Microfluidic platforms for biomarker analysis

Biomarkers have been described as characteristics, most often molecular, that provide information about biological states, whether normal, pathological, or therapeutically modified. They hold great potential to assist diagnosis and prognosis, monitor disease, and assess therapeutic effectiveness. While a few biomarkers are routinely utilised clinically, these only reflect a very small percentage of all biomarkers discovered. Numerous factors contribute to the slow uptake of these new biomarkers, with challenges faced throughout the biomarker development pipeline. Microfluidics offers two important opportunities to the field of biomarkers: firstly, it can address some of these developmental obstacles, and secondly, it can provide the precise and complex platform required to bridge the gap between biomarker research and the biomarker-based analytical device market. Indeed, adoption of microfluidics has provided a new avenue for advancement, promoting clinical utilisation of both biomarkers and their analytical platforms. This review will discuss biomarkers and outline microfluidic platforms developed for biomarker analysis.

Pan-serotype diagnostic for foot-and-mouth disease using the consensus antigen of nonstructural protein 3B

An amino acid consensus sequence for the seven serotypes of foot-and-mouth disease virus (FMDV) nonstructural protein 3B, including all three contiguous repeats, and its use in the development of a pan-serotype diagnostic test for all seven FMDV serotypes are described. The amino acid consensus sequence of the 3B protein was determined from a multiple-sequence alignment of 125 sequences of 3B. The consensus 3B (c3B) protein was expressed as a soluble recombinant fusion protein with maltose-binding protein (MBP) using a bacterial expression system and was affinity purified using amylose resin. The MBP-c3B protein was used as the antigen in the development of a competition enzyme-linked immunosorbent assay (cELISA) for detection of anti-3B antibodies in bovine sera. The comparative diagnostic sensitivity and specificity at 47% inhibition were estimated to be 87.22% and 93.15%, respectively. Reactivity of c3B with bovine sera representing the seven FMDV serotypes demonstrated the pan-serotype diagnostic capability of this bioreagent. The consensus antigen and competition ELISA are described here as candidates for a pan-serotype diagnostic test for FMDV infection.

Mathematical knowledge of some entrants to a pre-service education course

Standards for teaching emphasise the need for teachers to have deep content knowledge. To assess the mathematical knowledge of students enrolling in its B.Ed. program, the University of New England has introduced a mathematics diagnostic test. This work is the first stage of an ongoing research project into the numeracy needs of students entering the B.Ed. program. The test is a pen-andpaper test that replaces previous on-line, multiple-choice tests. This paper reports on the test results, discusses some common errors made by students and outlines the future direction of the research.

Mutation analysis of the Menkes gene

Menkes disease is a copper deficiency caused by mutations in the Menkes gene, which encodes a copper-transporting protein. This study identified the causative mutations in several Menkes patients, which provided a diagnostic test for relatives and identified critical regions of the Menkes protein. Further regions were identified through functional analysis of mutations introduced by in vitro mutagenesis.

HbA1c: chasing numbers or considering context?

Glycated haemoglobin (HbA_1c) reflects the average blood glucose level in the three months preceding the test. Changes in consecutive HbA_1c tests indicate deteriorating, or improved, glycaemic control. HbA_1c is considered to be the "gold standard" measure of blood glucose control and is often used as the basis for prescribing choices and other care decisions. A number of factors can affect the accuracy of the HbA_1c result, for example, the life span of red blood cells, assay methods and clinicians' awareness of possible interfering factors. The aim of this article is to outline how HbA_1c is used as a diagnostic test, how it is used to monitor glycaemic control and how it can guide management decisions. It is also important to emphasise the importance of considering HbA_1c in the context of the individual rather than as an isolated number.

Detection and measurement of Plasmodiophora brassicae

Clubroot, caused by Plasmodiophora brassicae, is one of the most important diseases of brassicas. Management of clubroot is difficult, and the best means of avoiding the disease include planting in areas where P. brassicae is not present and using plants and growing media free from pathogen inoculum. As P. brassicae is not culturable, its detection has traditionally relied on plant bioassays, which are time-consuming and require large amounts of glasshouse space. More recently, fluorescence microscopy, serology, and DNA-based methods have all been used to test soil, water, or plant samples for clubroot. The use of fluorescence microscopy to detect and count pathogen spores in the soil requires significant operator skill and is unlikely to serve as the basis for a routine diagnostic test. By contrast, serologic assays are inexpensive and amenable to high-throughput screening but need to be based on monoclonal antibodies because polyclonal antisera cannot be reproduced and are therefore of limited quantity. Several polymerase chain reaction (PCR)-based assays have also been developed; these are highly specific for P. brassicae and have been well-correlated with disease severity. As such, PCR-based diagnostic tests have been adopted to varying extents in Canada and Australia, but wide implementation has been restricted by sample processing costs. Efforts are underway to develop inexpensive serologic on-farm diagnostic kits and to improve quantification of pathogen inoculum levels through real-time PCR. Proper detection and quantification of P. brassicae will likely play an increasingly important role in the development of effective clubroot management strategies.

The molecular systematics of Leiopotherapon unicolor (Günther, 1859): testing for cryptic speciation in Australia’s most widespread freshwater fish

Leiopotherapon unicolor is the most widespread freshwater fish species in Australia. A comprehensive allozyme and mitochondrial DNA 16S rRNA data set was assembled from 141 specimens of L. unicolor collected Australia-wide in order to test for cryptic speciation in this far-ranging species. Surprisingly, little genetic diversity was observed within L. unicolor and provided no evidence for the existence of cryptic species within this lineage. In contrast, a small sample set of L. aheneus used as the outgroup showed two highly divergent haplotypes strongly suggestive of cryptic speciation. L. unicolor has a number of ecological and life history attributes that may explain the lack of significant genetic divergence over substantial geographical distances. The occurrence of other widespread fish and crustacean species that also display only limited genetic diversity indicate that climate conditions more favourable to dispersal across central and northern Australia than is suggested by the extent of present-day aridity have occurred in the relatively recent geological past.

Using the results of diagnostic testing to promote mathematical pedagogical knowledge

Teachers require a range of knowledge bases, including both content knowledge and pedagogical knowledge (Shulman, 1986). In recent times there have been calls from a variety of sources for teacher preparation courses to improve the mathematical knowledge of teachers, particularly primary teachers. These calls have been underlined by the recent formation of bodies such as the Institutes of Teachers in Victoria and NSW, as well as the development of teaching standards by professional bodies including the Australian Association of Mathematics Teachers. Rather than simply adopt a "back-to-basics" approach, work is required that uses the results of educational research to design courses that help pre-service students to understand how and why errors are made (by themselves and by children in their own classrooms). Diagnostic testing of preservice students is the first step in the process. However, it is not enough to simply test students and to remediate their misconceptions. Instead, the aim is to use the results of the testing to improve students' pedagogical knowledge as well as their subject content knowledge. This paper outlines one approach to the use of diagnostic testing with pre-service students and how the results can be used to assist in the development of pedagogical knowledge.

Carpal tunnel syndrome: diagnostic usefulness of sonography

PURPOSE: To prospectively evaluate accuracy of sonography for diagnosis of carpal tunnel syndrome (CTS) in patients clinically suspected of having the disease in one or both hands.
MATERIALS AND METHODS: A prospective cohort of 133 patients suspected of having CTS were referred to a teaching hospital between October 2001 and June 2002 for electrodiagnostic study. One hundred twenty patients (98 women, 22 men; mean age, 49 years; range, 19–83 years) underwent sonography within 1 week after electrodiagnostic study. Radiologist was blinded to electrodiagnostic study results. Seventy-five patients had bilateral symptoms; 23 patients, right-hand symptoms; and 22 patients, left-hand symptoms (total, 195 symptomatic hands). Cross-sectional area of median nerve was measured at three levels: immediately proximal to carpal tunnel inlet, at carpal tunnel inlet, and at carpal tunnel outlet. Flexor retinaculum was used as a landmark to margins of carpal tunnel. Optimal threshold levels (determined with classification and regression tree analysis) for areas proximal to and at tunnel inlet and at tunnel outlet were used to discriminate between patients with and patients without disease. Sensitivity, specificity, and false-positive and false-negative rates were derived on the basis of final diagnosis, which was determined with clinical history and electrodiagnostic study results as reference standard.
RESULTS: For right hands, sonography had sensitivity of 94% (66 of 70); specificity, 65% (17 of 26); false-positive rate, 12% (nine of 75); and false-negative rate, 19% (four of 21) (cutoff, 0.09 cm2 proximal to tunnel inlet and 0.12 cm2 at tunnel outlet). For left hands, sensitivity was 83% (53 of 64); specificity, 73% (24 of 33); false-positive rate, 15% (nine of 62); and false-negative rate, 31% (11 of 35) (cutoff, 0.10 cm2 proximal to tunnel inlet).
CONCLUSION: Sonography is comparable to electrodiagnostic study in diagnosis of CTS and should be considered as initial test of choice for patients suspected of having CTS.

Stat5 as a diagnostic marker for leukemia

The Jak-Stat-Socs pathway is an important component of cytokine receptor signaling. Not surprisingly, perturbation of this pathway is implicated in diseases of hematopoietic and immune origin, including leukemia, lymphoma and immune deficiencies. This review examines the role of a key component of this pathway, Stat5. This has been shown to be activated in a variety of leukemias and myeloproliferative disorders, including downstream of a range of key oncogenes where it has been shown to play an important role in mediating their effects. Therefore, Stat5 represents a useful pan-leukemia/myeloproliferative disorder diagnostic marker and key therapeutic end point, as well as representing an attractive therapeutic target for these disorders.

Process analysis of the solution strategies used for problems contained in the Minnesota Paper Board Form Board test

This thesis reviews progress toward an understanding of the processes involved in the solution of spatial problems. Previous work employing factor analysis and information processing analysis is reviewed and the emphasis on variations in speed and accuracy as the major contributers to individual differences is noted. It is argued that the strategy used by individuals is a preferable explanatory concept for identifying the cognitive substratum necessary for problem solving. Using the protocols obtained from subjects solving The Minnesota Paper Form Board (Revised), a test commonly regarded as measuring skill in spatial visualization, a number of different strategies are isolated. Assumptions as to the task variants which undergird these strategies are made and tested experimentally. The results suggest that task variants such as the size of the stimulus and the shape of the pieces interact with subject variables to produce the operating strategy. Skill in problem solving is revealed in the ability to structure the array, to hold a structured image and to reduce the number of answers requiring intensive processing. The interaction between task and subject variables results in appropriate or inappropriate strategies which in turn affect speed and accuracy. Results suggest that strategy formation and usage are the keys to explaining individual differences and an heuristic model is presented to explain the performance of individual subjects on the problems involved in the Minnesota Paper Form Board. The model can be used to predict performance on other tests; and as an aid to teaching subjects experiencing difficulties. The model presented incorporates strategy variation and is consequently mores complex than previously suggested models. It is argued that such complexity is necessary to explain the nature of a subject's performance and is also necessary to perform diagnostic evaluation. Certain structural -features of the Minnesota Paper Form Board are questioned and suggestions for improvement included. The essential explanatory function of the strategy in use makes the prevalent group administration approach suspect in the prediction of future performance in spatial or vocational activity.

Molecular systematics of the Australian freshwater shrimp, Paratya australiensis

Molecular genetic date was used to test the taxonomic hypothesis of a single species of the freshwater shrimp, Paratya australiensis. While the results identified significant cryptic diversity, clear species boundaries could not be defined. The results of this study defy a simple taxonomic conclusion but are suggestive of incipient speciation.

Molecular genetics and comparative genomics reveal RNAi is not functional in malaria parasites

Techniques for targeted genetic disruption in Plasmodium, the causative agent of malaria, are currently intractable for those genes that are essential for blood stage development. The ability to use RNA interference (RNAi) to silence gene expression
would provide a powerful means to gain valuable insight into the pathogenic blood stages but its functionality in Plasmodium remains controversial. Here we have used various RNA-based gene silencing approaches to test the utility of RNAi in malaria
parasites and have undertaken an extensive comparative genomics search using profile hidden Markov models to clarify whether RNAi machinery
exists in malaria. These investigative approaches revealed that Plasmodium lacks the enzymology required for RNAi-based ablation of gene expression
and indeed no experimental evidence for RNAi was observed. In its absence, the most likely explanations for previously reported RNAi-mediated knockdown are either the general toxicity of introduced RNA (with global down-regulation of gene expression) or a specific antisense effect mechanistically distinct from RNAi, which will need systematic
analysis if it is to be of use as a molecular genetic tool for malaria parasites.