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Control Sera (control + sera)
Selected AbstractsNovel ELISA systems for antibodies to desmoglein 1 and 3: correlation of disease activity with serum autoantibody levels in individual pemphigus patientsEXPERIMENTAL DERMATOLOGY, Issue 5 2010Enno Schmidt Please cite this paper as: Novel ELISA systems for antibodies to desmoglein 1 and 3: correlation of disease activity with serum autoantibody levels in individual pemphigus patients. Experimental Dermatology 2010; 19: 458,463. Abstract:, Pemphigus vulgaris (PV) and pemphigus foliaceus (PF) are intraepidermal blistering skin diseases. PV is characterised by autoantibodies directed against desmoglein (Dsg) 3 and in patients with the mucocutaneous variant also against Dsg 1, whereas in PF, only Dsg 1 is targeted. Here, ectodomains of Dsg 3 and Dsg 1 were recombinantly expressed in a human cell line (HEK293) and applied as authentic solid phases in ELISA test systems. Autoantibodies against Dsg 3 and/or Dsg 1 could be detected in 71 (100%) of 71 PV sera and against Dsg 1 in 48 (96%) of 50 PF sera. Control sera showed reactivity with Dsg 3 and Dsg 1 in 0.2% and 0.7%, respectively, of 401 healthy blood donors and in 2.1% of 48 randomly selected patients with bullous pemphigoid. No reactivity with Dsg 1 and 3 was detected in 21 patients with linear IgA disease. For both pemphigus variants, a statistically significant correlation between clinical severity and autoantibody levels was observed as demonstrated for 10 PV and 5 PF patients. In conclusion, the use of the ectodomains of Dsg 3 and 1 as target antigens expressed in a human cell line resulted in sensitive and specific ELISA systems for both diagnosis and monitoring of PV and PF. [source] Cytotoxic factor-autoantibodies: possible role in the pathogenesis of dengue haemorrhagic feverFEMS IMMUNOLOGY & MEDICAL MICROBIOLOGY, Issue 3 2001U.C. Chaturvedi Abstract During dengue virus infection a unique cytokine, cytotoxic factor (hCF), is produced that is pathogenesis-related and plays a key role in the development of dengue haemorrhagic fever (DHF). However, what regulates the adverse effects of hCF is not known. We have previously shown that anti-hCF antibodies raised in mice, neutralise the pathogenic effects of hCF. In this study we have investigated the presence and levels of hCF-autoantibodies in sera of patients with various severity of dengue illness (n=136) and normal healthy controls (n=50). The highest levels of hCF-autoantibodies (mean±S.D.=36±20 U ml,1) were seen in patients with mild illness, the dengue fever (DF), and 48 out of 50 (96%) of the sera were positive. On the other hand the hCF-autoantibody levels declined sharply with the development of DHF and the levels were lowest in patients with DHF grade IV (mean±S.D.=5±2 U ml,1; P=<0.001 as compared to DF). Only one of the 13 DHF grade IV patients had an antibody level above the ,cut-off' value (mean plus 3 S.D. of the control sera). The analysis of data with respect to different days of illness further showed that the highest levels of hCF-autoantibodies were present in DF patients at >9 days of illness. Moreover, the DF patients at all time points, i.e. 1,4, 5,8 and >9 days of illness had significantly higher levels of hCF-autoantibodies (P<0.001) than patients with DHF grade I, II, III and IV. In addition DHF grade I and grade II patients had significantly more positive specimens than DHF grade III and grade IV patients at all time points. These results suggest that elevated levels of hCF-autoantibodies protect the patients against the development of severe forms of DHF and, therefore, it may be useful as a prognostic indicator. [source] Improved diagnoses of autoimmune hepatitis using an anti-actin ELISAJOURNAL OF CLINICAL LABORATORY ANALYSIS, Issue 5 2008Vincent Aubert Abstract The presence of antismooth muscle antibodies is one of the diagnostic criteria of autoimmune hepatitis. We evaluated a new anti-F-actin ELISA test and compared it with indirect immunofluorescence assay (IIFA) for antismooth muscle antibodies (ASMA). Two hundred and nine serum samples (35 autoimmune hepatitis, 174 other hepatopathies and control sera) were tested by IIFA on mouse stomach kidney sections for ASMA and by the Quanta Lite Actin ELISA for anti-F-actin antibodies. ASMA were detected in 26 of 35 sera from autoimmune hepatitis (74%) as compared with 25 (71%) with anti-actin antibodies, as well as in 25 of 49 (51%) samples from viral hepatitis as compared with 7 (14%) with anti-actin antibodies. With regards to autoimmune hepatitis, though sensitivity (74.3 vs 71.4%) and negative predictive value (93.5 vs 93.9%) of ASMA and anti-actin ELISA were comparable, anti-actin ELISA was significantly better than ASMA IIFA in terms of specificity (89.7 vs 74.7%), and positive predictive value (58.1 vs 37.1%). Although frequently positive in HCV samples, a comparable sensitivity but better specificity makes the anti-actin ELISA a useful tool in combination with ASMA IIFA for the screening and diagnosis of autoimmune hepatitis. J. Clin. Lab. Anal. 22:340,345, 2008. © 2008 Wiley-Liss, Inc. [source] Development of recombinant OmpA and OmpB proteins as diagnostic antigens for rickettsial diseaseMICROBIOLOGY AND IMMUNOLOGY, Issue 7 2009Eun-Ju Do ABSTRACT In this study the diagnostic potential of Rickettsia conorii recombinant antigens was analyzed. For this, site-specific PCR primers were used to clone the OmpA and OmpB genes of R. conorii into pMAL-c2X plasmids. Six fragments of OmpA and four of OmpB were expressed as fusion proteins with maltose-binding protein in Escherichia coli. OmpA1350-1784, OmpB801-1269, and OmpB1227-1634 regions from truncated proteins were selected as diagnostic candidate antigens by ELISA using control sera. ELISA results of three antigens were compared to the results obtained by using a commercial ELISA kit which contained whole OmpA and OmpB antigens from R. conorii. For this analysis, 40 serum samples taken from febrile patients and uninfected controls were tested. Of the 20 R. conorii test results which were positive with the commercial kit, 18 were shown to be positive by ELISA using OmpA1350-1784 (a sensitivity of 90%). The specificity of the ELISA was 100%; all of the 20 samples shown to be negative using the commercial kit were also negative in our assay. The sensitivities of the ELISA using the OmpB801-1269 and OmpB1227-1634 were 90% and 95%, respectively. The specificities of the OmpB801-1269 and the OmpB1227-1634 were 100% and 95%, respectively. These results suggest that specific regions of OmpA and OmpB effectively detect antibodies against R. conorii, and the truncated recombinant antigens could be used for development of diagnostic tools for rickettsial disease. [source] Identification of immunogenic cell wall-associated proteins of Streptococcus suis serotype 2PROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 17 2008Anding Zhang Abstract Streptococcus suis serotype 2 (SS2) is a porcine and human pathogen with adhesive and invasive properties. The absence of suitable vaccine or virulent marker can be the bottleneck to control SS2 infection. An immunoproteome-based approach was developed to identify candidate antigens of SS2 for vaccine development. Hyperimmune sera, convalescent sera, and control sera were analyzed for reactivity by Western Blot against SS2 cell wall-associated proteins (WAPs) separated by 2-DE. A total of 34 proteins were identified by immunoproteomic analysis, of which 15 were recognized by both hyperimmune sera and convalescent sera, including most WAPs currently characterized as SS2 vaccine candidate antigens: muramidase-released protein (MRP), surface protein SP1 (Sao), and glyceraldehyde-3-phosphate dehydrogenase (GapdH). The novel immunogenic proteins may be developed as alternative antigens for further study of SS2 vaccine and diagnostics. [source] Mining biomarkers in human sera using proteomic toolsPROTEINS: STRUCTURE, FUNCTION AND BIOINFORMATICS, Issue 1 2004Rulin Zhang Abstract One of the major difficulties in mining low abundance biomarkers from serum or plasma is due to the fact that a small number of proteins such as albumin, ,2-macroglobulin, transferrin, and immunoglobulins, may represent as much as 80% of the total serum protein. The large quantity of these proteins makes it difficult to identify low abundance proteins in serum using traditional 2-dimensional electrophoresis. We recently used a combination of multidimensional liquid chromatography and gel electrophoresis coupled to matrix-assisted laser desorption/ionization-quadrupole-time of flight and Ion Trap liquid chromatography-tandem mass spectrometry to identify protein markers in sera of Alzheimer's disease (AD), insulin resistance/type-2 diabetes (IR/D2), and congestive heart failure (CHF) patients. We identified 8 proteins that exhibit higher levels in control sera and 36 proteins that exhibit higher levels in disease sera. For example, haptoglobin and hemoglobin are elevated in sera of AD, IR/D2, and CHF patients. The levels of several other proteins including fibrinogen and its fragments, alpha 2-macroglobulin, transthyretin, pro-platelet basic protein, protease inhibitors clade A and C, as well as proteins involved in the classical complement pathway such as complement C3, C4, and C1 inhibitor, were found to differ between IR/D2 and control sera. The sera levels of proteins, such as the 10 kDa subunit of vitronectin, alpha 1-acid glycoprotein, apolipoprotein B100, fragment of factor H, and histidine-rich glycoprotein were observed to be different between AD and controls. The differences observed in these biomarker candidates were confirmed by Western blot and the enzyme-linked immunosorbent assay. The biological meaning of the proteomic changes in the disease states and the potential use of these changes as diagnostic tools or for therapeutic intervention will be discussed. [source] Application of immunoproteomics to leptospirosis: towards clinical diagnostics and vaccine discoveryPROTEOMICS - CLINICAL APPLICATIONS, Issue 4 2007Uraiwan Kositanont Abstract Each of the currently available methods for serodiagnosis of leptospirosis, including the microscopic agglutination test (MAT), has its own drawback(s) when used in clinical practice. A new diagnostic test is therefore required for an earlier and more accurate diagnosis of leptospirosis. We applied immunoproteomics to define potential immunogens from five serovars of Leptospira reference strains. A leptospiral whole cell lysate from each serovar was used as the antigen to react with IgG and IgM in the sera from four patients with a positive MAT. Sera from four non-leptospirosis patients with a negative MAT were pooled and used as the negative control. 2-D Western blot analysis showed that the degree of immunoreactivity corresponded with the MAT titers. No immunoreactive spots were detected when the pooled control sera were used. A total of 24 protein spots immunoreacted with IgM and/or IgG from patients with leptospirosis. These immunoreactive proteins were identified by MALDI-TOF MS and were classified into five groups, including flagellar proteins, chaperones/heat shock proteins, transport proteins, metabolic enzymes, and hypothetical proteins. More immunoreactive spots were detected with anti-human IgG in the sera of all patients and with all the serovars of leptospires used. Some of the identified proteins immunoreacted only with IgG, whereas the others were detectable with both IgM and IgG. Among the immunoreactive proteins identified, FlaB proteins (flagellin and flagellar core protein) have been shown to have a potential role in clinical diagnostics and vaccine development. These data underscore the significant impact of immunoproteomics in clinical applications. [source] Chronic inflammatory demyelinating polyneuropathy sera inhibit axonal growth of mouse dorsal root ganglion neurons by activation of rho-kinase,ANNALS OF NEUROLOGY, Issue 5 2009Junko Taniguchi MS Clinical course and prognosis are variable among patients with chronic inflammatory demyelinating polyneuropathy (CIDP), whereas the extent of axonal degeneration is the major prognostic factor. We studied the effects of sera from CIDP patients on axonal growth in cultured mouse dorsal root ganglion neurons. Compared with control sera, CIDP sera prominently suppressed axonal outgrowth of dorsal root ganglion neurons and shortened axonal length. The inhibitory activity was abolished by adding Y27632, a Rho-kinase inhibitor. These findings suggest that CIDP sera inhibit axonal elongation by Rho-kinase activation, and some serum factors may be responsible for development of axonal degeneration in CIDP. Ann Neurol 2009;66:694,697 [source] Markers of oxidative and nitrosative stress in systemic lupus erythematosus: Correlation with disease activity,ARTHRITIS & RHEUMATISM, Issue 7 2010Gangduo Wang Objective Free radical,mediated reactions have been implicated as contributors in a number of autoimmune diseases, including systemic lupus erythematosus (SLE). However, the potential for oxidative/nitrosative stress to elicit an autoimmune response or to contribute to disease pathogenesis, and thus be useful when determining a prognosis, remains largely unexplored in humans. This study was undertaken to investigate the status and contribution of oxidative/nitrosative stress in patients with SLE. Methods Sera from 72 SLE patients with varying levels of disease activity according to the SLE Disease Activity Index (SLEDAI) and 36 age- and sex-matched healthy controls were evaluated for serum levels of oxidative/nitrosative stress markers, including antibodies to malondialdehyde (anti-MDA) protein adducts and to 4-hydroxynonenal (anti-HNE) protein adducts, MDA/HNE protein adducts, superoxide dismutase (SOD), nitrotyrosine (NT), and inducible nitric oxide synthase (iNOS). Results Serum analysis showed significantly higher levels of both anti,MDA/anti,HNE protein adduct antibodies and MDA/HNE protein adducts in SLE patients compared with healthy controls. Interestingly, not only was there an increased number of subjects positive for anti-MDA or anti-HNE antibodies, but also the levels of both of these antibodies were statistically significantly higher among SLE patients whose SLEDAI scores were ,6 as compared with SLE patients with lower SLEDAI scores (SLEDAI score <6). In addition, a significant correlation was observed between the levels of anti-MDA or anti-HNE antibodies and the SLEDAI score (r = 0.734 and r = 0.647, respectively), suggesting a possible causal relationship between these antibodies and SLE. Furthermore, sera from SLE patients had lower levels of SOD and higher levels of iNOS and NT compared with healthy control sera. Conclusion These findings support an association between oxidative/nitrosative stress and SLE. The stronger response observed in serum samples from patients with higher SLEDAI scores suggests that markers of oxidative/nitrosative stress may be useful in evaluating the progression of SLE and in elucidating the mechanisms of disease pathogenesis. [source] Deiminated Epstein-Barr virus nuclear antigen 1 is a target of anti,citrullinated protein antibodies in rheumatoid arthritisARTHRITIS & RHEUMATISM, Issue 3 2006Federico Pratesi Objective To test the hypothesis that deimination of viral sequences containing Arg,Gly repeats could generate epitopes recognized by anti,citrullinated protein antibodies (ACPAs) that are present in rheumatoid arthritis (RA) sera. Methods Multiple antigen peptides derived from Epstein-Barr virus (EBV),encoded Epstein-Barr nuclear antigen 1 (EBNA-1) were synthesized, substituting the arginines with citrulline, and were used to screen RA sera. Anti,cyclic citrullinated peptide antibodies were purified by affinity chromatography and tested on a panel of in vitro deiminated proteins. Their ability to bind in vivo deiminated proteins was evaluated by immunoprecipitation, using EBV-infected cell lines. Results Antibodies specific for a peptide corresponding to the EBNA-135,58 sequence containing citrulline in place of arginine (viral citrullinated peptide [VCP]) were detected in 50% of RA sera and in <5% of normal and disease control sera. In addition, affinity-purified anti-VCP antibodies from RA sera reacted with filaggrin-derived citrullinated peptides, with deiminated fibrinogen, and with deiminated recombinant EBNA-1. Moreover, anti-VCP antibodies immunoprecipitated, from the lysate of calcium ionophore,stimulated lymphoblastoid cell lines, an 80-kd band that was reactive with a monoclonal anti,EBNA-1 antibody and with anti,modified citrulline antibodies. Conclusion These data indicate that ACPAs react with a viral deiminated protein and suggest that EBV infection may play a role in the induction of these RA-specific antibodies. [source] Anti-La/SSB antibodies transported across the placenta bind apoptotic cells in fetal organs targeted in neonatal lupusARTHRITIS & RHEUMATISM, Issue 6 2002Hai B. Tran Objective To determine whether La and/or Ro epitopes on apoptotic cells in fetal organs that are targeted in neonatal lupus syndrome (NLS) are accessible for binding by autoantibodies in vivo, we traced the fate of transplacental autoantibodies in a murine passive transfer model. Methods Pregnant mice at day 15 of gestation (E15) were injected intraperitoneally with human anti-Ro/La,positive sera or control sera, and transplacental transfer of human autoantibodies was tested by enzyme-linked immunosorbent assay with recombinant antigens. Multiple cryostat sections at the level of the heart of E17 fetuses were visualized simultaneously for human IgG binding and apoptosis (TUNEL) under confocal microscopy. Serial paraffin sections of E17 and E19 fetuses were examined for histologic evidence of inflammation. Results Human IgG anti,52-kd Ro, anti,60-kd Ro, and anti-La autoantibodies were transported efficiently into the fetal circulation. Human IgG,apoptotic cell complexes were detected in the heart (atrial trabeculae and atrioventricular node), skin, liver, and newly forming bone of fetuses from mothers injected with anti-Ro/La sera but not control sera. The IgG binding was fetal-specific and organ-specific; transplacental autoantibodies did not bind to apoptotic cells in the fetal thymus, lung, brain, or gut. The complexes were not associated with an inflammatory reaction. Injection of mothers with affinity-purified anti-La autoantibodies (but not anti-Ro/La Ig depleted of anti-La) revealed an identical location of IgG binding to apoptotic cells in the fetuses. Conclusion This is the first study to demonstrate that transplacental anti-La autoantibodies bind specifically to apoptotic cells in selected fetal organs in vivo, similar to the organ involvement in NLS. We hypothesize that additional factors are required to promote proinflammatory clearance of IgG,apoptotic cell complexes and subsequent tissue damage. [source] Aberrant serum hyaluronan and hyaluronidase levels in sclerodermaBRITISH JOURNAL OF DERMATOLOGY, Issue 3 2004B.A. Neudecker Summary Background Scleroderma, or systemic sclerosis, is characterized by aberrations of extracellular matrix deposition. These changes parallel early stages of wound healing when increased deposition of hyaluronan (HA) and collagen occur. Both processes result ultimately in the formation of fibrotic scar tissue. Activities of HA synthase and hyaluronidase, the enzymes that synthesize and degrade HA, are critical in HA turnover. Both become elevated whenever increased matrix deposition occurs. HA deposition occurs early in wound healing, and increases are documented in the circulation of scleroderma patients. We postulated that elevated HA and hyaluronidase may both be indicators of early-stage disease in scleroderma, in parallel with early changes observed in wound healing. In an attempt to reduce HA accumulation and the associated fibrosis in scleroderma tissues, topical and intravenous hyaluronidase administrations have been used in the past as treatment modalities, with occasional success. This also suggested that hyaluronidase enzyme activity is involved in the disease process. It is now recognized that the hyaluronidases constitute an enzyme family. The somatic hyaluronidase Hyal-1 is the only activity present in human serum. Objectives To determine levels of HA and Hyal-1 in the sera of scleroderma patients at various stages of their disease. Methods Levels of HA and Hyal-1 activity were determined in 25 scleroderma patients. Subjects were separated into two groups, those in the early stage with duration of disease of 2 years or less, and late-stage patients with disease duration of more than 2 years. Results In early-stage scleroderma, levels of HA were elevated significantly, as predicted, in comparison with late-stage patients and controls. Late-stage levels of HA were comparable with those found in control sera. By contrast, levels of Hyal-1 activity were normal in early-stage patients, similar to those in controls, but were decreased in late-stage patients, falling even below those of controls. Conclusions We have confirmed that circulating levels of HA are elevated in scleroderma, but show for the first time that such elevations occur predominantly in early-stage disease. Patients with late-stage disease have decreased serum Hyal-1 activity, perhaps reflecting decreased levels of HA turnover. This study also represents the first time that hyaluronidase activity levels have been determined in scleroderma patients. [source] SEREX identification of new tumour-associated antigens in cutaneous T-cell lymphomaBRITISH JOURNAL OF DERMATOLOGY, Issue 2 2004T.B. Hartmann Summary Background Cutaneous T-cell lymphoma (CTCL) is a clonal lymphoproliferative disorder of mainly CD4+ T cells, with primary manifestation in the skin. Objectives To detect new CTCL-associated antigens for immunological therapies and to define their specificity in terms of RNA expression and seroreactivity. Methods A newly constructed CTCL cDNA phage library was screened and cross-reactivities against the detected clones were tested using 15 mycosis fungoides and six Sézary syndrome sera. The mRNA expression of the identified genes was analysed by reverse transcription,polymerase chain reaction (RT,PCR) using 22 tumour tissues, nine cell lines and up to 29 different types of normal tissue. Results We identified nine different tumour antigens (HD-CL-01 to HD-CL-09) of which seven clones had high homology to genes with known functions. Several of these genes had previously been associated with cancer, namely inositol 1,4,5-triphosphate 5-phosphatase, vimentin, aldose reductase and elongation factor-1,. Variations in the deduced protein sequences were observed in three cases, mostly due to variations in protein length. The individual clones were recognized by up to 56% of patients' sera, while control sera were negative except in one case. Using RT,PCR, we found a frequent expression of these new tumour antigens in tumour specimens (26,100%). In contrast to humoral specificity, specific mRNA was also detected in selected normal tissues (29,89%). Conclusions SEREX (serological identification of antigens by recombinant expression cloning) identified multiple tumour-associated antigens in CTCL. The serological specificity and the high percentage of reactive sera of CTCL patients against several clones suggest these genes as potential targets for diagnostic and prognostic purposes. [source] A luminescent bioassay for thyroid blocking antibodiesCLINICAL ENDOCRINOLOGY, Issue 3 2001N. J. Jordan Thyroid blocking antibodies (TBAb) have a role in the development of hypothyroidism and in the neonate are responsible for transient hypothyroidism. Specific measurement of TBAb requires a bioassay, but current methods are lengthy and cumbersome. We describe a rapid luciferase-based method for the detection of TBAb using the lulu* cell line which is suitable for the provision of a clinical service Chinese hamster ovary (CHO) cells were transfected with human TSH-R together with G418 resistance and a cAMP responsive luciferase construct. Stable pools of transfected cells were selected and clones identified by limiting dilution. Clone lulu* gave the best response to stimulation by TSH and was used to develop a bioassay for TBAb. The luminescent bioassay conditions have been optimized and validated using 12 serum samples from patients found to be TBAb positive in a bioassay using an established method quantifying cAMP by radioimmunoassay (RIA). The effect of thyroid stimulating antibodies (TSAb) on the calculation of Inhibition Index (InI) using two previously described formulae have been investigated and we have used serum containing both TSAb and TBAb to investigate detection of TBAb in samples containing more than one type of activity. Lulu* displays a dose dependent increase in luciferase expression in response to stimulation with bovine (b) TSH which is more effective in serum free medium than in salt free buffer. TSH stimulated luciferase expression can be inhibited by TBAb in either serum or an immunoglobulin preparation. Using optimized assay conditions, challenging 10% serum against 1 U/l bTSH in culture medium, we have tested 31 euthyroid sera to determine a reference range: InI values >23% were considered positive. Twelve samples previously shown to contain TBAb by an established method quantifying cAMP by RIA were positive by the luciferase-based assay. Of control sera, 20/20 systemic lupus erythematosus, 13/14 rheumatoid arthritis, 12/12 multinodular goitre were negative. We demonstrated that if more complex formulae are used to calculate InI, false positive TBAb results can be obtained in samples containing only TSAb. Finally, when sera contain both TSAb and TBAb, the net activity of stimulating and blocking antibodies is detected in the bioassay. Where TSAb are also present, analysis of serum may be required at several dilutions to detect TBAb. We describe the production of a new cell line, lulu*, and its use to develop a luminescent bioassay for TBAb suitable for clinical use. Comparing two established methods of calculating TBAb, we found that they do not give identical results. In light of this, the high prevalence reported for TBAb in some studies has to be considered with caution. [source] Basophil Activation Test and specific IgE measurements using a panel of recombinant natural rubber latex allergens to determine the latex allergen sensitization profile in childrenPEDIATRIC ALLERGY AND IMMUNOLOGY, Issue 2 2006María L. Sanz There are no documented studies that describe natural rubber latex (NRL) sensitization in children with a history of surgical intervention but without any congenital malformation (urogenital anomalies, spina bifida, etc.), although some authors have studied NRL allergy in children without a history of surgical intervention. The aim of this work was to evaluate the sensitization profile to single NRL allergens in children without spina bifida and without repeated surgical interventions, by using different recombinant and natural latex allergens in two analytical techniques: specific serum immunoglobulin E (IgE) quantification and flow cytometry determination of activated basophils expressing CD63, after stimulating cells from patients with NRL allergens. A total of 23 patients and 10 healthy children were selected. Conjunctival and in-use NRL provocation tests were carried out, as well as specific IgE determination in all patients' and controls' sera with the recombinant NRL allergens: rHev b 1, rHev b 2, rHev b 3, rHev b 5, rHev b 6.01, rHev b 6.02, rHev b 8, rHev b 9 and rHev b 11 and with NRL (k82) using appropriate ImmunoCAPs. The Basophil Activation Test (BAT) was performed with whole latex extract and with the recombinant allergens rHev b 5 and rHev b 6.01, as well as with the natural allergen Hev b 6.02. The sensitivity and the specificity of NRL-specific IgE (k82) were 100%. Positive IgE responses to rHev b 5 were found in sera of 10 children, to rHev b 6.01 in 16 and for rHev b 6.02 in 15 children's sera. Specific IgE to rHev b 8 was found in four sera of the children. We only found significant differences in sensitization to rHev b 5 in children with two or more surgical interventions compared with the non-intervened group or those with only one intervention. Specific IgE in sera of children with latex-fruit syndrome recognized rHev b 6.02, but not to rHev b 11. The patients sensitized to Hev b 8, Hev b 9 and/or Hev b 11 were atopic. The four patients presenting a positive response to the NRL profilin Hev b 8 were allergic to pollen. The BAT against whole NRL extract was positive in 22 of 23 children; against rHev b 5 in 14 of the patients studied; against rHev b 6.01 in seven cases and against nHev b 6.02 in 19 children. In all the control subjects, the results using this technique were negative. If combined rHev b 5, rHev b 6.01 and nHev b 6.02 together, BAT could detect 20 of the 23 children with latex allergy. The combined use of ImmunoCAP with all the recombinant NRL allergens and BAT with rHev b 5, rHev b 6.01 and nHev b 6.02, enabled the identification of NRL allergy in 22 of 23 patients. There is a positive and significant correlation between sensitization to Hev b 5 and the number of interventions. BAT and allergen-specific IgE determination could be used as first-line in vitro diagnostic tests in patients with NRL allergy. [source] | |||||||||||||