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Results for “Biomarker variability”

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PubMed1997

Fluctuations in GAD65 antibodies after clinical diagnosis of IDDM in young children.

OBJECTIVE: To investigate whether the presence of GAD antibodies at onset of IDDM correlates to a more aggressive rate of beta-cell destruction after clinical onset. RESEARCH DESIGN AND METHODS: We studied GAD antibodies at onset of disease, after 1 year, and after 6 years in 33 consecutively referred children (mean age 8.08, range 1.7-16.3). In a subset of 11 patients, GAD antibodies were studied very frequently. The correlation between GAD antibodies and clinical parameters, including glycosylated hemoglobin, residual insulin secretion, and insulin dosage, was evaluated. RESULTS: GAD antibody titers were highly variable. Four patients became GAD antibody positive weeks to years after clinical onset. Other patients switched between testing positive and negative for GAD antibodies shortly after clinical onset. No correlation was found between the presence of GAD antibodies and the rate of beta-cell destruction, but patients with high GAD antibody indexes at onset had significantly higher glycosylated hemoglobin levels. CONCLUSIONS: GAD antibodies at clinical onset do not predict the rate of beta-cell destruction in young children with newly diagnosed IDDM. The highly variable GAD antibody levels suggest variation of the autoimmune process.

Autoantibodies↗

PubMed2001

Genotype and phenotype in susceptibility to coal workers' pneumoconiosis. the use of cytokines in perspective.

The readily available technique of screening for gene polymorphisms could be used to explain inter-individual variability in a classic occupational interstitial lung disease, such as coal workers' pneumoconiosis (CWP). The objective of this paper is to describe candidate genes selected from the wide pool of cytokines and growth factors, and to discuss the applications and pitfalls when using them as biomarkers for susceptibility to CWP. The selection of candidate genes is mainly based on observed phenotypic changes in bronchoalveolar lavage (BAL) fluid or BAL cells of patients with CWP, or on animal experiments that use quartz as the fibrogenic agent. This paper also reviews the studies that have been performed to validate tumour necrosis factor genotype and phenotype with respect to CWP. Finally, it is proposed that a multiple marker approach to susceptibility to CWP should be used. This involves the measurement of two cytokines (tumour necrosis factor and transforming growth factor-beta) to improve denomination of high- and low-risk groups.

Biomarkers↗

PubMed2005

The investigation of primary rectal cancer by surgeons: current pattern of practice.

OBJECTIVE: Selection of the optimal treatment strategy for patients with rectal cancer requires appropriate investigation, but published guidelines provide no clear consensus. We examined the current practice pattern for the investigation of primary rectal cancer by general surgeons in the province of Ontario, Canada. METHODS: A telephone interview was completed by 124 surgeons in Ontario who manage patients with rectal cancer, who indicated the investigations they routinely perform in assessment before treatment. An exploratory cluster analysis was used to identify surgeon-related variables that predicted the pattern of investigation; these were tested in univariate and multivariate analyses. RESULTS: Cluster analysis identified 3 distinct groups of surgeons based on patterns of test usage. Univariate analysis showed that the use of chest radiography, computed tomography of the abdomen and pelvis, and ultrasound of the abdomen varied significantly with the surgeon's subspecialty training, practice location and years in practice. Regression analysis confirmed that each of these 3 variables independently predicted the pattern of preoperative investigation. There were no significant predictors of the use of colonoscopy, sigmoidoscopy or carcinoembryonic antigen level. Over half of surgeons reported that they would have ordered additional imaging tests but did not because of lack of availability. The perceived desirability of additional tests, endorectal ultrasound in particular, varied with training. CONCLUSIONS: The current practice pattern for the preoperative investigation of primary rectal cancer by general surgeons varies significantly with specific surgeon-related variables, with potential impact on the management and outcome of patients with rectal cancer.

Adult↗

PubMedDate not supplied

Analysis of factors that affect short-term blood pressure variability in patients with chronic renal failure.

Recent reports suggest the relationship of short-term blood pressure (BP) variability to cardiovascular target organ damage. In this study, short-term BP variability was assessed as the standard deviation of daytime and nighttime BP in 36 hospitalized patients with chronic renal failure (CRF) who underwent ambulatory BP monitoring. Positive correlations were observed between body mass index (BMI) and daytime systolic and diastolic BP variability, BMI and nighttime diastolic BP variability, cholesterol and daytime systolic BP variability, cholesterol and nighttime systolic and diastolic BP variability, nocturnal decline in BP and nighttime diastolic BP variability, and plasma concentration of norepinephrine (p-NE) and nighttime systolic BP variability. In multivariate linear regression analyses, BMI showed the strongest association with daytime and nighttime diastolic BP variability (p < .005 and p < .05). On the other hand, cholesterol and p-NE were the primary determinants of daytime and nighttime systolic BP variability, respectively (p < .01 and p < .0005). Interestingly, CRF patients with ischemic heart disease (IHD) had significantly increased daytime systolic and diastolic BP variability and nighttime systolic BP variability (p < .05 or less). Furthermore, logistic regression analysis demonstrated that nighttime systolic BP variability was an independent risk factor of IHD in patients with CRF (odds ratio 1.50 [95% confidence interval 1.01 to 2.25]; p < .05). Taken together, short-term BP variability is suggested to be affected by BMI, cholesterol, and p-NE in CRF patients. Furthermore, sympathetic nerve overactivity may be involved in cardiovascular complications in CRF patients through the increase in nighttime systolic BP variability.

Aged↗

PubMed2006

Plasma level of CXC-chemokine CXCL12 is increased in rheumatoid arthritis and is independent of disease activity and methotrexate treatment.

OBJECTIVE: Several actions of the chemokine CXCL12 have potential relevance for rheumatoid arthritis (RA). Interaction with CXCR4, the unique receptor for CXCL12, stimulates angiogenesis, mononuclear cell trafficking into the joints, lymphoid-tissue-like rearrangement of T cells within the synovium, and chondrocyte release of cartilage-degrading metalloproteinases. We investigated the level of CXCL12 in plasma (p-CXCL12) as a marker of RA diagnosis, RA disease activity, and response to methotrexate (MTX) treatment. METHODS: A prospective study including 36 patients with RA (ACR criteria) of at least 6 months' duration, and 50 sex and age matched healthy controls. ELISA for CXCL12 was performed on plasma prior to and after 16 and 28 weeks of MTX treatment in the patients with RA and once in controls. RESULTS: The p-CXCL12 was 1855 +/- 145 pg/ml in RA patients and 1273 +/- 79 pg/ml in controls (p < 0.001). During the 28 weeks of MTX treatment, the ACR disease activity variables decreased, whereas the p-CXCL12 level remained constant and increased. P-CXCL12 was not correlated to any ACR disease activity variable at any time (p > 0.05). CONCLUSION: Patients with RA had a significantly and constantly increased p-CXCL12 level compared to controls. The p-CXCL12 level was independent of any ACR disease activity variables, as well as response to MTX treatment.

Adult↗

PubMedDate not supplied

Atypical (anaplastic) meningioma: relationship between histologic features and recurrence--a clinicopathologic study.

Two-hundred-and-eighty meningiomas of the surgical pathological files of the Taipei Veterans General Hospital from the period 1976-1986 were reviewed by the authors without prior knowledge of clinical circumstances or outcome. Thirty-four cases were regarded as atypical or anaplastic based on high cellularity, pleomorphism and the presence of mitotic figures with 6 cases showing only the above features and the remaining 28 displaying in addition one or more of the following "ominous" variables: papillary formation, necrosis and invasion of the underlying brain. With a median follow-up of three years after surgery the recurrence rate was 44% (15 cases) for this group of tumors whereas the remaining 246 histologically benign meningiomas had a 6% recurrence rate during the same period. This difference was statistically significant (p less than 0.0001). Once the atypical or anaplastic character of a meningioma was established, no difference in the recurrence rate was found related to the number of mitoses and whether the additional ominous variables were present, alone or in conjunction with others. Immunostaining for vimentin, S100, fibronectin and EMA showed variable results in the 34 atypical meningiomas but without significant difference between those that recurred within 3 years and the ones that did not.

Adolescent↗

PubMed1998

Do prognostic marker studies on core needle biopsy specimens of breast carcinoma accurately reflect the marker status of the tumor?

Core needle biopsies (CNB) are often used for the diagnosis of breast lesions. In some breast cancer patients, e.g., those treated with preoperative chemotherapy, the CNB specimen might be the only pretreatment tissue sample available for studies of prognostic and predictive markers. Our purpose was to evaluate whether marker studies performed on CNB specimens accurately reflect the marker status of the tumor. Immunostaining for five commonly used prognostic and predictive markers was performed on both CNB and subsequent excision specimens from 56 consecutive patients who had a CNB with carcinoma followed by excision of the tumor. None of the patients received radiotherapy or chemotherapy between the CNB and the excision. Paraffin sections of the CNB and excision specimens were immunostained for bcl-2, estrogen receptor (ER), c-erbB-2, and p53. These markers were scored as positive or negative. Microvessel density (MVD) was scored as a continuous variable on sections immunostained for Factor VIII-related antigen by calculating the average number of microvessels in three 224x fields of highest tumor vascularity ("hot spots"). Immunostaining results for bcl-2, ER, c-erbB-2, and p53 on the CNB and the corresponding excision specimens were 100% concordant. Although there was significant correlation between MVD on the CNB specimens and the corresponding excisions (r = 0.507, P = 0.0002), the mean MVD on the CNB and corresponding excision specimens differed by more than 10% in 85.7% of cases, with differences ranging from 4.3 to 233.3%. MVD was higher in the CNB than in the excision specimens in 30 (61.2%) of 49 cases. In conclusion, in all of the cases studied, accurate results for the dichotomously scored markers bcl-2, ER, c-erbB-2, and p53 were obtained on CNB specimens. In contrast, in most cases, MVD, which was scored as a continuous variable, could not be reliably assessed on the CNB specimen.

Biomarkers, Tumor↗

PubMed1988

Lymphocyte proliferative response to PHA and anti-CD3/Ti monoclonal antibodies, T cell surface marker expression, and serum IL-2 receptor levels as biomarkers of age and health.

Alteration of T cell surface marker expression with a decrease of CD3 positive cells relative to the number of CD4 and CD8 positive cells, diminished in vitro proliferative response to mitogenic stimuli like PHA and antibodies to the CD3/Ti complex, and increase in serum IL-2 receptor levels, are among the changes in immunologic parameters that have been associated with advanced age. To distinguish between effects of the primary aging process and diseases of aging not known to be directly related to immune function, we investigated these variables in two well characterized populations of elderly donors (greater than 70 years) and a young adult control group (less than 35 years). The first group of older donors reported no evidence of significant chronic or recent acute illness and saw a physician only for routine medical care. The second group was randomly selected from individuals seen in a geriatric medicine clinic for diagnoses that included osteoarthritis and cardiopulmonary disorders. Altered surface marker expression and increased serum IL-2 receptor levels were seen only in the second group. On the other hand, lymphocyte proliferative responses to PHA, Leu 4 (anti-CD3) and a monoclonal antibody to the beta-chain of the T cell antigen receptor (WT31) were significantly decreased in both populations. Because we would expect primary aging to affect even extremely fit individuals of advanced age, we conclude that decrease in T cell proliferative response may represent a biomarker of primary aging in man. The alteration in surface marker expression and increased IL-2R levels in serum appear to be effects secondary to non-immunologic disease rather than aging.

Adult↗

PubMed2006

Finding regions of significance in SELDI measurements for identifying protein biomarkers.

MOTIVATION: There is a well-recognized potential of protein expression profiling using the surface-enhanced laser desorption and ionization technology for discovering biomarkers that can be applied in clinical diagnosis, prognosis and therapy prediction. The pre-processing of the raw data, however, is still problematic. METHODS: We focus on the peak detection step, where the standard method is marked by poor specificity. Currently, scientists need to inspect individual spectra visually and laboriously in order to verify that spectral peaks identified by the standard method are real. Motivated by this multi-spectral process, we investigate an analytical approach-called RS for 'regions of significance'-that reduces the data to a single spectrum of F-statistics capturing significant variability between spectra. To account for multiple testing, we use a false discovery rate criterion for identifying potentially interesting proteins. RESULTS: We show that RS has better operating characteristics than several existing methods and demonstrate routine applications on a number of large datasets.

Algorithms↗

PubMed2007

Using exposure-response and biomarkers to streamline early drug development.

Biomarkers (BMs) are biological measures of PD drug effects or disease markers that may represent clinically significant patient outcomes, either efficacy or toxicity. Their use in drug development, especially as an integral part of PK/PD modeling, has become a popular strategy for optimizing development time and resources. This approach supports quantitative integration of information across different species and throughout the clinical phases I-III. If the BM is based on the mechanism of action (MOA) of the drug, it is expected to follow an exposure-response relationship (E-R). If it is also involved in causal pathways in the pathophysiology of the disease (POD), it may become a surrogate marker (SM). SMs allow prediction of clinical outcomes for different dosing regimens of drug candidates and patient individualization of treatment in clinical practice. Appropriate evaluation of BMs by mechanistic, epidemiological, and clinical pharmacology studies as part of the drug development process allow scientists to establish clinically relevant ER. In early drug development, known ERs for BMs facilitate translation of in vitro findings to in vivo consequences, interspecies PK/PD comparisons, and streamlining of dose-finding phase I and II studies, as well as assessment of new dosing regimen candidates for their likely clinical efficacy and safety, extrapolation of clinical study results to special populations (e.g., pediatrics), and interpretation of exposure differences found in food, drug interaction and special populations studies. Recently, two novel BMs, namely, P50, a measure of ex vivo/in vitro whole blood oxygen affinity and S(pO2), i.e., in vivo pulse oximetry, were used in the development of an allosteric synthetic hemoglobin modifier (SAM), efaproxiral, as PD endpoints; these BMs are based on the MOA of SAMs. Early use of these BMs established excellent in vitro/in vivo PK/PD correlations, appropriate interspecies PK and PD scaling as well as PD-guided phase I and II dose-finding studies. This approach allowed appropriate translation of in vitro and preclinical information along with early identification of sources of PK/PD variability. Frontloading drug development with the identification and use of mechanism-based (MOA/POD) BMs constitutes a rational strategy to quantitatively integrate PK/PD information and optimize dose finding.

Allosteric Site↗

PubMed2006

Biomarkers of potential prognostic significance in diffuse large B-cell lymphoma.

Diffuse large B-cell lymphoma (DLBCL) is a biologically heterogeneous disease for which the current approach to treatment is only successful for 50% of patients. The prognostic value of various clinical and biological factors in predicting treatment outcome is discussed in this paper. A review of the English literature was performed including original articles and relevant reviews from MEDLINE that addressed the topics of DLBCL biology and potential prognostic factors. The International Prognostic Index is, to date, the most successful clinical model for predicting outcome. In addition, a rapidly expanding list of molecules has been identified by conventional and newer diagnostic methods that may be of significant prognostic value. Gene expression profiling has led to the discovery of new biological subtypes of DLBCL based on patterns of gene expression, and a host of new genes that may play important roles in this disease. Various derangements in apoptosis, cell-cycle regulation, differentiation, and signal transduction have been noted, while the host environment and immune response also appear to modify clinical outcome. Although to our knowledge, the fundamental abnormalities underlying DLBCL remain elusive, progress is being continuously made to further the understanding of the biological heterogeneity of this disease and the use of various clinical and biological variables to predict treatment outcome. The goal is to be able to identify subgroups of patients at high risk of treatment failure and develop more effective treatment based on specific biological defects that may represent new rational therapeutic targets.

Biomarkers, Tumor↗

PubMedDate not supplied

Pesticide exposure and creatinine variation among young children.

Pesticide exposure may differentially impact young children; they live closer to the ground and take in greater amounts of food relative to body mass than older children or adults. We are using an organophosphate (OP) urinary biomarker screen (gas chromatography with flame photometric detection, GC/FPD) to evaluate pesticide exposure among 154 children < or = 6 years of age living in a heavily farmed border (US-Mexico) community. The screen detects diethylphosphates (DEPs) and dimethylphosphates (DMPs) above a reference range of 1000 non-occupationally exposed individuals (DL=25 microg/g creatinine, Cr). At least one metabolite was detected for 33% of the subjects; many samples contained multiple biomarkers. DEP was detected in 5% of the subjects. DMP and DMTP were frequently measured (25% and 26%, respectively). Biomarker concentrations are adjusted by the body's metabolism of Cr as an indicator of urine dilution. Cr concentrations were examined separately to evaluate their effect on internal dose measures. Cr concentrations were significantly different by season (K-W=0.83, P=0.022). Significant differences exist between the autumn:spring (P=0.038) Cr concentrations and between summer:autumn (P=0.041) Cr concentrations based on Mann-Whitney U=1070.5, z=-2.041, (P=0.041). Our analysis of NHANES III data did not reflect seasonal Cr differences for 6 year olds. No younger children were included. Absorbed daily dose (ADD) estimates were calculated for children with the highest concentrations of metabolite. Calculations are theoretical values assuming that the entirety of a given metabolite was metabolized from a single pesticide. Several class appropriate pesticides were evaluated. For the children with the highest levels, almost all estimated ADDs exceeded the RfD. Although the actual metabolite concentrations dropped appreciably, ADD were still exceeded RfDs at the 95th percentile. The urinary OP screen was effective in identifying subjects with atypical internal doses. Daily Cr yield is a critical component in ADD calculations. Cr variability produces differences in internal dose measurement and estimates of ADD independent of exposure. Cr variability among young children needs to be examined, and caution should be applied when evaluating Cr adjusted internal doses for children.

Agriculture↗

PubMed2004

Novel chromatic and structural biomarkers of diet in carotenoid-bearing plumage.

Previous attempts to establish a link between carotenoid-based plumage reflectance and diet have focused on spectral features within the human visible range (400-700 nm), particularly on the longer wavelengths (550-700 nm) that make these plumages appear yellow, orange or red. However, carotenoid reflectance spectra are intrinsically bimodal, with a less prominent but highly variable secondary reflectance peak at near-ultraviolet (UV; 320-400 nm) wavelengths visible to most birds but not to normal humans. Analysis of physical reflectance spectra of carotenoid-bearing plumages among trophically diverse tanagers (Thraupini, Emberizinae, Passeriformes) indicated that both the absolute and relative (to long visible wavelengths) amounts of short waveband (including UV) reflectance were lower in more frugivorous species. Striking modifications to the branched structure of feathers increased with frugivory. These associations were independent of phylogenetic relatedness, or other physical (specimen age, number of carotenoid-bearing patches) or ecological (body size, elevation) variables. By comparison, reflectance at longer visible wavelengths ('redness') was not consistently associated with diet. The reflectance patterns that distinguished frugivores should be more apparent to UV-sensitive birds than to UV-blind humans, but humans can perceive the higher plumage gloss produced by modified gross feather structure. Basic aspects of carotenoid chemistry suggest that increases in pigment concentration and feather dimensions reduce short waveband reflectance by the plumages of frugivores.

Animals↗

PubMed1998

The clinical predictivity of biomarkers of stage III-IV epithelial ovarian cancer in a prospective randomized treatment protocol.

BACKGROUND: The aim of this study was to define the clinical relevance of functional biomarkers, prospectively assessed in a randomized clinical protocol, in patients with Stage III-IV epithelial ovarian cancer. The protocol compared cisplatin with polychemotherapy that included cisplatin and cyclophosphamide. METHODS: In a subset of 168 patients with invasive epithelial ovarian cancer cell proliferation was determined by the 3H-thymidine labeling index, DNA ploidy was assessed by flow cytometry, and the expression of p53, bcl-2, and glutathione S-transferase-pi (GST-pi) was evaluated by immunohistochemistry using the antibodies PAb1801, anti-bcl-2, and GST-pi, respectively. RESULTS: Cell proliferation, DNA ploidy, and the expression of p53, bcl-2, and GST-pi were generally unrelated to one another and unrelated to clinicopathologic features, except for an association between DNA ploidy and the rate of cell proliferation. All biologic variables except bcl-2 were slightly related to tumor grade. DNA ploidy emerged as a predictor of clinical complete response and 3-year overall survival, regardless of treatment type or residual disease. Conversely, except for a favorable outcome for patients with tumors not expressing bcl-2 who were treated with cisplatin, no definitive patterns of predictivity for short term or long term clinical outcomes were observed for the other biomarkers studied. CONCLUSIONS: DNA ploidy appears to be the most clinically relevant biomarker for epithelial ovarian cancer. More information is needed to understand the role of the other markers studied in this tumor type.

Aged↗

PubMed2005

Ethoxyresorufin-O-deethylase (EROD) activity in dab (Limanda limanda) as biomarker for marine monitoring.

GOAL, SCOPE AND BACKGROUND: Ethoxyresorufin-O-deethylase (EROD) is a well-accepted marker for biological effects in fish and is, therefore, part of numerous monitoring programmes. EROD activity was measured in livers of dab (Limanda limanda) from the German Bight (North Sea) from 1995 to 2003. The aim of the present long-term study was (a) time trend monitoring of EROD activity of dab from the German Bight and (b) to elucidate the needs for a successful application of EROD activity as an early warning system. METHODS: EROD activities were determined fluorimetrically in dab liver microsomes, using resorufin as an external standard. The limit of detection (LD) and the limit of quantification (LQ) were calculated. Results were referred to protein concentrations. RESULTS AND DISCUSSION: EROD activities of 610 female dab caught in different seasons between January 1995 and August 2003 were analyzed individually. Activities varied from < LD to 1768 pmol/(min mg protein) and showed an annual cycle as well as significant differences between the years. Highest EROD activities were observed in early summer and lowest activities during the winter period. In autumn 2002, significantly elevated EROD activities were detected, possibly related to effects of the River Elbe flood event. Two scenarios with different EROD baseline data are presented to discuss the prerequisites for the use of EROD as a monitoring tool. The comparison of these scenarios underlines the importance of appropriate season-specific baseline data. CONCLUSION: The use of EROD as an early warning tool for contaminant effects in dab in the German Bight has different prospects during the year, because, due to the high background variability, elevated EROD activities are less easy to detect in spring/ summer than during the remaining times of the year. RECOMMENDATION AND OUTLOOK: The availability of site-specific data on the EROD baseline level, its random variation and its annual cycle is a necessary prerequisite for monitoring. If monitoring is to be carried out only for a limited time period of the year, a season with low background variability in EROD activity (autumn) should be chosen to avoid the need for a compensation of the temperature-triggered shift in sexual cycles and the resulting changes in EROD activity.

Animals↗

PubMed2005

Cytochrome P450 1A1 and 1B1 in human blood lymphocytes are not suitable as biomarkers of exposure to dioxin-like compounds: polymorphisms and interindividual variation in expression and inducibility.

Cytochrome P450 1A1 (CYP1A1) and 1B1 (CYP1B1) are phase I enzymes, the expression of which can be affected by many environmental compounds, including dioxins and dioxin-like compounds. Because CYP1A1 and CYP1B1 expression can easily be determined in peripheral blood lymphocytes, it is often suggested as biomarker of exposure to these compounds. In this study we investigated the interindividual differences in constitutive and induced CYP1A1-catalyzed ethoxyresorufin-O-deethylase (EROD) activity and CYP1A1 and CYP1B1 gene expression in human blood lymphocytes in a group of ten non-smoking females. Freshly isolated lymphocytes were cultured in medium containing the mitogen PHA and were exposed to the most potent dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or the less potent dioxin-like polychlorinated biphenyl 126 (PCB126). In addition, we determined the occurrence of the CYP1A1 MspI and CYP1B1 Leu432Val polymorphisms. All individuals showed a concentration-dependent increase of EROD activity by TCDD, which was significantly correlated with an increase in CYP1A1, but not CYP1B1 expression. The maximum induced EROD activity by TCDD was very different among the individuals, but the EC50 values were about the same. PCB126 also caused a concentration-dependent increase of EROD activity, but was a factor 100-1000 less potent than TCDD among the individuals. The allele frequencies for CYP1A1 MspI and CYP1B1 Leu432Val reflected a normal Caucasian population and in this study the polymorphisms had no apparent effect on the expression and activity of these enzymes. Our study shows a large interindividual variability in constitutive and induced EROD activity, and CYP1A1 and CYP1B1 expression in human lymphocytes. In addition, dioxin concentrations at which effects were observed in our in vitro study are about 10-fold higher than the human blood levels found in vivo, indicating that EROD activity and CYP1A1 and CYP1B1 expression in human lymphocytes might not be applicable as biomarkers of exposure to dioxin and dioxin-like compounds.

Adult↗

PubMed1999

Model systems to study the parameters determining the success of phage antibody selections on complex antigens.

Phage antibody display technology offers a powerful tool for the isolation of specific antibodies to defined target antigens. Most selection strategies described to date have relied on the availability of purified and often recombinant antigen, providing the possibility to perform selections on a well-defined antigen source. However, when the target antigen cannot be purified (e.g., an integral membrane protein), or if the antigen is unknown (e.g., when searching for novel markers on cells or tissues), panning of phage antibody libraries has to be performed on complex antigen sources such as cell surfaces or tissue sections, or even by in vivo selection methods. This provides a series of technical and experimental challenges. One focus of our research is to select antibodies directed to novel cancer-induced antigens expressed by tumours and by the tumour vasculature. To understand the parameters governing selection on complex antigen sources and to assess the efficiency of these phage library selections, we have set up two model selection systems in which both tumour cells and vascular endothelial cells serve as target "antigen". We describe a model based on phage antibodies directed to the tumour antigen epithelial glycoprotein-2, to compare phage antibody selections on a range of different antigen sources including purified and recombinant antigen, whole live cells, tissue cryosections and in vivo grown solid tumours. Secondly, we describe a model based on a phage antibody directed against the endothelial cell inducible adhesion molecule E-selectin. We compare selections on cultured cell monolayers with selections on cell suspensions immobilised on columns, to determine which selection approach is most suitable for the identification of novel tumour endothelial cell markers. Our data provide insight into the efficiency and thus potency of different selection strategies and show that there are very large differences in the recovery and enrichment of binding phage between the different methods tested. Our results further demonstrate the feasibility of phage antibody selections on whole, intact cells and show that these may sometimes compare favourably to selections on purified antigen. Selections on endothelial cells immobilised on columns compare favourably with selections on cell-monolayers; the most favourable conditions for both selection procedures are described. The implications of our data for phage antibody selections on these different complex antigen sources using either non-immune or immune phage antibody repertoires are discussed. The use of model systems such as the ones described here will help to determine optimal experimental conditions for phage library selections on complex antigens and aid in developing more powerful selection procedures for target discovery.

Animals↗

PubMed2003

Imposex in the indigenous Nassarius kraussianus (Mollusca: Neogastropoda) from South African harbours.

Nassarius kraussianus populations in the Durban and Richards Bay harbours (along the eastern seaboard of southern Africa) and in the Knysna lagoon (southern Cape coast) showed imposex development, confirming bioavailability of tributyltin (TBT) in these waterways. Incidence of imposex (which ranged from 29% to 100%) and relative penis length of females (RPL; which ranged from 1.3% to 55.2%) were markedly variable among populations from different sites in each waterway, indicative of localised effects of TBT contamination. Shell length of the snails was apparently not correlated with contamination level, as assessed from imposex measurements. While population imposex features are easy to determine and often provide a sensitive biomarker of TBT exposure, the discussion outlines reservations when extrapolating these to general ecological situations. Additionally, this study represents the first published report on the topic for sub-Saharan, African harbours, some of which seem remarkably less affected than suggested for other world regions.

Animals↗