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DNA size fractionation is an essential tool in molecular biology and is used to isolate targets in a mixture characterized by a broad molecular-weight distribution. Microfluidics was thought to provide the opportunity to create devices capable of enhancing and speeding up the classical fractionation processes. However, this conjecture met limited success due to the low mass or volume throughput of these technologies. We describe the μLAF (μ-laboratory for DNA fractionation) technology for DNA size selection based on the stacking of molecules on films of ∼100 μm in thickness with 105cm-2pores ∼2 μm in diameter. Size selection is achieved by controlling the regime of electrohydrodynamic migration through the temporal modulation of an electric field. This technology allows the processing of milliliter-scale samples containing a DNA mass of several hundreds of ng within ∼10 min and the selection of DNA in virtually any size window spanning 200 to 1000 bp. We demonstrate that one operation suffices to fractionate sheared genomic DNA in up to six fractions with collection efficiencies of ∼20–40% and enrichment factors of ∼1.5–3-fold. These performances compare favorably in terms of speed and versatility to those of the current standards.

Abstract: Two species of hookworms (Uncinaria lucasi and Uncinaria hamiltoni) have been formally described from pinnipeds, but dissimilar types are noted from these hosts. This report is the first description of hookworms (Uncinaria spp.) from the New Zealand sea lion, Phocarctos hookeri. The nematodes were collected from dead pups on Enderby Island (Auckland Islands, 50°30', 166°17') during January and February, 2004. Standard measurements of male and female hookworms were obtained, providing a general morphometric characterization of the hookworm species in P. hookeri. Considerable variations in the body length of adult hookworms were noted within the same host. The arrangement of some of the bursal rays differs from that described for U. lucasi and U. hamiltoni.

Diagnosis of essential thrombocythemia (ET) is controversial and remains mainly an exclusion diagnosis. Endogenous megakaryocyte colony (EMC) formation have been largely evaluated to identify specific criteria for ET, but results are impeded by the lack of medium standardization. We evaluated megakaryocyte (MK) colony formation in a serum-free collagen-based medium, without cytokine and in the presence of various concentrations of thrombopoietin (TPO). Thirty-six bone marrows from patients diagnosed with ET (n = 11), polycythemia vera (PV; n = 12), reactive thrombocytosis (RT; n = 6) and healthy donors (n = 7) were assessed. We demonstrate that 11 out 11 of the ET patients had spontaneous megakaryocyte colony-forming unit (CFU-MK) formation, in contrast to none of the RT patients and healthy donors. MK progenitors from ET patients remained responsive to TPO, because exogenous addition of TPO significantly increased cloning efficiency. Moreover, at low doses of TPO (0.5 ng/ml and 5 ng/ml), the number of positive cultures and mean number of TPO stimulated CFU-MK were significantly higher in cultures of cells from patients with ET than in patients with RT. In summary, we have described a standardized serum-free, collagen-based assay that allows differential diagnosis of ET and RT, according to endogenous CFU-MK formation and sensitivity to TPO.