Analysis of sensitization to carboxymethylcellulose: Identification of high risk group using ELISA and histamine release experiment

Objective and Design: Carboxymethylcellulose (CMC) has been considered to be inert and is commonly used as an additive in medicines, foods and cosmetics. However, we experienced a patient who developed an anaphylactic reaction to CMC after an upper gastrointestinal examination using a barium meal containing CMC. Therefore, we examined the incidence of sensitization by CMC in healthy subjects, and categorized the high risk group prone to developing anaphylactic response to CMC.Methods: An ELISA for detecting CMC-specific IgE antibody was developed using serum from the patient as a positive control. In the ten subjects exhibiting high anti-CMC IgE among 387 normal populations, histamine release from isolated leukocytes was performed.Results: Five of ten subjects with a high IgE titer showed a significant CMC-induced histamine release from leukocyte preparations in vitro as observed in the patient, and were classified as high risk group. There was a correlation between sensitization by CMC and that by Japanese cedar pollen. The incidence of sensitization in females was 2.4 fold higher than that in males.Conclusions: The combination of ELISA and histamine release experiment made it possible to identify the high risk group for developing anaphylactic response. The administration of high dose CMC as a suspending agent in barium sulfate or injectable corticosteroids to this group should be avoided to prevent anaphylactic reactions in the clinic.Received 18 August 2003; returned for revision 29 September 2003; accepted by M. J. Parnham 10 December 2003     

Detection by ELISA of IgA and IgM antibodies in secretion and IgM antibodies in serum in primary lower respiratory syncytial virus infection

Twenty-six infants and children with primary lower RS virus infection, diagnosed by the detection of RS virus in nasopharyngeal secretion (NPS) by use of immunofluorescent antibody (FA) technique, were studied with respect to the presence of IgA and IgM antibodies. Samples of NPS and serum obtained during the first 3-4 months following the beginning of illness, were investigated. Employing a reverse ELISA technique, we found IgM antibodies in the acute, but not during the convalescent, phase of illness in NPS from 20 of the patients and in serum from 21 of the patients. The majority of the IgM antibody conversions observed occurred in NPS as well as in serum on days 5-8 following the illness. RS virus IgA antibodies, also detected by a reverse ELISA technique, were demonstrated in NPS in 22 of the patients, with antibody conversions being found in 19 of the patients on days 5-8 following the beginning of the illness. Two patients still had IgA antibodies in NPS approximately 3 months FSOI. By comparison, RS virus was detected in acute-phase NPS by double-antibody sandwich ELISA in 25 of the 26 patients investigated.     

Humoral Immunity to Dietary Antigens in Atopic Dermatitis

Enzyme-linked immunosorbent assays (ELISA) employing a biotin-avidin amplification step are described for the quantification of human serum IgG antibodies to the dietary antigens ovalbumin (OA) and beta-lactoglobulin (BLG). The analytical quality of these assays was acceptable. Antibodies were measured in 16 patients with mild or moderate atopic dermatitis (AD), in 31 patients with a history of AD, and in closely matched controls. Levels of serum anti-OA antibodies did not differ in patients and controls, whereas anti-BLG antibodies tended to be higher in patients with mild or moderate AD than in controls (P less than 0.05).     

Factors influencing the sensitivity of herpes simplex virus detection in clinical specimens in a simultaneous enzyme-linked immunosorbent assay using monoclonal antibodies

A rapid simultaneous enzyme-linked immunosorbent assay (ELISA) using monoclonal antibodies was investigated for herpes simplex virus (HSV) detection. All HSV isolated (n = 127) were detected, whereas no response was obtained with HSV negative preparations. Equivalent results were obtained from 275 of 277 clinical specimens in the monoclonal ELISA and in an ELISA using polyclonal antibodies, confirming that appropriately selected monoclonal antibodies may be as efficacious as polyclonal antibodies in antibody-based assays. In clinical specimens, the rate of HSV detection (sensitivity) relative to tissue culture isolation was low for both assays, and the major factor responsible for this was the low concentration of virus present in some specimens. The sensitivity of ELISA obtained in routine use varied with different panels of unselected specimens and was related to the speed of development of the cytopathic effect. These results emphasise the need for caution in assigning a definitive sensitivity level to ELISA tests evaluated on different panels of specimens.     

Enzyme-Linked Immunosorbent Assay for Determination of Major Excrement Allergens of House Dust Mite Species D. pteronyssinus, D. farinae and D. microceras

Species-specific enzyme-linked immunosorbent assays (ELISA) for major excrement allergens (Dp42, Df6 and Dm6) of D. pteronyssinus, D. farinae and D. microceras in house dust were established, using immunoabsorbed, monospecific rabbit antibodies, coupled to horse radish peroxidase. The limit of detection was 13, 4 and 38 ng/ml, respectively. The coefficient of variation for the entire procedure, including dust sieving (212 micron) and extraction was 5-16% for allergen levels above 1000 ng/g dust. Allergen concentration by ELISA correlated well with the number of mite bodies identified and counted by microscopy in 31 dusts (r = 0.88, 0.86 and 0.82 for combined Dermatophagoides sp., D. pteronyssinus and D. farinae group, resp.) Dermatophagoides allergen was recorded in 21/22 mattress dusts (median: 26,000 ng/g; maximum: 290,000 ng/g). D. pteronyssinus allergen occurred in largest amounts (median 7,500 ng/g) followed by D. microceras (median 650 ng/g) and D. farinae (median 240 ng/g).     

High frequency of Coxsackie-B-virus-specific IgM in children developing type I diabetes during a period of high diabetes morbidity

Twenty-four consecutive children with newly diagnosed insulin-dependent (type I) diabetes mellitus (IDDM) were investigated for a history of infectious disease. Thirteen of the 24 (54%) patients reported symptoms of acute infection within two months before diabetes was diagnosed. The mean age was 8.5 years and 15 (63%) of the patients were girls. No clear seasonal variation in onset was seen. Coxsackie B (CB)-virus-specific IgM responses were detected by reverse radioimmunoassay (RIA) in 16 of the 24 (67%) patients on the day of diagnosis of IDDM. The highest titre was usually recorded at that time, but with some the highest titre was found with a second serum obtained three to seven weeks after diagnosis. Thereafter the titres declined, and after six months IgM was detected only in a few patients. Thirteen patients displayed monotypic IgM responses, whereas three patients showed ditypic responses. Among the former, IgM was recorded against Coxsackie B4 (CB4) in four, B5 (CB5) in three, B1 (CB1) in two, B2 (CB2) in two, and B3 (CB3) in two patients. The ditypic responses were against CB2 and CB3, CB3 and CB4, and CB5. No CB-virus-specific IgM was detected in sera, found during the same period, from age-matched nondiabetic children without evidence of infection. In neutralisation (NT) tests, antibodies to the homotypic virus were found in 12 of the 16 diabetic patients showing CB-virus-specific at the time of diagnosis. A significant rise in NT titre was demonstrated in three of these patients. No significant clinical difference was noted between IgM positive and IgM negative patients.(ABSTRACT TRUNCATED AT 250 WORDS)     

Production and characterization of antibody against fusarochromanone

Antibody against fusarochromanone (TDP‐1) was obtained from rabbits after immunization with TDP‐1 conjugated to bovine serum albumin (BSA). An indirect enzyme‐linked immunosorbent assay (ELISA) using TDP‐1‐ovalbumin conjugate as the antigen coated on to the microtiter plate was used for monitoring the antibody liter. For toxin detection, a direct competitive ELISA in which TDP‐1 was conjugated to horseradish peroxidase (HRP) was used. Competitive direct ELISA revealed that the antibody had about 5.6 and 4.5 times greater binding efficiency for monoacetyl fusarochromanone (TDP‐2) and diacetylated TDP‐1 than TDP‐1. The concentration causing 50% inhibition of binding of TDP‐1‐HRP to the antibody by TDP‐1, TDP‐2 and diacetyl‐TDP‐1 were 2.8, 0.5 and 0.62 ng/ml, respectively. For the analysis of fusarochromanones in wheat and barley, the toxins were first extracted from the commodities with 100% methanol. A small aliquot of the extract was dried, acetylated, diluted in buffer and then analyzed directly by ELISA. The overall recovery for fusarochromanone in the wheat and barley samples spiked with TDP‐1 in the concentration range of 20 to 500 ppb were found to be 97% and 103.4% with cv of 15% and 11.2% for barley and wheat, respectively.     

Reactivity of a monoclonal antibody produced to the histidine-rich protein of Plasmodium lophurae with Plasmodium falciparum

Monoclonal antibodies were produced against the histidine-rich protein of Plasmodium lophurae and tested for reactivity with Plasmodium falciparum antigens. One anti-histidine-rich protein monoclonal antibody showed immunological cross-reactivity with polypeptides of P. falciparum synthesized in vivo and in vitro.     

Antibodies to HTLV-I in populations of the southwestern Pacific

Sera collected from 1,102 individuals in 14 populations of the southwestern Pacific between 1956 and 1979 were tested by ELISA for antibodies to human T-cell leukemia virus type I (HTLV-I). Selected sera were also tested by particle agglutination and immunoblotting. Six of the populations had prevalences of antibodies greater than 4%, two populations had prevalences greater than 15%. Six populations had antibody prevalences of 2% or less. Three populations from the coast and northern islands of New Guinea had high prevalences of antibodies, while three New Guinea highland groups had virtually none. One population from the Solomon Islands had a high prevalence, while two others had very low prevalences. Two populations from small remote islands in Vanuatu both had high prevalences. Pacific sera did not neutralize a standard strain of virus readily neutralized by Japanese, European, and American sera. We conclude that infections with HTLV-I, some acquired more than 20 years ago, are widespread throughout the southwestern Pacific, even in several very isolated populations, although others have been spared. Some strains of HTLV-I in populations of the Pacific may have substantially different envelope proteins from prototype strains of America, Europe, and Japan.     

Primary and recurrent cytomegalovirus infections have different effects on human herpesvirus-6 antibodies in immunosuppressed organ graft recipients: absence of virus cross-reactivity and evidence for virus interaction.

The relationship between serum antibodies to human herpesvirus-6 (HHV-6) and cytomegalovirus (CMV) infection was studied in immunosuppressed adult organ graft recipients all of whom had IgG to both HHV-6 and Epstein-Barr virus capsid antigen (EBVCA) before operation and who had received an organ or organs from HHV-6 seropositive donors. In primary CMV infection the titre of IgG to HHV-6 rose substantially (between 32- and 512-fold) in eight out of eight patients whereas IgG to EBVCA only rose 32-fold in two patients. Moreover, the HHV-6 responses coincided closely with the CMV seroconversion. Serum absorption studies gave no evidence for antibody cross-reaction between CMV and HHV-6 because the CMV antibody titre could be reduced specifically without affecting HHV-6 antibody titres and vice versa. In recurrent CMV infection, HHV-6 antibody levels rose (32-fold) in three out of eight patients but these changes did not coincide with the CMV antibody response. Similarly, in the complete absence of CMV infection, five out of eight patients showed antibody rises to HHV-6 (between four- and 16-fold). IgG titres to EBVCA were stable in both these groups of patients. It is concluded that there is serological evidence (rising titre greater than or equal to four-fold) for genuine HHV-6 reactivation or, alternatively, for reinfection in 16 out of the 24 patients. This phenomenon was most frequent in primary CMV infection where the largest HHV-6 antibody responses were seen probably because of an, as yet, undetermined interaction between the two viruses.