We analysed data from 80 patients who tested positive for SARS‐CoV‐2 RNA who had previously been HLA typed to support transplantation. Data were combined from two adjacent centres in Manchester and Leeds to achieve a sufficient number for early analysis. HLA frequencies observed were compared against two control populations: first, against published frequencies in a UK deceased donor population ( n = 10,000) representing the target population of the virus, and second, using a cohort of individuals from the combined transplant waiting lists of both centres ( n = 308), representing a comparator group of unaffected individuals of the same demographic. We report a significant HLA association with HLA‐ DQB1*06 (53% vs. 36%; p < .012; OR 1.96; 95% CI 1.94–3.22) and infection. A bias towards an increased representation of HLA‐A*26, HLA‐DRB1*15, HLA‐DRB1*10 and DRB1*11 was also noted but these were either only significant using the UK donor controls, or did not remain significant after correction for multiple tests. Likewise, HLA‐A*02, HLA‐B*44 and HLA‐C*05 may exert a protective effect, but these associations did not remain significant after correction for multiple tests. This is relevant information for the clinical management of patients in the setting of the current SARS‐CoV‐2 pandemic and potentially in risk‐assessing staff interactions with infected patients.
Immunodepletion of clinical fluids to overcome the dominance by a few very abundant proteins has been explored but studies are few, commonly examining only limited aspects with one analytical platform. We have systematically compared immunodepletion of 6, 14, or 20 proteins using serum from renal transplant patients, analysing reproducibility, depth of coverage, efficiency, and specificity using 2-D DIGE (‘top-down’) and LC-MS/MS (‘bottom-up’). A progressive increase in protein number (≥2 unique peptides) was found from 159 in unfractionated serum to 301 following 20 protein depletion using a relatively high-throughput 1-D-LC-MS/MS approach, including known biomarkers and moderate–lower abundance proteins such as NGAL and cytokine/growth factor receptors. On the contrary, readout by 2-D DIGE demonstrated good reproducibility of immunodepletion, but additional proteins seen tended to be isoforms of existing proteins. Depletion of 14 or 20 proteins followed by LC-MS/MS showed excellent reproducibility of proteins detected and a significant overlap between columns. Using label-free analysis, greater run-to-run variability was seen with the Prot20 column compared with the MARS14 column (median %CVs of 30.9 versus 18.2%, respectively) and a corresponding wider precision profile for the Prot20. These results illustrate the potential of immunodepletion followed by 1-D nano-LC-LTQ Orbitrap Velos analysis in a moderate through-put biomarker discovery process.
Screening for and treating asymptomatic bacteriuria are common in KTRs despite uncertainties around the benefits and harms. In an era of antimicrobial resistance, further studies are needed to address the diagnosis and management of asymptomatic bacteriuria in these patients.
Background: Neutrophil gelatinase-associated lipocalin (NGAL) is a promising biomarker for acute kidney injury that is beginning to be used in clinical practice in addition to research studies. The current study describes an independent validation and comparison of five commercially available NGAL assays, focusing on urine samples. This is an essential step in the translation of this marker to clinical use in terms of allowing valid inter-study comparison and generation of robust results. Methods: Two CE (Conformité Europé enne)-marked assays, the NGAL Test (BioPorto) on Siemens ADVIA w 1800 and the ARCHITECT Urine NGAL assay on i2000SR (Abbott Laboratories), and three research-use-only (RUO) ELISAs (R&D Systems, Hycult and BioPorto) were evaluated. Imprecision, parallelism, recovery, selectivity, limit of quantitation (LOQ), vulnerability to interference and hook effect were assessed and inter-assay agreement was determined using 68 urine samples from patients with various renal diseases and healthy controls. Results: The Abbott and R&D Systems assays demonstrated satisfactory performance for all parameters tested. However for the other three assays evaluated, problems were identified with LOQ (BioPorto/ADVIA w ), parallelism (BioPorto ELISA) or several parameters (Hycult). Between-method agreement varied with the Hycult assay in particular being markedly different and highlighting issues with standardization and form of NGAL measured. Conclusions: Variability exists between the five NGAL assays in terms of their performance and this should be taken into account when interpreting results from the various clinical or research studies measuring urinary NGAL.
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