CTCs Can Make Better Samples Than Biopsies

Although circulating tumor cells (CTCs) and cell-free DNA (cfDNA) may be “bagged” in liquid biopsy samples, they may not stay intact through all the phases of an analytical workflow. If these biomarkers suffer degradation—during storage, transportation, nucleic acid extraction, or other sample preparation steps—they may skew assays toward erroneous or equivocal results. Plainly, this is no way to investigate a patient’s cancer, make a case for an individualized therapy, or monitor attempts at correction—to say nothing of guarding against treatment resistance or cancer recurrence.

An Integrated Platform for Isolation, Processing, and Mass Spectrometry-based Proteomic Profiling of Rare Cells in Whole Blood

Abstract Isolation and molecular characterization of rare cells (e.g. circulating tumor and stem cells) within biological fluids and tissues has significant potential in clinical diagnostics and personalized medicine. The present work describes an integrated platform of sample procurement, preparation, and analysis for deep proteomic profiling of rare cells in blood. Microfluidic magnetophoretic isolation of target […]

A rapid, standardized protein extraction method using adaptive focused acoustics for identification of mycobacteria by MALDI-ToF MS

Mycobacterial identification using MALDI-ToF MS (MALDI) has been hindered by inadequate extraction methods. Adaptive Focused Acoustics™ uses concentrated ultrasonic energy to achieve cellular disruption. Using this technology, we developed a rapid mycobacterial inactivation/protein extraction method for MALDI- based identification. Agreement for identification to the species level versus conventional identification was stratified by log confidence cut-offs of ≥2.0, ≥ 1.8, or ≥1.7. A total of 182mycobacterial isolates were tested. Complete inactivation of all species/strains was achieved after 2 min. Using a log confidence cut-off of ≥2.0, overall
agreement for the commercial method (CM) was 41.7% versus 66.7% for the novel method (NM). For the CM, agreement increased to 66.7% and 83.3% using log confidence cut-offs of ≥1.8 and ≥1.7, respectively; for the NM, agreement was 100% for both cut-offs with all isolates. With no alteration to the existing database, overall
agreement for the NM was 83.4%, largely due to low scores for clinical isolates of M. chelonae and M.mucogenicum. Addition of spectra froma single clinical strain of each species to the existing database increased overall agreement to 93.1%.

Trans-Pacific-RAD-Seq Population Genomics Confirms Introgressive Hybridization in Eastern Pacific Pocillopora Corals

Discrepancies between morphology-based taxonomy and phylogenetic systematics are common in Scleractinian corals. In Pocillopora corals, nine recently identified genetic lineages disagree fundamentally with the 17 recognized Pocillopora species, including 5 major Indo-Pacific reef-builders. Pocillopora corals hybridize in the Tropical Eastern Pacific, so it is possible that some of the disagreement between the genetics and taxonomy may be due to introgressive hybridization. Here we used 6769 genome-wide SNPs from Restriction-site Associated DNA Sequencing (RAD-Seq) to conduct phylogenomic comparisons among three common, Indo-Pacific Pocillopora species – P. damicornis, P. eydouxi and P. elegans – within and between populations in the Tropical Eastern Pacific (TEP) and the Central Pacific. Genome-wide RADSeq comparisons of Central and TEP Pocillopora confirm that the morphospecies P. damicornis, P. eydouxi and P. elegans are not monophyletic, but instead fall into three distinct genetic groups. However, hybrid samples shared fixed alleles with their respective parental species and, even without strict monophyly, P. damicornis share a common set of 33 species-specific alleles across the Pacific. RAD-Seq data confirm the pattern of one-way introgressive hybridization among TEP Pocillopora, suggesting that introgression may play a role in generating shared, polyphyletic lineages among currently recognized Pocillopora species. Levels of population differentiation within genetic lineages indicate significantly higher levels of population differentiation in the Tropical Eastern Pacific than in the Central West Pacific.

RAD-Seq derived markers flank the shell colour and banding loci of the Cepaea nemoralis supergene

Studies on the classic shell colour and banding polymorphism of the land snail Cepaea played a crucial role in establishing the importance of natural selection in maintaining morphological variation. Cepaea is also a pre-eminent model for ecological genetics because the outward colour and banding phenotype is entirely genetically determined, primarily by a ‘supergene’ of at least five loci. Unfortunately, progress in understanding the evolution and maintenance of the Cepaeapolymorphism stalled, partly because of a lack of genetic markers. With a view to re-establish Cepaea as a prominent model of molecular ecology, we made six laboratory crosses of Cepaea nemoralis, five of which segregated for shell ground colour (C) and the presence or absence of bands (B). First, scoring of colour and banding in 323 individuals found no recombination between the C and B loci of the supergene. Second, using restriction site–associated DNA sequencing (RAD-Seq) of two parents and 22 offspring, we identified 44 anonymous markers putatively linked to the colour (C) and banding (B) loci. The genotype of eleven of the most promising RAD-Seq markers was independently validated in the same 22 offspring, then up to a further 146 offspring were genotyped. The closest RAD-Seq markers scored are within ~0.6 centimorgan (cM) of the C-B supergene linkage group, with the combined loci together forming a 35.8 cM linkage map of markers that flank both sides of the Cepaea C-B supergene.

Contact-free Encapsulation of Proteins in PLGA Nanoparticles by Focused Ultrasound

 40th Annual Meeting and Exposition of the Controlled Release Society, July 21-24, 2013, Honolulu, Hawaii

Schiller, Hanefeld, Schneider, Weigandt, Lehr.

Freeze Fracture Electron Microscopic Examination of Liposomes Produced by Covaris AFA Technology

Kakumanu and Jain 2013. Drug Delivery Technology 13:46-50.

In this paper, Covaris researchers analyze the structure of liposomes produced by AFA using ffEM.