Leg ulcers in older people: a national study addressing variation in diagnosis, pain and sleep disturbance.

BACKGROUND: Leg ulcers commonly emerge as a symptom of other comorbidities, often in older people. As a consequence of the ulcer, pain and sleep disturbances might occur. Due to the complex illness, the responsibility of treatment is unclear between health caregivers. The interaction between ulcer type, sleep and pain has not previously been investigated. This study aimed to explore pain in older men and women (65 years and older) with different diagnoses of leg ulcers and to investigate the associations of sleep disturbances and pain in people with leg ulcer diagnosis. METHODS: The study used a cross-sectional design and data from the Swedish Registry of Ulcer Treatment, collected between May 2009 and December 2013. One thousand and eight hundred and twenty four people were included, and 62.9% were women. The mean age was 83.4 years (SD 8.8). For the analyses, the chi-square test, Mann-Whitney U-test, t-test, one-way ANOVA and logistic regression was performed. Pain was measured by the Numeric Rating Scale (NRS), and sleep disturbances was assessed dichotomously. RESULTS: We found the prevalence of pain intensity ≥ 5 on the NRS to be 34.8% in those reporting pain. Additionally, the pain intensity was associated with the number of ulcers (p = 0.003). Sleep disturbances were associated with pain (p < 0.001) and were found in 34.8% of the total sample. Although more women than men reported pain and scored higher on the NRS, no significant gender difference in sleep disturbances was found (p = 0.606). The mean NRS scores did not differ significantly between the ulcer types; however, arterial and venous-arterial ulcers increased the risk of sleep disturbances, as did higher pain scores. CONCLUSIONS: The majority of the participants were of advanced age (>80 years) and frequently suffered from pain and sleep disturbances. Further research is needed regarding pain, sleep and wound healing in the oldest old with leg ulcers. Ulcer pain sometimes appears to receive less attention in ulcer management, as do sleep disturbances, implying that individual needs might not be satisfactorily met. National guidelines in managing leg ulcers, which also consider consequences such as sleep disturbances, pain and discomfort, are needed.

Nine surface plasmon resonance assays for specific protein quantitation during cell culture and process development.

Quantitation of protein is essential during pharmaceutical development, and a variety of methods and technologies for determination of total and specific protein concentration are available. Here we describe the development of a streamlined assay platform for specific quantitation assays using surface plasmon resonance (SPR) technology. A total of nine different assays were developed using similar conditions, of which eight assays were for quantitation of different human blood plasma proteins (IgG, IgG1-4 subclasses, IgA, transferrin, and albumin) from a chromatography-based IgG plasma process. Lastly, an assay for monitoring the concentration of a recombinant monoclonal antibody during 13 days of CHO cell culturing was developed. Assay performances were compared with enzyme-linked immunosorbent assay (ELISA), nephelometry, ARCHITECT, and Cobas c501. SPR assays were shown to have higher sensitivity than analysis using nephelometry, ARCHITECT, and Cobas and to have significantly lower analysis and hands-on time compared with ELISA. Furthermore, the SPR assays were robust enough to be used for up to 12 days, allowing specific protein concentration measurement of a sample to be completed at line within 10 min. Using the same platform with only few varied parameters between different assays has saved time in the lab as well as for evaluation and presentation of results.

Sensitive methods for evaluation of antibodies for host cell protein analysis and screening of impurities in a vaccine process.

BACKGROUND: Host cell proteins (HCP) should be carefully monitored in vaccine production. To achieve a reliable HCP estimation, a mixture of polyclonal antibodies (pAbs) with broad affinity would be of preference. Sensitive evaluations of the pAbs are therefore of value. METHODS: Column purification of specific HCPs with affinity to the anti-HCP pAbs was compared with Western blotting of the anti-HCP pAbs binding to filter bound total lysate. The anti-HCP pAbs were used in an HCP quantification analysis using surface plasmon resonance (SPR). Host cell derived impurities from an influenza vaccine process were analyzed using 2-D DIGE analysis. RESULTS: The Western blotting showed a similar HCP binding pattern of anti-HCP pAbs from immunizations using two adjuvants: CFA/IFA and AbISCO(®). From the column purification of HCPs, total proteins detectable were similar for all anti-HCP pAbs; however the immune response pattern differed significantly for the anti-HCP pAbs from the AbISCO(®) immunization. In the SPR HCP quantification assay the standard curve ranged from 0.3 to 40 µg/ml. The advantage of SPR compared with ELISA was the decreased hands on time and that the sample number was not limiting. The 2-D DIGE showed that most of the HCPs were removed at the clarification and virus capture step. DISCUSSION: Column purification of HCPs with affinity to the anti-HCP pAbs increased the sensitivity of affinity analysis compared with Western blotting and opened the possibility of further analysis. The anti-HCP pAbs did not interact with proteins in the virus; simplifying analysis of process samples using SPR. 2-D DIGE analysis gave a direct study of the impurity profile with the advantage of independence from antibody performance.

Toward an alternative compatibilizer for PLA/cellulose composites: grafting of xyloglucan with PLA.

Poly(L-lactic acid) (PLLA) chains were grafted on xyloglucan substrates via ring-opening polymerization of the L-lactide monomer. Different parameters such as the nature of the substrate (native or modified xyloglucan) and the substrate/monomer ratios were varied in the synthesis to achieve different lengths of the grafted chains. A range of experimental techniques including infrared spectroscopy and nuclear magnetic resonance were used to characterize the final product. Thermal analysis showed that the glass transition temperature of xyloglucan was decreased from 252 °C to 216 °C following the grafting of PLLA. The grafting of less hydrophilic chains from xyloglucan also affected the interaction with water: the PLLA-grafted xyloglucan was insoluble in water and the moisture uptake could be decreased by about 30%. Xyloglucan adsorbs strongly to cellulose; therefore such a graft copolymer may improve the compatibility between cellulose fibers and PLLA. The PLLA-grafted xyloglucan may be useful as a novel compatibilizer in fiber-reinforced PLLA composites.

Comparison and evaluation of a Point-of-care device (CoaguChek XS) to Owren-type prothrombin time assay for monitoring of oral anticoagulant therapy with warfarin.

BACKGROUND: The standardized test used for evaluating the effect of warfarin is the prothrombin time (PT) which is measured and expressed in international normalized ratio (INR). Regular control of treatment intensity is required since inappropriate dosage increases the risk for complications. Portable point-of-care analytical instruments for measurement of capillary whole blood PT have been available for the last decades. The purpose of this study was to compare and evaluate INR values obtained by the point-of-care device CoaguChek XS, to Owren PT in a hospital setting. MATERIALS AND METHODS: In 397 warfarin-treated patients, capillary whole blood was analyzed with the CoaguChek XS and the results were compared to analysis of venous plasma samples with the Owren PT assay. To study reproducibility and rule out preanalytical errors, a subgroup of 152 patients had two capillary blood samples analyzed with the CoaguChek XS. RESULTS: In 397 patients, with a median age(+/-2SD) of 69.0(50-88) years, there was a positive correlation between results from the CoaguChek XS and the Owren-type PT assay (r=0.94;p<0.001) and concordance of 88.2%. The mean INR difference (S.D) was 0.02 (0.22). Comparison of the 152 double samples analyzed with the CoaguChek XS, produced a positive correlation of 0.99; p<0.001. CONCLUSIONS: The CoaguChek XS presents reproducible, highly comparable results with Owren PT at therapeutic levels of INR. The CoaguChek XS seems to produce better results than the earlier CoaguChek S, probably due to a new method of PT measurement where levels of fibrinogen and haematocrit do not affect the outcome.

Synthesis of RNA using 2'-O-DTM protection.

tert-Butyldithiomethyl (DTM), a novel hydroxyl protecting group, cleavable under reductive conditions, was developed and applied for the protection of 2'-OH during solid-phase RNA synthesis. This function is compatible with all standard protecting groups used in oligonucleotide synthesis, and allows for fast and high-yield synthesis of RNA. Oligonucleotides containing the 2'-O-DTM groups can be easily deprotected under the mildest possible aqueous and homogeneous conditions. The preserved 5'-O-DMTr function can be used for high-throughput cartridge RNA purification.

Cartridge-based high-throughput purification of oligonucleotides for reliable oligonucleotide arrays.

A novel, cartridge-based procedure for the efficient and irreversible detritylation of oligonucleotides is reported. This method, combined with a process for the elimination of depurinated fragments produces, in a highly parallel fashion, oligonucleotides with better purity than those traditionally obtained using reversed-phase high-performance liquid chromotography purification. Our combined detritylation and purification methodology compares favorably with commercial cartridge-based purification systems. The benefits of working with pure oligonucleotides, with regard to higher signal and better signal linearity, are shown in array-based hybridization experiments.

Functional inactivation of the SR family of splicing factors during a vaccinia virus infection.

SR proteins are essential splicing factors required for constitutive splicing and function as key regulators of alternative RNA splicing. We have shown that SR proteins purified from late adenovirus-infected cells (SR-Ad) are functionally inactivated as splicing enhancer or splicing repressor proteins by a virus-induced partial de-phosphorylation. Here, we show that SR proteins purified from late vaccinia-virus-infected cells (SR-VV) are also hypo-phosphorylated and functionally inactivated as splicing regulatory proteins. We further show that incubating SR-Ad proteins under conditions that restore the phospho-epitopes to the SR proteins results in the restoration of their activity as splicing enhancer and splicing repressor proteins. Interestingly, re-phosphorylation of SR-VV proteins only partially restored the splicing enhancer or splicing repressor phenotype to the SR proteins. Collectively, our results suggest that viral control of SR protein activity may be a common strategy used by DNA viruses to take control of the host cell RNA splicing machinery.