Recombinant single chain variable fragment antibodies (scFv) against Pro(144)-Leu(155) fragment of human protein C

The aim of this work was to obtain the recombinant single chain variable fragments of antibodies (scFv) against human protein C, the key component of blood anticoagulation system. For this purpose a peptide that mimics a Pro144-Leu155 sequence of protein C was synthesized and the murine immune scFv library against this peptide was constructed. The protein C specific scFv 9E were selected from the constructed library by the phage-display method. The scFv 9E dissociation constant was found to be 2∙10-9 М. It was shown that scFv 9E were suitable for protein C detection by ELISA and Western blotting. Selected scFv could be further used for protein C investigation and for the development of quantitative methods for protein C detection in human blood

VIBRATIONAL SUM FREQUENCY GENERATION STUDIES OF SELF ASSEMBLED MONOLAYERS ON GOLD FOR NEURON IMMOBILIZATION

Author Institution: Department of Chemistry, Wayne State University, MI 48202, USA; Smart Sensors and Integrated Microsystems (SSIM), Department of Electrical and Computer Engineering, Department of Biomedical Engineering, Wayne State University, MI 48202, USAThe significant challenge in the development of miniaturized biocompatible neuroprosthetic devices lies in understanding how neuronal networks grow on and interact with the implant surface.Self Assembled Monolayers (SAMs) have emerged as promising candidates for defining material features on miniature scales. To assess the suitability of SAMs for neuronal growth, we have investigated correlations of neuron adhesion with the chemical structure at the SAM surface characterized by Sum Frequency Generation (SFG) vibrational spectroscopy. SFG measurements were performed on three different types of SAMs: C{_1_1} amino terminated SAM, C{_1_1} carboxy terminated SAM, and 1:1 mixture, prepared on gold substrates with different surface roughness. The CH-stretch region is dominated by CH${_2}$ stretch modes for all three SAMs regardless of the surface roughness, indicating significant amount of ${gauche }$ defects in the alkane chains. The frequencies of the methylene symmetric and asymmetric stretch transitions are close to those observed in liquid alkane chains. This also suggests that the monolayers are in a disordered liquid-like state. The C=O stretch region for carboxy terminated SAM shows one broad peak assigned to hydrogen bonded monomer COOH, 1767 cm$^{-1}$. Spectra of mixed amino: carboxy SAMs show transitions of both the COOH group and the asymmetric stretch of the COO$^{-}$, 1654 cm$^{-1}$, indicating partial ionization and homogeneous mixing without phase segregation of amino and carboxy terminated domains. Neuronal cell growth was monitored using Fluorescent Microscopic imaging, showing that only amino-terminated SAMs support growth of neuronal networks. A practical conclusion of this study is that the neuron adhesion is not critically affected by the surface roughness or details of the molecular ordering and orientation of the terminal amino groups, but only on the chemical functionality displayed at the surface

Prevalence of Bacterial Contamination of Casting Material in a Pediatric Population

Surgical site infection is a relatively common and devastating complication following pediatric orthopedic surgery. Many infections have been determined to be the result of settled airborne particles on surgical equipment and the sterile field. Fiberglass casts are commonly used orthopedic fixation devices before and after surgery; however, fiberglass casting material is expelled during the removal process and represents an uninvestigated area for the possibility of cast saw dust as a source of airborne bacterial contamination in an operating room setting. This study evaluates the prevalence and distribution of microbiota on 90 pediatric casts by collecting and culturing fiberglass cast material from 90 pediatric casts. Bacterial identification was performed using a Bruker Biotyper Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry device. 81 out of 90 casts (90%) showed evidence of microbial contamination. Isolated species were very diverse and ranged from normal skin flora to opportunistic pathogens. The 5 most commonly isolated organisms were Acinetobacter pittii, Enterobacter cloacae, Micrococcus luteus, Staphylococcus epidermidis, and Staphylococcus hominis. Further investigation is required to determine if casting material is truly a cause of surgical site infection

Increased levels of the HER1 adaptor protein Ruk(l)/CIN85 contribute to breast cancer malignancy

The adaptor protein regulator for ubiquitous kinase/c-Cbl-interacting protein of 85kDa (Ruk/CIN85) was found to modulate HER1/EGFR signaling and processes like cell adhesion and apoptosis. Although these features imply a role in carcinogenesis, it is so far unknown how and by which molecular mechanisms Ruk/CIN85 could affect a certain tumor phenotype. By analyzing samples from breast cancer patients, we found high levels of Rukl/CIN85 especially in lymph node metastases from patients with invasive breast adenocarcinomas, suggesting that Rukl/CIN85 contributes to malignancy. Expression of Rukl/CIN85 in weakly invasive breast adenocarcinoma cells deficient of Rukl/CIN85 indeed converted them into more malignant cells. In particular, Rukl/CIN85 reduced the growth rate, decreased cell adhesion, enhanced anchorage-independent growth, increased motility in both transwell migration and wound healing assays as well as affected the response to epidermal growth factor. Thereby, Rukl/CIN85 led to a more rapid and prolonged epidermal growth factor-dependent activation of Src, Akt and ERK1/2 and treatment with the Src inhibitor PP2 and the PI3K inhibitor LY294002 abolished the Rukl/CIN85-dependent changes in cell motility. Together, this study indicates that high levels of Rukl/CIN85 contribute to the conversion of breast adenocarcinoma cells into a more malignant phenotype via modulation of the Src/Akt pathway