SummaryUsing a multistep polymerase chain reaction method, we have produced a construct in which a cDNA sequence encoding the extraceUular domain of the human 55-kD tumor necrosis factor (TNF) receptor is attached to a sequence encoding the Fc portion and hinge region of a mouse IgG1 heavy chain through an oligomer encoding a thrombin-sensitive peptide linker. This construct was placed downstream from a cytomegalovirus promoter sequence, and expressed in Chinese hamster ovary cells. A secreted protein, capable of binding TNF and inactivating it, was produced by the transfected cells. Molecular characterization revealed that this soluble version of the TNF receptor was dimeric. Moreover, the protein could be quantitatively cleaved by treatment with thrombin. However, the monovalent extracellular domain prepared in this way has a greatly reduced TNF inhibitory activity compared with that of the bivalent inhibitor. Perhaps because of its high affinity for TNF, the chimeric protein is far more effective as a TNF inhibitor than are neutralizing monodonal antibodies. This molecule may prove very useful as a reagent for the antagonism and assay of TNF and lymphotoxin from diverse species in health and disease, and as a means of deciphering the exact mechanism through which TNF interacts with the 55-kD receptor.T he recent cloning of the 55-kD (1-3) and 75-kD (3-5) TNF receptors has opened the way for further studies of TNF effects and signal transduction. Moreover, it appears that truncated receptor molecules, lacking the transmembrane or cytoplasmic domains, are capable of interacting with TNF, and therefore have been isolated from urine (6, 7) and serum (2) as TNF inhibitors. We considered that derivatives of such molecules might prove useful as antagonists of TNF action in vivo, as high affinity ligands to be applied as the basis of a more sensitive assay for TNF, and as reagents to be used in defining the molecular interaction between TNF and its receptor.Unfortunately, truncated forms of the TNF receptor are highly unstable in vivo, and therefore are poor substitutes for antibodies as a means of antagonizing TNF action in living animals. The naturally occurring TNF receptor fragments are univalent and therefore have an avidity that is effectively far lower than that of a bivalent ligand. The production of large quantities of a truncated receptor by recombinant means has, in our hands, been problematic, since the protein is produced in an inactive and insoluble form in bacteria (Peppel, K., and B. Beutler, unpublished observations). This difficulty may arise from the highly cysteine-rich structure of the receptor-binding domain (1, 2). When produced in roammalian cells through recombinant techniques, the soluble receptor fragment is active, but produced at low levels, and therefore difficult to purify (Beutler, B., and T. Brown, unpublished observations).To circumvent these problems, we have engineered a chimeric protein in which the extracellular domain of the TNF receptor, which normally engages the TNF molecu...

In the last 5 years, the availability of powerful DSP and Communications design software, and the emergence of relatively affordable devices that receive and digitize RF signals, has brought Software Defined Radio (SDR) to the desktops of many communications engineers. However, the more recent availability of very low cost SDR devices such as the RTL-SDR, costing less than $20, brings SDR to the home desktop of undergraduate and graduate students, as well as both professional engineers and the maker communities. Since the release of the various open source drivers for the RTL-SDR, many in the digital communications community have used this device to scan the RF spectrum and digitise I/Q signals that are being transmitted in the range 25MHz to 1.75GHz. This wide bandwidth enables the sampling of frequency bands containing signals such as FM radio, ISM signals, GSM, 3G and LTE mobile radio, GPS and so on. In this paper we will describe the opportunity and operation of the RTL-SDR, and the development of a hands-on, open-course for SDR. These educational materials can be integrated into core curriculum undergraduate and graduate courses, and will greatly enhance the teaching of DSP and communications theory, principles and applications. The lab and teaching materials have recently been used in Senior (4th year Undergraduate) courses and are available as open course materials for all to access, use and evolve

We report the first and complete nucleotide sequence of a human gamma 3 heavy chain constant region gene (Cy3). This gene displays the same organization than the others Cy genes and exhibits normal RNA splice and polyadenylation sites. A comparison of its primary sequence with those of Cy 1, Cy2 and Cy4 genes confirms the high degree of homology (95%) of the human family in both coding and non-coding regions, and the divergence of the hinge region. The Cy3 gene we sequenced codes for a Gm(b) y3 chain (EZZ).Comparison with other known protein sequences reveals that only two specific aminoacids are involved in the Gm(b) and Gm(g) allotypes, which suggests an important part of the spatial configuration in the allotypic specificities.

Detection of specific DNA sequences is central to modern molecular biology and also to molecular diagnostics where identification of a particular disease is based on nucleic acid identification. Many methods exist, and fluorescence spectroscopy dominates the detection technologies employed with different assay formats. This study demonstrates the use of surface-enhanced resonance Raman scattering (SERRS) to detect specific DNA sequences when coupled with modified SERRS-active probes that have been designed to modify the affinity of double- and single-stranded DNA for the surface of silver nanoparticles resulting in discernible differences in the SERRS which can be correlated to the specific DNA hybridization event. The principle of the assay lies on the lack of affinity of double-stranded DNA for silver nanoparticle surfaces; therefore, hybridization of the probe to the target results in a reduction in the SERRS signal. Use of locked nucleic acid (LNA) residues in the DNA probes resulted in greater discrimination between exact match and mismatches when used in comparison to unmodified labeled DNA probes. Polymerase chain reaction (PCR) products were detected using this methodology, and ultimately a multiplex detection of sequences relating to a hospital-acquired infection, namely, methicillin-resistant Staphylococcus aureus (MRSA), demonstrated the versatility and applicability of this approach to real-life situations.