A label-free mass spectrometry method for the quantification of protein isotypes

Successful quantitative mass spectrometry (MS) requires strategies to link the mass spectrometer response to the analyte abundance, with the response being dependent on more factors than just analyte abundance. Label-dependent strategies rely on the incorporation of an isotopically labeled internal standard into the sample. Current label-free strategies (performed without internal standards) are useful for analyzing samples that are unsuitable for isotopic labeling but are less accurate. Here we describe a label-free technique applicable to analysis of products from related genes (isotypes). This approach enables the invariant tryptic peptide sequences within the family to serve as “built-in” internal standards and the isotype-specific peptide sequences to report the amount of the various isotypes. A process of elimination segregates reliably trypsin-released standard and reporter peptides from unreliably released peptides. The specific MS response factors for these reporter and standard peptides can be determined using synthetic peptides. Analysis of HeLa tubulin digests revealed peptides from βI-, βII-, βIII-, βIVb-, and βV-tubulin, eight of which were suitable; along with five standard peptides for quantification of the β-tubulin isotypes. To show the utility of this method, we determined that βI-tubulin represented 77% and βIIItubulin represented 3.2% of the total HeLa β-tubulin

Tension Pneumocephalus Related to Spontaneous Skull Base Dehiscence in a Patient on BiPAP

Spontaneous pneumocephalus is an uncommon phenomenon that may develop in patients with occult skull base defects. There have been reports of pneumocephalus occurring spontaneously in the setting of continuous positive airway pressure (CPAP) use (1). Tension pneumocephalus represents a neurosurgical emergency where intracranial air is trapped with increasing pressures resulting in neurological deterioration (2). Previous literature has also documented the growing understanding of how obesity, elevated intracranial pressure (ICP), obstructive sleep apnea (OSA), and cortical skull thinning are associated with spontaneous tegmen dehiscence and cerebrospinal fluid (CSF) leakage (3). Other mechanisms for spontaneous CSF leak include aberrant arachnoid granulation, congenital skull base dehiscences, increased abdominal and thoracic pressure resulting in reduced cerebral venous drainage, and age-related cortical thinning. In this report, we present the case of a bilevel positive airway pressure (BiPAP) user with an undiagnosed spontaneous tegmen dehiscence who developed spontaneous tension pneumocephalus

Microscopy with ultraviolet surface excitation for rapid slide-free histology.

Histologic examination of tissues is central to the diagnosis and management of neoplasms and many other diseases, and is a foundational technique for preclinical and basic research. However, commonly used bright-field microscopy requires prior preparation of micrometre-thick tissue sections mounted on glass slides, a process that can require hours or days, that contributes to cost, and that delays access to critical information. Here, we introduce a simple, non-destructive slide-free technique that within minutes provides high-resolution diagnostic histological images resembling those obtained from conventional haematoxylin-and-eosin-histology. The approach, which we named microscopy with ultraviolet surface excitation (MUSE), can also generate shape and colour-contrast information. MUSE relies on ~280-nm ultraviolet light to restrict the excitation of conventional fluorescent stains to tissue surfaces, and it has no significant effects on downstream molecular assays (including fluorescence in situ hybridization and RNA-seq). MUSE promises to improve the speed and efficiency of patient care in both state-of-the-art and low-resource settings, and to provide opportunities for rapid histology in research

Diets of and trophic relationships among dominant marine nekton within the northern California Current ecosystem

In this study we analyzed the diets of 26 nekton species collected from two years (2000 and 2002) off Oregon and northern California to describe dominant nekton trophic groups of the northern California Current (NCC) pelagic ecosystem. We also examined interannual variation in the diets of three nekton species. Cluster analysis of predator diets resulted in nekton trophic groups based on the consumption of copepods, euphausiids, brachyuran larvae, larval juvenile fishes, and adult nekton. However, many fish within trophic groups consumed prey from multiple trophic levels—euphausiids being the most widely consumed. Comparison of diets between years showed that most variation occurred with changes in the contribution of euphausiids and brachyuran larvae to nekton diets. The importance of euphausiids and other crustacean prey to nekton indicates that omnivory is an important characteristic of the NCC food web; however it may change during periods of lower or higher upwelling and ecosystem production