28 research outputs found
Proceedings of the Thirteenth International Society of Sports Nutrition (ISSN) Conference and Expo
Meeting Abstracts: Proceedings of the Thirteenth International Society of Sports Nutrition (ISSN) Conference and Expo Clearwater Beach, FL, USA. 9-11 June 201
Restriction Endonuclease-Mediated Selective Polymerase Chain Reaction : A Novel Assay for the Detection of K-ras Mutations in Clinical Samples
The enriched polymerase chain reaction (PCR) assay has been used extensively in the detection of ras gene mutations in many types of human malignancies. Although it is very sensitive, it has a number of features that limit its use in the routine diagnostic laboratory. The aim of this study was to develop a novel enriched PCR strategy, in which the concurrent activity of the restriction enzyme BstNI and Taq polymerase allowed the amplification of mutant K-ras while inhibiting the formation of wild-type product. This restriction endonuclease-mediated selective PCR assay uses three sets of primers, together with BstNI, in the reaction mix, and the amplification products are analyzed by gel electrophoresis. The reliability of the restriction endonuclease-mediated selective PCR assay to detect activated K-ras was determined in a variety of clinical samples, including 139 fresh colorectal carcinomas and 113 paraffin-embedded blocks from 80 separate tumors of the colon and rectum, pancreas, breast, or kidney. Codon 12 mutations of the K-ras oncogene were identified in DNA from both fresh and paraffin-embedded tumors in a rapid, sensitive, and reproducible manner. Mutations were detected in 33 (24%) of the fresh colorectal cancers and 16 (20%) of the paraffin-embedded tumors. These results were 97% concordant in cases in which paraffin blocks and fresh specimens from the same tumor were available for analysis. We conclude that restriction endonuclease-mediated selective PCR is a sensitive, rapid, and robust assay for the detection of point mutations in a variety of clinical samples. Importantly, there is no need for manipulation of the sample once the PCR has been set up, and therefore, the chance of contamination is significantly reduced. In contrast to previous assays, restriction endonuclease-mediated selective PCR is not labor intensive, and its format is suitable for use in routine diagnostic laboratory
Size and Phase Control of Cubic Lyotropic Liquid Crystal Nanoparticles
The
effective use of lyotropic liquid crystalline dispersions,
such as cubosomes, as drug delivery vehicles requires that they have
tailored physical characteristics that suit specific therapeutics
and external conditions. Here, we have developed phytantriol-based
cubosomes from a dispersion of unilamellar vesicles and show that
we can control their size as well as the critical packing parameter
(CPP) of the amphiphilic bilayer through regulation of temperature
and salt concentration, respectively. Using the anionic biological
lipid 1,2-dipalmi-toylphosphatidylserine (DPPS) to prevent the cubic
phase from forming, we show that the addition of phosphate buffered
saline (PBS) results in a transition from small unilamellar vesicles
to the cubic phase due to charge-shielding of the anionic lipid. Using
dynamic light scattering, we show that the cubosomes formed following
the addition of PBS are as small as 30 nm; however, we can increase
the average size of the cubsosomes to create an almost monodisperse
dispersion of cubosomes through cooling. We propose that this phenomenon
is brought about through the phase separation of the Pluronic F-127
used to stabilize the cubosomes. To complement previous work using
the salt-induced method of cubosome production, we show, using synchrotron
small-angle X-ray scattering (SAXS), that we can control the CPP of
the amphiphile bilayer which grants us phase and lattice parameter
control of the cubosomes