Cytokeratin immunoreactivity in lobular intraepithelial neoplasia.

Eighteen commercially available antibodies reactive against different cytokeratin proteins were tested on classic examples of lobular intraepithelial neoplasia (LIN) and of ductal intraepithelial neoplasia (DIN) of the breast. About 90% of higher-grade DIN (AIDH and DCIS) show no or substantially diminished reaction with clone 34betaE12 (specified as reactive against keratins 1, 5, 10, and 14 as determined by the manufacturer), while the cells of LIN were found to express the antigen reactive with this antibody. To determine which of these four keratins are present in the cells of LIN, antibodies reactive against these individual four keratins were tested. None of the four antibodies to keratins 1, 5, 10, or 14 reacted with the cells of LIN. To investigate this further, 13 additional monoclonal antibodies to various other keratin proteins were tested on the cells of LIN. Those that successfully reacted with the cells of LIN were further tested on the cells of DIN. All of the individual antibodies reactive with the cells of LIN were also reactive with the cells of DIN to a degree, with clone RCK108 (reactive against keratin 19) coming the closest to demonstrating the reactivity seen with 34betaE12. We conclude that the reactivity seen in the cells of LIN with 34betaE12 is due to either (a) a crossreaction with keratin 19 that is slightly less prominent than the reaction of the individual clone RCK108, (b) a crossreaction with a keratin protein that was not tested (3, 11, 12), (c) a crossreaction with a protein closely resembling keratin in formalin-fixed, paraffin-embedded tissue, or (d) the detection of a mutated or truncated form of keratin 1, 5, 10, or 14 that cannot be detected by the individual monoclonal antibody.     

Focus: A Multifaceted Battle Against Cancer: Extracting Knowledge from Chemical Imaging Data Using Computational Algorithms for Digital Cancer Diagnosis

Fourier transform infrared (FTIR) spectroscopic imaging is an emerging microscopy modality for clinical histopathologic diagnoses as well as for biomedical research. Spectral data recorded in this modality are indicative of the underlying, spatially resolved biochemical composition but need computerized algorithms to digitally recognize and transform this information to a diagnostic tool to identify cancer or other physiologic conditions. Statistical pattern recognition forms the backbone of these recognition protocols and can be used for highly accurate results. Aided by biochemical correlations with normal and diseased states and the power of modern computer-aided pattern recognition, this approach is capable of combating many standing questions of traditional histology-based diagnosis models. For example, a simple diagnostic test can be developed to determine cell types in tissue. As a more advanced application, IR spectral data can be integrated with patient information to predict risk of cancer, providing a potential road to precision medicine and personalized care in cancer treatment. The IR imaging approach can be implemented to complement conventional diagnoses, as the samples remain unperturbed and are not destroyed. Despite high potential and utility of this approach, clinical implementation has not yet been achieved due to practical hurdles like speed of data acquisition and lack of optimized computational procedures for extracting clinically actionable information rapidly. The latter problem has been addressed by developing highly efficient ways to process IR imaging data but remains one that has considerable scope for progress. Here, we summarize the major issues and provide practical considerations in implementing a modified Bayesian classification protocol for digital molecular pathology. We hope to familiarize readers with analysis methods in IR imaging data and enable researchers to develop methods that can lead to the use of this promising technique for digital diagnosis of cancer.     

Perfusion culture apparatus for suspended mammalian cells

A variety of processes have been proposed for mammalian cell culture in the commercial production of useful substances (e.g., monoclonal antibodies, therapeutic and diagnostics proteins). Among them, the perfusion culture of suspended non-immobilized cells is the most advantageous. Perfusion culture can be classified by the separation process of suspended cells from the culture mixture into three types, namely filtration, gravitational settling and centrifugation. From a commercial point of view, the present situation and technical problems of suspended-cell perfusion culture will be reviewed based on the three types, The recent development of perfusion culture has been carried out mainly on the filtration separation process, but the centrifugation process seems to have a promising future because of operation stability and scale-up feasibility. The reasons will be explained in details.     

An intelligent system for estimating frog community calling activity and species richness

Over the past decade, dramatic declines in frog populations have been noticed worldwide. To examine this decline, monitoring frogs is becoming increasingly important. Compared to traditional field survey methods, recent advances in acoustic sensor technology have greatly extended spatial and temporal scales for monitoring animal populations. In this paper, we examine the problem of monitoring frog populations by analysing acoustic sensor data, where the population is reflected by community calling activity and species richness. Specifically, a novel acoustic event detection (AED) algorithm is first proposed to filter out those recordings without frog calls. Then, multi-label learning is used to classify each individual recording with six acoustic features: linear predictive coding coefficients, Mel-frequency cepstral coefficients, linear-frequency cepstral coefficients, acoustic complexity index, acoustic diversity index, and acoustic evenness index. Next, frog community calling activity and species richness are estimated by accumulating the results of AED and multi-label learning, respectively. Finally, ordinary least squares regression (OLS) is conducted to reveal the relationship between frog populations (frog calling activity and species richness) and weather variables (maximum temperature and rainfall). Experimental results demonstrate that our proposed intelligent system can significantly facilitate the effort to estimate frog community calling activity and species richness with comparable accuracies. The statistical results of OLS indicate that rainfall pattern has a lagged impact on frog community calling activity (significant in the first day after rainy day) and species richness (significant in the fourth day after rainy day). Temperature is shown to affect species richness but is less likely to change calling activity.     

Nucleotide sequence of the ethidium efflux gene from Escherichia coli

The nucleotide sequence of the gene specifying the ethidium efflux system of Escherichia coli has been determined. The translated open reading frame has identified a membrane-bound polypeptide of 110 amino acids (11,960 Da) which shares 42% identity with a staphylococcal protein specifying resistance to ethidium.     

The different effects of PEG 6000 and NaCI on leaf development are associated with differential inhibition of root water transport

The inhibitory effects of PEG on whole-plant growth can exceed the effects of other osmolytes such as NaCI, and this has been ascribed to toxic contaminants, or to reduced oxygen availability in PEG solutions. We investigated another possibility, namely that PEG has an additional inhibitory effect on root water transport which in turn affects leaf development. The effects on first-leaf growth of applications of PEG 6000 or isoosmotic NaCI to the roots were determined using hydroponically grown maize (Zea mays L.) seedlings. Leaf growth rates were inhibited within minutes of PEG application to the roots and remained inhibited for days. The inhibitory effects on growth of NaCI, and also of KCl and mannitol, were much smaller. The comparative effects of NaCI and PEG on root water transport were determined by assaying pressurized flow through excised roots. PEG induced a 7-fold greater inhibition of flow through live roots than NaCI. Killing of the roots by heat treatment, to reduce cell membrane resistances to solute penetration, nearly doubled the flow rate for roots in NaCI, but not for roots in PEG. We suggest that the greater viscosity of PEG solutions, as compared with NaCI, may be a primary factor contributing to the additional inhibition of water flow through live and killed roots. PEG did not have additional effects on leaf turgor but had a 3 times greater inhibitory effect than NaCI on the irreversible extensibility of the leaves and induced 16 times more leaf accumulation of the growth inhibitory stress hormone abscisic acid (ABA). We conclude that greater inhibition of root water transport by PEG 6000, as compared with NaCI, leads to additional reductions in extensibility, additional ABA accumulation, and a greater inhibition of leaf growth.     

Measurement of 7-dehydrocholesterol and cholesterol in hair can be used in the diagnosis of Smith-Lemli-Opitz syndrome

7-dehydrocholesterol (7-DHC) and cholesterol (CHOL) are biomarkers of Smith-Lemli-Opitz Syndrome (SLOS), a congenital autosomal recessive disorder characterized by elevated 7-DHC level in patients. Hair samples have been shown to have great diagnostic and research value, which has long been neglected in the SLOS field. In this study, we sought to investigate the feasibility of using hair for SLOS diagnosis. In the presence of antioxidants (2,6-ditert-butyl-4-methylphenol and triphenylphosphine), hair samples were completely pulverized and extracted by micro-pulverized extraction in alkaline solution or in n-hexane. After microwave-assisted derivatization with N,O-Bis(trimethylsilyl)trifluoroacetamide, the analytes were measured by GC-MS. We found that the limits of determination for 7-DHC and CHOL were 10 ng/mg and 8 ng/mg, respectively. In addition, good linearity was obtained in the range of 50–4000 ng/mg and 30–6000 ng/mg for 7-DHC and CHOL, respectively, which fully meets the requirement for SLOS diagnosis and related research. Finally, by applying the proposed method to real hair samples collected from 14 healthy infants and two suspected SLOS patients, we confirmed the feasibility of hair analysis as a diagnostic tool for SLOS. In conclusion, we present an optimized and validated analytical method for the simultaneous determination of two SLOS biomarkers using human hair.