipid antibody (aPL) in normal human sera (NHS) are specifically inactivated at 56 degrees C. The degree of binding in ELISA by unmasked aPL in NHS was equivalent to that in non-heated, aPL-reactive autoimmune SLE sera. Previously "negative" SLE sera also reacted equally strongly in the aPL ELISA when similarly heat-inactivated. Isotype studies by ELISA of the heat-potentiated aPL in 36 NHS revealed the presence of specific IgG (34/36), IgM (11/36) and IgA (24/36) aPL antibodies. 11/36 (31%) NHS had all three aPL isotypes while 13/36 (36%) had both IgG and IgA antibodies to phospholipid.
The Epstein-Barr virus nuclear antigen I (EBNA I) is the only latent EBV antigen consistently expressed in malignant tissues of the nasopharynx. A 20-amino-acid synthetic peptide, p107 contains a major epitope of EBNA I. We tested sera from 210 patients with nasopharyngeal carcinoma (NPC) and from 128 normal individuals (NHS) for IgA antibodies to p107 using an enzyme-linked immunosorbent assay (ELISA). Whereas 191/210 (91%) of NPC patients had IgA antibodies to p107, only 17/128 (13.3%) of NHS had such antibodies and only 6/57 (10.5%) of sera from patients with malignancies other than NPC had IgA-p107 reactivity. Thirty-nine salivary samples from 46 NPC patients (84.8%) also contained IgA-p107 antibodies whereas only 3/42 (7.1%) of normal saliva samples were IgA-p107 positive. The results suggest that IgA antibodies to EBNA I may become a useful, easily measurable, marker for NPC.
Margosa Oil is an extract of the seed of the Neem tree and is widely used as a traditional medicine by Indians in India, Sri Lanka, Burma, Thailand, Malaysia and Indonesia. Used mainly for external applications, it is often administered orally to neonates and infants regularly in small amounts. Margosa Oil causes toxic encephalopathy particularly in infants and young children. The usual features are vomiting, drowsiness, tachypnea and recurrent generalised seizures. Leucocytosis and metabolic acidosis are significant laboratory findings. Management is aimed primarily towards the control of convulsions although supportive management is equally important. Prognosis is usually good but fatalities and neurological deficits have been reported. We report here two infants with Margosa Oil poisoning presenting with encephalopathy.
The anti-phospholipid antibody (aPL) in 26 heat-inactivated normal human sera (NHS) was tested for IgG subclass in ELISA. The specific antibody in NHS included all four IgG antibody subclasses, as well as IgA. The incidence of IgG subclasses ranged from 50% (13/26) for IgG1 to 92% (24/26) for IgG2. Specific IgA anti-phospholipid antibody (aPL) was detected by ELISA in 38% (28/73) of normal human saliva. The salivary IgA aPL bound preferentially to anionic phospholipids including cardiolipin, phosphatidylserine and phosphatidic acid but not to phosphatidylcholine or sphingomyelin. Unlike aPL in normal human sera, aPL in saliva was predominantly not associated with the previously described heat-labile inhibitor of aPL. This may indicate a role of salivary IgA aPL in local immunity by binding to cross-reactive bacterial cell surface components including phospholipids.
An ELISA using the Epstein-Barr virus nuclear antigen 1 (EBNA 1) was found to detect selectively specific IgA in sera from patients with nasopharyngeal carcinoma (NPC). The antigen, p107, was a 20-amino acid synthetic peptide, representing a major epitope of EBNA 1.267/294 (90.8%) of NPC patients had IgA antibodies to p107 but in normal individuals, only 41/577 (7.1%) had IgA/p107. In sera from patients with other cancers, 11/77 (14.3%) had IgA/p107 reactivity. 124 IgA/VCA positive and 86 IgA/VCA negative NPC sera were also tested for IgA/p107 binding in ELISA. The majority of IgA/VCA positive sera (117) also contained IgA/p107 antibodies. Of interest was the detection of 74/86 IgA/p107 reactive sera in the IgA/VCA negative group. The results suggest that the IgA/p107 ELISA could become a useful, complementary screening assay to the IgA/VCA immunofluorescence test for detection of NPC.
Epstein-Barr virus (EBV) receptors (EBV/C3d receptors) were detected, using the monoclonal antibody HB5, on 23 ectocervical and 5 endocervical biopsies of the uterine cervix. Elevated IgA titers against the viral capsid antigen and early antigen of EBV were also found in the cervical secretions from cervical carcinoma patients (83%), compared with samples from patients with cervical intraepithelial neoplasia (75%), herpes simplex virus-infected patients (0%), and gynecologic patients with nonmalignant conditions (0%). EBV DNA was present in 63% of cervical carcinoma biopsies detected by in situ hybridization. These observations suggest a positive association between EBV and carcinoma of the cervix.
Cerebral infarction in the young is likely to be non-atheromatous. While in previous studies no cause has been found in 40% to 50% of patients, an increasing role for haemorheological factors is becoming apparent. Among these, an association between antiphospholipid antibodies (aPLs) and ischaemic cerebrovascular disease is now well-recognised. This entity has not been previously reported in Malaysian patients. In a study of 80 patients with stroke below the age of 50 years who were seen at the University Hospital, Kuala Lumpur, between January 1982 and May 1992, 3 patients with ischaemic cerebral infarction were found to have aPLs. aPLs was detected using ELISA method for anticardiolipin antibodies (aCLs), and presence of lupus anticoagulant (LA) was established by kaolin clotting time, thromboplastin inhibition test and platelet neutralisation procedure. Only 1 patient had active systemic lupus erythematous. Cerebrovascular events were recurrent in one of the 2 non-lupus patients. aPL-related stroke should be considered in young patients who have cerebral ischaemia occurring without obvious cause. More cases are likely to emerge in Malaysia with active screening.
The BamHI Z EBV replication activator (ZEBRA) protein is involved in the switch from latency to productive cycle of Epstein-Barr virus. A recombinant ZEBRA protein was synthesized and assessed in enzyme-linked immunosorbent assay (ELISA) for serum IgG response in nasopharyngeal carcinoma (NPC) patients. In 100 NPC serum samples that were positive for IgA to the EBV viral capsid antigen (VCA), 75% had IgG anti-ZEBRA antibodies. In contrast, only 3/83 (3.6%) serum samples from healthy donors and 2/50 (4%) from other cancers were positive for IgG to ZEBRA. Interestingly, in a selected group of 100 NPC sera negative for IgA to VCA, 25% contained IgG anti-ZEBRA antibodies. This suggests that the ELISA for IgG anti-ZEBRA may also identify earlier cases of NPC not detected by the conventional immunofluorescence test for IgA to VCA.
Inhibition studies were carried out to study possible cross-reactivity between a peptide fragment of the Epstein-Barr virus nuclear antigen, EBNA-1, and keratin/collagen. The 20-amino acid peptide (pAG), derived from a glycine-alanine repeat region of EBNA-1, uniquely makes up about one-third of the viral protein and is a dominant IgA antigenic epitope in patients with nasopharyngeal carcinoma (NPC). A small percentage of normal human sera (NHS) also binds pAG and this reactivity is examined in this study. Ten percent (2/20) and 13.4% (2/15) of IgA-pAG-positive NPC sera and NHS, respectively, were significantly inhibited by keratin in a competitive ELISA system. Conversely, 31.6% (6/19) and 30.8% (4/13) of IgA-keratin-positive NPC sera and NHS, respectively, were significantly inhibited by pAG. This indicated minimum cross-reactivity between IgA serum antibodies to EBNA-1 and keratin. Using collagen as inhibitor, none of 18 and only 2/13 IgA-pAG-positive NPC sera and NHS, respectively, were inhibited. In the collagen ELISA system, only 2/19 (10.5%) and 4/25 (16%) of IgA-collagen-positive NPC sera and NHS, respectively, were inhibited with pAG. Therefore, cross-reactivity with collagen was also low. IgA-pAG-positive NHS may therefore not be a false positive phenomenon, but whether it may represent an early serological profile related to NPC carcinogenesis remains to be determined.
The use of maternal age alone to identify pregnant mothers at risk of a fetus with Down's syndrome has recently been supplemented by maternal serum screening using biochemical markers such as alpha-protein, human chorionic gonadotrophin and oestriol. These tests have been reported to increase the sensitivity of antenatal detection of such fetuses from 35% to 67% with a false positive rate of 5%. However, these maternal serum markers may be affected by maternal weight, the smoking history of mothers and diabetes mellitus. Furthermore, such sensitivities are achieved only when gestational age is assessed accurately by ultrasound. Many further studies need to be carried out before the introduction of maternal serum screening into routine obstetric practice in Singapore. These include studies on the incidence of Down's syndrome in the local population, studies on the distribution of these serum markers in the second trimester of pregnancy, sensitivities and positive predictive values of such a test in the local population as well as the socio-economic implications of implementing such a screening test in the local obstetric population.
The linear antigenic epitopes of the Epstein-Barr virus replication activator protein (ZEBRA), recognised by specific serum IgG in nasopharyngeal carcinoma (NPC), were determined. This was achieved by synthesizing the entire amino acid sequence of ZEBRA as a set of 29, 22-residue peptides with an overlap of 14 amino acids. The ZEBRA peptides were tested in enzyme-linked immunosorbent assay (ELISA) for IgG binding in sera from 37 selected NPC patients who had IgG antibodies to the native ZEBRA protein. The most immunogenic epitope was peptide 1 at the amino-terminal end with 36 of the sera reactive against it. Further analysis of peptide 1, using the multipin peptide-scanning technique, defined a 10-amino-acid sequence FTPDPYQVPF, which was strongly bound by IgG. Two other regions of ZEBRA were also identified as immunodominant IgG epitopes, namely peptide 11 (amino acids 82-103) and peptide 19/20 (amino acids 146-175) with 8-13 of the NPC sera reactive against the peptides. The number of peptides reactive with individual NPC serum varies from 1 to 6 or more and there is some correlation between a greater number of peptide (at least 4) bound and a higher (at least 1:40) titre of serum IgA to viral capsid antigen. The immunodominant ZEBRA peptide 1 could be utilised in IgG ELISA for the detection of NPC.
We investigated the aeroallergens affecting 200 asthmatics from the University Hospital in Kuala Lumpur, Malaysia and found 164 (82%) patients with skin prick test (SPT) reactivity to one or more of a panel of 14 allergens, which included indoor and outdoor animal and plant aeroallergens. Reactivity was most frequent to the indoor airborne allergens, with 159 (79.5%) reacting to either or both house dust mite (Dermatophagoides) species and 87 (43.5%) to cockroach. The SPT reactivity to house dust mites corresponded with the finding that patients found house dust to be the main precipitant of asthmatic attacks.