Solid-phase Immunoassay Research Articles

Serological evidence of exposure to Herpes Simplex Virus type 1 is associated with cognitive deficits in the CATIE schizophrenia sample

Cognitive impairment is a core feature of schizophrenia. Previous studies have indicated that exposure to neurotropic infectious agents such as Herpes Simplex Virus type 1 may contribute to cognitive deficits and neuroanatomical abnormalities in individuals with schizophrenia. We examined the association between exposure to neurotropic infectious agents and cognitive function in 1308 participants in the Clinical Antipsychotic Trials of Intervention Effectiveness (CATIE) trial. This sample included all of the individuals in the CATIE trial for whom baseline blood samples were available. Cognition was evaluated at baseline by a test battery which yielded composite scores in the domains of processing speed, verbal memory, vigilance, reasoning, and working memory as well as a summary neurocognitive score. Solid phase immunoassay techniques were used to measure IgG class antibodies to Herpes Simplex Virus type 1 (HSV-1), Herpes Simplex Virus type 2 (HSV-2), Cytomegalovirus (CMV), and to Toxoplasma gondii ( T gondii) in the sera of the study individuals. We found a significant association between the neurocognitive summary score and antibodies to HSV-1 but not to HSV-2, CMV, or T. gondii. There was also a significant association between HSV-1 exposure and the Verbal Memory, Vigilance, and Processing Speed composite scores. HSV-1 may modulate the neurocognitive function of individuals with schizophrenia through its ability to establish latency in the central nervous system and undergo periodic reactivation. A better understanding of the role of HSV-1 may lead to better methods of treatment for the cognitive impairments associated with schizophrenia.

Supercritical Angle Fluorescence Immunoassay Platform

An inexpensive and easy-to-use immunoassay platform for the sensitive detection of analytes is presented. It comprises single-use polymer test tubes and a compact fluorescence reader. The optics for the capture of supercritical angle fluorescence (SAF) has been built into the tubes allowing for the extremely sensitive readout of solid phase immunoassays in real time and without washing steps. One-step sandwich immunoassays with interleukin 2 (IL-2) were carried out with capture antibodies immobilized in the tubes. At a turn around time of 12 min, the limit of detection for IL-2 was 0.27 pM (4.5 pg/mL) and the linear range covered 3 orders of magnitude. The developed technology is also adaptable to well plates and has great potential of replacing the work-intensive and time-consuming enzyme-linked immunosorbant assay (ELISA).

Donor human leukocyte antigen specific antibodies predict development and define prognosis in transplant glomerulopathy

The pathogenesis of transplant glomerulopathy (TG) remains unclear, with evidence of human leukocyte antigen (HLA) antibodies as important contributors to the disease. We studied the risk factors and the associations of HLA antibodies in the development of TG. Sixty-one cases with morphologic features of TG were identified and compared with contemporaneous matched patients (without TG) from a 17-year period, all undergoing renal biopsy in a single center. Univariate risk factors for TG were previous glomerulitis [odds ratio (OR) 3.3, 95% confidence interval (95% CI) [1.2–9.4], p = 0.025), delayed graft function (OR 2.3 [1.0–5.1], p = 0.042), HLA class I presensitization defined by Luminex solid-phase immunoassays (OR 5.0 [2.3–11.0]. p < 0.001), and de novo posttransplant development of donor HLA specific antibody (DSA) (OR 4.7 [1.7–13.2], p = 0.002). Only DSA remained significantly associated with TG after adjustment (OR 3.8 [1.1–12.9], p = 0.032). DSA was detected in >50% of TG patients, suggesting HLA antibodies play a critical role in TG pathogenesis. TG patients with DSA had increased risk of graft loss (median graft survival 4.4–5.2 years), whereas patients with morphologic features of TG without DSA had similar graft survival compared with the non-TG group (median graft survival 15 years). Thus, DSA is a useful predictor for graft failure in TG patients.

Competitive solid-phase immunoassay of gastrin in serum using time-resolved fluorometry

Background. A competitive solid-phase assay for the measurement of gastrin in serum using time-resolved fluorescence was developed as an alternative to conventional radioimmunoassay (RIA) technology. Methods. The assay depends on the competitive binding of unlabelled versus Eu-labelled gastrin to specific gastrin antibodies – bound to anti-rabbit IgG immobilized on polystyrene microtitration strips. The bound Eu3+-label was dissociated from the bound gastrin and converted to a fluorescent β-diketone chelate which was measured by fluorometry with time-resolution. Results. Using a sample volume of 50 μl the lower limit of detection was below10 pmol/L. Dilution of samples showed an excellent linearity. Spiking with gastrin-17 in known concentrations showed a recovery of 103% indicating that there is no bias inherent in the assay. The method correlated fully with the routine in-house radioimmunoassay in the concentration range 10–400 pmol/L (slope = 0.98 with r2 = 0.98). Thus the reference interval for clinical samples does not require modification when changing from one method to the other. Conclusion. We have described a convenient and accurate competitive assay for measurement of gastrin in serum based on solid-phase technology using time-resolved fluorometric detection as a realistic alternative to the established state-of-the-art RIA-technology.

Integrated microfluidic bioprocessor for solid phase capture immunoassays

A microfluidic device for solid-phase immunoassays based on microparticle labeling is developed using microvalve-control structures for automated sample processing. Programmable microvalve control in a multilayer structure provides automated sample delivery, adjustable hydrodynamic washing and compatibility with a wide range of substrates. Capture antibodies are derivatized on glass surfaces within the processor using an APTES patterning method, and magnetic microspheres conjugated with a secondary detection antibody are used as labels in a capture-sandwich format. In this microfluidic processor, washing force can be precisely controlled to remove the nonspecifically bound microparticles. Automated microfluidic immunoassays are demonstrated for mouse immunoglobulin (IgG) and human prostate specific antigen (PSA) with limits of detection of 1.8 and 3 pM, respectively. The sample processor architecture is easily parallelized for high-throughput analysis and easily interfaced with various assay substrates.

Immune activation by casein dietary antigens in bipolar disorder

Inflammation and other immune processes are increasingly linked to psychiatric diseases. Antigenic triggers specific to bipolar disorder are not yet defined. We tested whether antibodies to bovine milk caseins were associated with bipolar disorder, and whether patients recognized different epitopes of the casein protein than control individuals. Anti-bovine casein immunoglobulin G (IgG) levels were measured with solid-phase immunoassays in 75 individuals with bipolar disorder and 65 controls. Epitope recognition was evaluated in immunoassays by cross neutralization with anti-bovine casein polyclonal antibodies of defined reactivity. Group-specific reactivity and associations with symptom severity scores were detected with age-, gender-, and race-controlled regression models. Individuals with bipolar disorder had significantly elevated anti-casein IgG (t-test, p ≤0.001) compared to controls. Casein IgG seropositivity conferred odds ratios of 3.97 for bipolar disorder [n=75, 95% confidence interval (CI): 1.31-12.08, p ≤0.015], 5.26 for the bipolar I subtype (n=56, 95% CI: 1.66-16.64, p ≤0.005), and 3.98 for bipolar disorder with psychosis (n=54, 95% CI: 1.32-12.00, p ≤0.014). Lithium and/or antipsychotic medication did not significantly affect anti-casein IgG levels. Casein IgG measures correlated with severity of manic (R(2) =0.15, 95% CI: 0.05-0.24, p ≤0.02) but not depressive symptoms. Unlike controls, sera from individuals with bipolar disorder did not inhibit binding of casein-reactive animal sera (t-test/χ(2) , p ≤0.0001). Anti-casein IgG associations with bipolar I diagnoses, psychotic symptom history, and mania severity scores suggest that casein-related immune activation may relate to the psychosis and mania components of this mood disorder. Case-control differences in epitope recognition implicate disease-related alterations in how the casein molecule is digested and/or how resulting casein-derived structures are rendered immunogenic.

Altered IgE epitope presentation: A model for hypoallergenic activity revealed for Bet v 1 trimer

In order to reduce side effects in the course of allergen specific immunotherapy hypoallergenic allergen derivatives with reduced IgE reactivity have been made by genetic engineering. In contrast to other recombinant hypoallergenic allergen derivatives which showed reduced IgE reactivity, a recombinant trimer of the major birch pollen allergen Bet v 1 showed reduced allergenic activity despite preserved IgE reactivity. We studied rBet v 1 trimer by SDS-PAGE, mass spectrometry, circular dichroism and gel filtration. Furthermore we investigated IgE and IgG reactivity of the rBet v 1 trimer in solid and liquid phase assays and compared its allergenic activity with that of rBet v 1 wildtype using basophil activation assays. In solid phase immunoassays rBet v 1 trimer exhibited even stronger IgE reactivity than the rBet v 1 wildtype, whereas both proteins were equally well recognized by Bet v 1-specific IgG antibody probes. In fluid phase IgE experiments rBet v 1 trimer inhibited IgE reactivity to rBet v 1 wildtype but showed a more than 10-fold reduced allergenic activity compared to the rBet v 1 monomer. By analytical gel filtration it was demonstrated that, despite its monomeric appearance in SDS-PAGE the trimer occurred in fluid phase in the form of defined high molecular weight (>600kDa) aggregates whereas rBet v 1 wildtype strictly appeared as monomeric protein. The results indicate that the hypoallergenic nature of the rBet v 1 trimer is due to formation of defined high molecular weight aggregates which may be responsible for an altered presentation of IgE epitopes in a form with reduced capacity to crosslink effector-cell bound IgE. We thus provide evidence for a novel mechanism for hypoallergenic activity.

Relationship between Total Tear and Serum IgE in Allergic Conjunctivitis

Background: Recently, a new commercial test for total tear IgE based on immunochromatography (Allerwatch®) was developed. We examined the relationship between total tear and serum IgE levels with the Allerwatch test. Methods: A nonrandomized cross-sectional study was conducted in 40 patients with allergic conjunctivitis, 35 age- and sex-matched healthy control subjects, and 6 patients with epidemic keratoconjunctivitis (EKC). The total tear IgE score was determined with the Allerwatch test (0, 1 and 2), and the serum total IgE level was measured by solid-phase immunoassay (Phadezym PRIST) in all subjects. Results: Total tear IgE was assayed within 10 min of collection in all samples. The total tear IgE-positive rate was significantly higher in the allergic group than in the control and EKC groups (92.5 vs. 0.0 vs. 0.0%; p < 0.00001). Sensitivity and specificity were 0.925 and 1.000, respectively. The total IgE score and log (total serum IgE) were also higher in the allergic group than in the control and EKC groups [total IgE score: 1.48 ± 0.63 vs. 0.00 ± 0.00 vs. 0.00 ± 0.00, p < 0.00001; log (serum total IgE): 2.23 ± 0.50 vs. 0.80 ± 0.31 vs. 0.79 ± 0.37, p < 0.00001]. The total tear IgE score was significantly correlated with log (total serum IgE) in the allergic group (r = 0.712, p < 0.00001). Conclusions: These results show that this rapid test for the measurement of total tear IgE is easy to perform on an outpatient basis and may be helpful in the management of ocular allergy.

Integrated optical biosensor for in-line monitoring of cell cultures

An analytical detection platform was developed to evaluate the induced toxicity in cell cultures exposed to foreign agents like growth factors or nanoparticles. Connecting a biosensing detection device to the cell culture flasks allows analyzing the composition of cell medium in real-time. The analysis relies on the quantification of inflammatory cytokines released by cells into the cell culture medium, by means of solid-phase immunoassays analyzed with the wavelength interrogated optical sensing (WIOS) instrument. A fluidic system for in situ measurements allows detecting cytokines in real-time, with a sensitivity of 1–100 ng/mL depending on the cytokine. In addition, integration of an in-line optical absorbance measurement unit, in combination with the standard AB cell proliferation assay, provides information on the cell viability in the culture. Fluidic connections between the cell culture flasks, the optical biosensor and the absorbance measurement unit simultaneously allow quantifying up to three cytokines (interleukin 8, interleukin 6 and the monocyte chemotactic protein), assessing cellular proliferation, and thus discriminating between naïve cells and cells exposed to foreign agents such as growth factors (tumor necrosis factor alpha) or nanoparticles. This analytical tool presents a high potential for assessing the cytotoxicity of nanoparticles and other chemicals in vitro.

Molecularly Imprinted Polymers for Ochratoxin A Extraction and Analysis

Molecularly imprinted polymers (MIPs) are considered as polymeric materials that mimic the functionality of antibodies. MIPs have been utilized for a wide variety of applications in chromatography, solid phase extraction, immunoassays, and sensor recognition. In this article, recent advances of MIPs for the extraction and analysis of ochratoxins are discussed. Selection of functional monomers to bind ochratoxin A (OTA) with high affinities, optimization of extraction procedures, and limitations of MIPs are compared from different reports. The most relevant examples in the literature are described to clearly show how useful these materials are. Strategies on MIP preparation and schemes of analytical methods are also reviewed in order to suggest the next step that would make better use of MIPs in the field of ochratoxin research. The review ends by outlining the remaining issues and impediments.

Reactivity with dichotomous determinants of Ro 60 stratifies autoantibody responses in lupus and primary Sjögren's syndrome

Analysis of B cell determinants of Ro 60 exposed on the surface of apoptotic cells (apotopes) or intracellular epitopes provides insight into the structural forms of the autoantigen that break immune tolerance. This study was initiated to compare anti-Ro 60 responses in systemic lupus erythematosus (SLE) and primary Sjögren's syndrome (SS) against membrane-bound and intracellular forms of Ro 60. The reactivity of autoantibodies from patients with SLE and primary SS to Ro 60 apotopes and epitopes was assessed by multiparameter flow cytometry and solid-phase immunoassay. Anti-Ro 60 IgG was eluted from early apoptotic cells or recombinant Ro 60 immobilized on nitrocellulose, and binding to membrane-bound and intracellular forms of Ro 60 was quantitated by flow cytometry. An immunodominant apotope, which was recognized by IgG from a subset of SLE patients with anti-Ro, but not anti-La, autoantibodies, was mapped to a region forming a helix-loop-helix at the apical tip of the Ro 60 molecule. Immobilization of this region to the solid phase exposed an epitope that was recognized by IgG from primary SS and SLE patients whose sera had both anti-Ro and anti-La autoantibodies. Autoantibodies eluted from either the surface of apoptotic cells or the Ro 60 epitope on the solid phase were non-cross-reactive and specifically recognized membrane-bound or cytoplasmic forms of Ro 60. This is the first example of a dichotomy of human autoantibody responses against mutually exclusive determinants linked to a single domain of a systemic autoantigen and supports a model in which tolerance is broken by different immunogenic forms of Ro 60.

Detection of proteases using an immunochemical method with haptenylated–gelatin as a solid-phase substrate

A simplified method for the measurement of proteases utilising solid-phase substrates incorporating an ELISA end-point detection step is described. Gelatin-hapten conjugates adsorbed onto polystyrene surfaces were found to be efficient substrates for proteases. Digestion of the solid-phase protein-hapten complexes resulted in proportional desorption of the attached conjugates and decrease in the detectable hapten species. Gelatin-cholic acid conjugates, affinity-purified sheep anti-cholic acid antibody-HRP and a chromogenic substrate were incorporated into a convenient and highly sensitive solid-phase immunochemical method. The detectable signal is inversely proportional to enzyme activity. Bacterial proteases (alpha-chymotrypsin Type II, Type IX from Bacillus polymyxa, Type XIV from Streptomyces griseus, Type XXIV from Bacillus licheniformens) were assayed. Dose-response curves for enzyme activities were measured within ranges of 0-550 microunits mL(-1) for chymotrypsin, 0-12 microunits mL(-1) for type IX, 0-35 microunits mL(-1) for type XIV and 0-100 microunits mL(-1) for type XXIV. The detection limits of the proteases studied were 89 microunits mL(-1) for chymotrypsin, 0.26 microunits mL(-1) for type IX, 5.8 microunits mL(-1) for type XIV and 6.5 microunits mL(-1) for type XXIV. It was demonstrated that the two-step immunochemical method combines the simplicity and sensitivity of solid-phase enzyme immunoassays, the broad specificity of gelatin as a protease substrate and the flexibility of the solid-phase format.

Sex and Ethnic Differences in 47 Candidate Proteomic Markers of Cardiovascular Disease: The Mayo Clinic Proteomic Markers of Arteriosclerosis Study

BackgroundCardiovascular disease (CVD) susceptibility differs between men and women and varies with ethnicity. This variability is not entirely explained by conventional CVD risk factors. We examined differences in circulating levels of 47 novel protein markers of CVD in 2561 men and women of African-American (AA) and non-Hispanic White (NHW) ethnicity, enrolled at geographically distinct sites.Methodology/Principal FindingsParticipants (1,324 AAs, mean age 63.5 y, 71% women; 1,237 NHWs, mean age 58.9 y, 57% women) belonged to sibships ascertained on the basis of hypertension. Solid-phase immunoassays and immunoturbidometric, clot-based, chromogenic, and electrophoretic assays were used to measure the 47 protein markers in plasma or serum. Marker levels were log transformed and outliers were adjusted to within 4 SD. To identify markers independently associated with sex or ethnicity, we employed multivariable regression analyses that adjusted for conventional risk factors, prior history of CVD, medication use and lifestyle factors (physical activity, alcohol consumption and education). Generalized estimating equations were used to correct for intrafamilial correlations. After adjustment for the above covariates, female sex was associated with higher levels of 29 markers and lower levels of 6 markers. Female sex was independently associated with higher levels of several inflammatory markers as well as lipoproteins, adipokines, natriuretic peptides, vasoconstrictor peptides and markers of calcification and thrombosis. AA ethnicity was associated with higher levels of 19 markers and lower levels of 6 markers, including higher levels of several inflammatory makers, higher leptin and lower adiponectin levels, lower levels of vasodilator-natriuretic peptides, higher levels of vasoconstrictor-antidiuretic peptides and markers of calcification and thrombosis.Conclusions/SignificancePlasma levels of several novel protein markers of CVD differ significantly in the context of sex and ethnicity. These results have implications for individualized CVD risk assessment.

A highly sensitive electrochemiluminescence immunoassay for the neurofilament heavy chain protein

Background The loss of neurological function is closely related to axonal damage. Neurofilament subunits are concentrated in neurons and axons and have emerged as promising biomarkers for neurodegeneration. Electrochemiluminescence (ECL) based assays are known to be of superior sensitivity and require less sample volume than conventional ELISAs. Methods We developed an ECL based solid-phase sandwich immunoassay to measure the neurofilament heavy chain protein (NfH SMI35) in CSF. We employed commercially available antibodies as previously used in a conventional ELISA (Petzold et al., 2003; Petzold and Shaw, 2007). The optimised and validated assay was applied in a reference cohort and defined patient groups. Results Analytical sensitivity (background plus three SD) of our assay was 2.4 pg/ml. The mean intra-assay coefficient of variation (CV) was 4.8% and the inter-assay CV 8.4%. All measured control and patient samples produced signals well above background. Patients with multiple sclerosis (MS) (median 46.2 pg/ml, n = 95), amyotrophic lateral sclerosis (ALS) (160.1 pg/ml, n = 50), mild cognitive impairment/Alzheimer's disease (MCI/AD) (65.6 pg/ml, n = 20), Guillain–Barre syndrome (GBS) (91.0 pg/ml, n = 20) or subarachnoid hemorrhage (SAH) (345.0 pg/ml, n = 20) had higher CSF NfH SMI35 values than the reference cohort (27.1 pg/ml, n = 73, p < 0.0001 for each comparison). Conclusion The new ECL based assay for NfH SMI35 in CSF is superior in terms of sensitivity, precision and accuracy to previously published methods (Petzold et al., 2003; Shaw et al., 2005; Teunissen et al., 2009). The improved performance and small sample volume requirement qualify this method in experimental settings and clinical trials designed to perform a number of tests on limited amounts of material.

Antiallograft antibodies: relevance, detection, and monitoring

Solid-phase immunoassays increase the accuracy of assessing pretransplant immunologic risk and facilitate posttransplant prediction and diagnosis of antibody-mediated rejection (AMR). This review will describe methods available for antibody analyses, discuss the types of targets of AMR and the characteristics of pathogenic alloantibodies, and provide guidelines for the application of antibody tests in the prediction of rejection risk and diagnosis of rejection. The presence of human leukocyte antigen-specific antibodies increases the risk of AMR, but the clinical relevance of low antibody levels is questioned with reports of stable graft function in their presence. Posttransplant monitoring has been shown to provide early diagnosis of AMR permitting preemptive intervention. Antibodies to other alloantigens and autoantigens are being implicated as potential targets for both acute and chronic AMR. Certain limitations and interfering factors have also been recognized that should be recognized in the interpretation of solid-phase antibody assay results. Contemporary technology is clearly advancing the detection of various antibodies that can contribute to AMR, but continued work is needed to elucidate the relevance of very low levels of human leukocyte antigen-specific antibody and the importance of antibodies to other alloantigens and autoantigens.

Immunogenetics and immunology in transplantation

Advances in immunogenetics and histocompatibility have facilitated the clinical transplantation of solid organs and tissues. Improved definition of HLA antigens, alleles, and haplotypes has clarified the diversity of the HLA system among different racial/ethnic populations. Knowledge of allele and haplotypes frequencies derived from these studies can be applied to the estimates of transplant compatibility and the likelihood finding suitable donors. Humoral sensitization that has been a major barrier to transplantation is being successfully treated with desensitization and paired donor exchanges. Sensitive solid phase immunoassays for HLA-specific antibodies permit analyses that guide both desensitization and donor exchanges through antibody monitoring, virtual crossmatches, and detection of cryptic sensitization. Ongoing studies are investigating the mechanisms underlying the down-regulation of donor HLA-specific antibodies.

Virtual crossmatch by identification of donor-specific anti-human leukocyte antigen antibodies by solid-phase immunoassay: A 30-month analysis in living donor kidney transplantation

Selection of donors for kidney transplantation depends on accurate prediction of risk factors for immunologic rejection. Historically, cytotoxicity crossmatch (CXM) examining lysis of donor cells by preformed anti-human leukocyte antigen (HLA) antibodies (Abs) has been considered the best predictor of immunologic rejection. However, there is much interest in defining anti-HLA Ab specificity in recipient sera by immunoassay to predict crossmatch results and aid in donor selection. Current immunoassays for anti-HLA Abs are highly sensitive, though correlation between Abs detected by immunoassay and their functional relevance in CXM and subsequent transplantation is not well defined. In this study, we retrospectively examined the predictive value of detection of donor-specific anti-HLA Abs (DSA) by Luminex Single Antigen assay from 149 consecutive living donor kidney transplant recipients. We demonstrate that detection of DSA by immunoassay accurately predicted negative crossmatch and graft survival. However, this approach had limited sensitivity for predicting positive crossmatch, attributable to either limited typing of donor HLA-DQ and -DP alleles or due to non-HLA Abs. False-positive prediction of CXM correlated with detection of “weak” Abs with low mean fluorescence intensity (MFI < 2000). Furthermore, we found that a ratio of the MFI of the DSA bead to the MFI of the positive control bead was a better method for identifying weak DSA that did not result in CXM-positive reactions. Interestingly, patients with weak DSA and negative CXM had equivalent graft survival over an 18 month follow-up period, suggesting that weak DSA may not preclude transplantation.

Serum Markers of Apoptosis in Traumatic and Ischemic Brain Injury

Objective: to study the time course of changes and relationship of the serum indicators of apoptotic processes in neurore-suscitation patients. Subjects and methods. Thirty-eight neuroresuscitation patients, including 14 patients with severe brain injury (SBI) (mean age 41.4±4.3 years) and 24 patients with strokes (mean age 53.8±2.5 years), were examined. The group of patients with strokes was divided into 2 subroups: 1) 11 patients with ischemic strokes (IS) and 2) 13 with hemorrhagic strokes (HS). The Glasgow coma scores for admission consciousness loss were 7.6±0.8 in the SBI group and 9.5±0.7 in the stroke group; mortality was 28.6 and 37.5%, respectively. A control group included 16 subjects (mean age 47.9±3.8 years). The investigators measured the serum levels of FAS antigen and its ligand (sAPO-I/FAS and sFAS-L), cas-pase-1/ICE, sCD40 (Bender MedSystem, Austria) and hTRAIL (Biosource, Belgium) by solid-phase immunoassay in neuroresuscitation patients on days 1, 7, and 14 of the acute period of diseases. They used statistical methods, such as Wilcoxon-Mann-Whitney U-test, Spearman’s rank correlation test. Results. A reduction in hTRAIL was observed in all the groups. There was a decrease in serum sCD40 in strokes on days 1 to 14 and in SBI on days 7 to 14. An increase in caspase 1/ICE was seen in HS in the first 24 hours, in IS on days 1 to 7, and in SBI on days 1 to 14. The most pronounced rise in caspase-1/ICE was induced by ischemic brain lesion within the first week of disease. A prolonged increase up to 2 weeks was noted in SBI. No rise in serum FAS-L was found in the examinees. The time course of changes in sAPO-I/FAS was different in all the groups. The most marked, moderate, and none reductions were revealed in HS, IS, and SBI, respectively. There was a pronounced serum sAPO-I/FAS increase in SBI within the first 24 hours. Assessment of correlations between the serum indicators of apoptosis revealed that there were differences in the association between the indices under study in all the patient groups and in the control group. Conclusion. There are general features and differences in the time course of changes in serum apoptotic markers and their association in the acute period of SBI, IS, and HS. Key words: severe brain injury, stroke, apoptosis, APO-I/FAS, FAS-L, caspase-1/ICE, CD40, hTRAIL.

Immunogenicity studies of guayule and guayule latex in occupationally exposed workers

Type I Hevea brasiliensis rubber latex allergy is managed by avoidance, using synthetic and alternative latex (such as Parthenium argentatum, guayule) products. This study investigates the ability of high-dose occupational exposure to guayule shrub/homogenate/latex to induce guayule-specific antibody responses in employees (Yulex Corporation). Participants completed an allergy history/guayule exposure questionnaire and provided annual blood samples from 2006 to 2008. Sera were analyzed for IgG and IgE anti-guayule (protein from homogenate, commercial-grade latex and non-ammoniated total plant proteins) using solid phase immunoassays (negative = IgG < 1 μg/ml, IgE < 1 ng/ml). Guayule-specific IgG antibody (range: 2.0–9.7 μg/ml) was detected in 3 of 16 (19%) highly exposed employees in the pilot plant and R&D/applications laboratory. Antibody levels related to relative cumulative-years (e.g. >3) of reported guayule homogenate/latex exposure. Equivocal IgG antibody responses (1.0–2.0 μg/ml) were detected in 2 of 5 (40%) of administrators with infrequent guayule homogenate/latex contact. No guayule-specific IgE antibody or guayule-associated allergic reactions were detected. We conclude that protein from guayule and in guayule latex can be immunogenic but not allergenic in occupationally exposed workers.

61-P: Interference in solid phase immunoassays: Identifying candidates prior to testing

61-P: Interference in solid phase immunoassays: Identifying candidates prior to testing