Cell-Free DNA BCTs prevent gDNA contamination that may occur due to nucleated cell disruption during sample storage and shipping. This novel blood collection tube provides a method for obtaining stable cfDNA samples for rare target detection and accurate analysis while mitigating the threat of gDNA contamination.

Using Streck's Cell-free DNA BCT tubes, it is possible to preserve the original proportion of fetal cell-free DNA for extended times as well as minimize the post-sampling maternal cell-free DNA background. Preserved in this way, fetal cell-free DNA can be amplified by WGA technology to be used in prenatal diagnostic tests.

BackgroundCell-free DNA (cfDNA) circulating in blood is currently used for noninvasive diagnostic and prognostic tests. Minimizing background DNA is vital for detection of low abundance cfDNA. We investigated whether a new blood collection device could reduce background levels of genomic DNA (gDNA) in plasma compared to K3EDTA tubes, when subjected to conditions that may occur during sample storage and shipping.MethodsBlood samples were drawn from healthy donors into K3EDTA and Cell-Free DNA™ BCT (BCT). To simulate shipping, samples were shaken or left unshaken. In a shipping study, samples were shipped or not shipped. To assess temperature variations, samples were incubated at 6°C, 22°C, and 37°C. In all cases, plasma was harvested by centrifugation and total plasma DNA (pDNA) assayed by quantitative real-time polymerase chain reaction (qPCR).ResultsShaking and shipping blood in K3EDTA tubes showed significant increases in pDNA, whereas no change was seen in BCTs. Blood in K3EDTA tubes incubated at 6°C, 22°C, and 37°C showed increases in pDNA while pDNA from BCTs remained stable.ConclusionsBCTs prevent increases in gDNA levels that can occur during sample storage and shipping. This new device permits low abundance DNA target detection and allows accurate cfDNA concentrations.

A pigment-dispersing hormone (PDH) from eyestalks of the fiddler crab Ucapugilator has been purified by gel filtration, ion-exchange chromatography, partition chromatography, and reversed-phase liquid chromatography. Based on automated gas-phase sequencing and subsequent identification ofcarboxyl-terminal amide, we have assigned the primary structure of this peptide as Asn-Ser-Glu-Leu-Ile-AsnSer-Ile-Leu-Gly-Leu-Pro-Lys-Val-Met-Asn-Asp-Ala-NH2. We have confirmed the sequence by synthesizing this peptide and demonstrating that the synthetic PDH and the native PDH display identical chromatographic behavior and biological activity. This hormone is a member of a family of invertebrate neuropeptides that includes a light-adapting/pigment-dispersing octadecapeptide hormone from the prawn Pandalus borealis. In assays for melanophore pigment dispersion in destalked fiddler crabs, Uca PDH was 21-fold more potent than Pandalus PI)H. These two hormones share a hexapeptide core sequence (residues 5-10: -Ile-Asn-Ser-Ile-Leu-Gly-) as well as the amino-and carboxyl-terminal residues but differ at positions 3, 4, 11, 13, 16, and 17. These results point to speciesrelated or group-specific structural differences among crustacean PDHs. MATERIALS AND METHODSAnimals. Fiddler crabs (U. pugilator), collected from Panacea, Florida, were maintained in the laboratory in a semiterrestrial system, with sand and recirculating seawater, at 220C under a 14-hr light/10-hr dark regime and were fed on dry catfish food. Eyestalks freshly ablated from Uca were lyophilized and stored in a desiccator at -20'C until use.Bioassay. Aliquots of initial extract and of the fractions from purifications, as well as solutions of purified and synthetic peptides, were assayed for melanophore pigment dispersion in destalked fiddler crabs. Samples to be assayed were dissolved in crustacean physiological saline (15) and injected into destalked crabs (50 gl per crab). The chromatophoral stages were scored and the activity values and relative potencies were calculated as described (11). During purification the desired fractions were each tested on 4 or 5 crabs. In tests for relative potencies of native and synthetic peptides, each dose was assayed on 15 crabs (three assays, 5 crabs per assay).Extraction and Purification. In this separation effort, 8500 eyestalks (17 g) were powdered, added to 200 ml of boiling H20, and stirred for 10 min. After the mixture was cooled and the residue had settled, the supernatant was siphoned off for further processing. The residue was reextracted five times, each with 200 ml of H20 at 70-90'C. The cooled combined extract was acidified to 5% HOAc and centrifuged at 10,000 x g for 20 min at 40C. The supernatant was mixed with pretreated Bio-Rex 70 (1 g of resin/15 ml of extract). The resin pretreatment included sequential equilibration with 0.5 M HC1, 50% HOAc, and 5% HOAc. The mixture was stirred for 6 hr at 20'C and poured into a column. After the unadsorbed material passed through the column, the resin was washed with one c...

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