The detection and characterisation of antibodies in human blood is a key for clinical diagnostics and risk assessmentn for autoimmunity, infectious diseases and transplantation. Antibody titre derived from immunoassays is a commonly used measure for antibody response, but this metric does not resolve readily the two fundamental properties of antibodies in solution, namely their affinity and concentration. This difficulty originates from the fact that the fundamental parameters describing the binding interaction, affinity and ligand concentration, are convoluted into the titre measurement; moreover, the difficulty of controlling the surface concentration and activity of the immobilised ligand can make it challenging to distinguish between avidity and affinity. To address these challenges, we developed microfluidic antibody affinity profiling, an assay which allows the simultaneous determination of both affinity and antibody concentration, directly in solution, without surface immobilisation or antibody purification. We demonstrate these measurements in the context of alloantibody characterisation in organ transplantation, using complex patient sera, and quantify the concentration and affinity of alloantibodies against donor Human Leukocyte Antigens (HLA), an extensively used clinical biomarker to access the risk of allograft rejection. These results outline a path towards detection and in depth profiling of antibody response in patient sera.
Four independent structures of human interleukin-4, two determined by nuclear magnetic resonance techniques and two by X-ray diffraction, have been compared in detail. The core of this four helix bundle protein is very similar in all the structures but there are some differences in loop regions that are known to be mobile in solution. Careful comparison of the experimental data sets and the methods of analysis of the different laboratories has provided clues to the sources of most of the differences, and also answered some general questions about the accuracy of protein structure determination by these two techniques.
The ability to determine the identity of specific proteins is a critical challenge in many areas of cellular and molecular biology, and in medical diagnostics. Here, we present a microfluidic protein characterisation strategy that within a few minutes generates a three-dimensional fingerprint of a protein sample indicative of its amino acid composition and size and, thereby, creates a unique signature for the protein. By acquiring such multidimensional fingerprints for a set of ten proteins and using machine learning approaches to classify the fingerprints, we demonstrate that this strategy allows proteins to be classified at a high accuracy, even though classification using a single dimension is not possible. Moreover, we show that the acquired fingerprints correlate with the amino acid content of the samples, which makes it is possible to identify proteins directly from their sequence without requiring any prior knowledge about the fingerprints. These findings suggest that such a multidimensional profiling strategy can lead to the development of novel method for protein identification in a microfluidic format.
Social surveys and interviews are staple methods within health research. One of the perceived merits of the postal questionnaire is the anonymity it affords to participants, enabling people to provide honest accounts, particularly in relation to sensitive topics. Interviewing can potentially introduce bias to participants' accounts because of a compulsion to provide socially desirable responses. Here we examine these assumptions through a comparison of questionnaire and interview accounts of the help-seeking experiences of people with symptoms of cancer.Public discourses of early diagnosis of cancer are increasingly commonplace, particularly after the ‘Be Clear on Cancer’ campaigns, which reinforced the importance of consulting quickly when experiencing cancer symptoms. This study aimed to explore the help-seeking experiences of people with symptoms of lung or colorectal cancer by inviting patients to complete a questionnaire about their symptom onset and first consultation with a health care practitioner. A sub sample of these participants were interviewed about their help-seeking experiences, with the interviews taking place within 8 weeks of the questionnaires being returned.We found that the reported length of the help-seeking interval (time from first symptom to first consultation with a health care practitioner) differed in questionnaire and interview accounts for the majority of participants. Whilst we may have expected participants to report longer intervals in the questionnaire, because of its perceived ability to reduce social desirability bias, we found that the converse was true; for most of the cases where there was a discrepancy in interval length between questionnaire and interview, longer help-seeking intervals were reported in the interview.We shall consider possible explanations for these unexpected results, suggesting that the concept of ‘public accounts’ and ‘private accounts’ provides insight into these discrepant participant responses. The formality of the questionnaire and the closed nature of questioning may encourage participants to report more socially acceptable behaviours in order to conform to public discourses around early help-seeking and early diagnosis. Whereas in interviews, participants were able to report more deviant accounts because they were within the private setting (their homes) and were able to narrate their stories and detail their reasoning.
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