4C). Differential loading of the antigen on the resin could result

in different OAg chain conformations and a different exposure of the epitopes and/or determine a different interaction with the corresponding antibodies. The present study describes the successful coupling of OAg from S. Typhimurium to NHS-Sepharose in order to produce an affinity matrix that is capable of purifying specific antibodies from polyclonal human serum. The method can be applied to OAg from different bacteria and it does not modify OAg chain epitopes. Columns were filled with different OAg–ADH preparations with consistent results and columns were used at least 10 times with no deleterious effect on recovery of antibodies. This process could potentially be adapted for the purification of antibodies against other bacterial polysaccharides, as well as for the large scale purification of antibodies against Salmonella OAg. GSK2126458 datasheet As such, the method will be useful for the investigation of the protective capacity of OAg antibodies with different fine specificities in the context of

immunity to NTS, both in HIV-infected African adults and in individuals immunised with OAg-based vaccines. We selleck thank Simona Rondini for the extraction of OAg from S. Typhimurium D23580. We are grateful to Robert Heyderman and the Malawi-Liverpool-Wellcome Trust Clinical Research Programme for S. Typhimurium D23580. We thank Simona Rondini, Yunshan Goh and Adam Cunningham for helpful discussions. This work was supported by a PhD studentship from the Medical Research Council, UK (to CMO’S) and a Clinical Research Fellowship from GlaxoSmithKline (to CAM). The authors Molecular motor declare that they have no conflict of interest. “
“Assessing cytokine profiles in small

tissue biopsies presents a significant technical challenge, particularly the quantification of multiple cytokines when some are present at low concentrations. Multiplex methods using Luminex technology may offer an attractive solution. However these are often developed using soluble materials such as sera or cell culture supernatants spiked with recombinant cytokines and standard protocols are not available for tissue samples. Luminex assays use multiple sets of polystyrene or paramagnetic beads or ‘microspheres’ — see Vignali (2000) and Houser (2012). Each set is fluorescently colour-coded to be identifiable on a dedicated flow cytometer or other platform and pre-coated with antibody to capture a specific cytokine or other analyte, around which a sandwich immunoassay is built. Different bead sets can be combined to enable simultaneous measurement of multiple cytokine concentrations in a single sample against standard curve preparations. These assays require substantially less sample than traditional enzyme-linked immunosorbent assays (ELISAs) – typically 25–50 μL for multiple analytes compared with 200 μL for a single analyte – yet may offer similar sensitivity to Luminex (Vignali, 2000, Biagini et al.