MALDI-TOF High Mass Calibration up to 200 kDa Using Human Recombinant 16 kDa Protein Histidine Phosphatase Aggregates

Background: Protein histidine phosphatase (PHP) is an enzyme which removes phosphate groups from histidine residues. It was described for vertebrates in the year 2002. The recombinant human 16 kDa protein forms multimeric complexes in physiological buffer and in the gas phase. High-mass calibration in matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) has remained a problem due to the lack of suitable standards. Large proteins can hardly be freed of their substructural microheterogeneity by classical purification procedures so that their use as calibrants is limited. A small adduct-forming protein of validated quality is a valuable alternative for that purpose. Methodology/Principal Findings: Three major PHP clusters of,113, 209 and.600 kDa were observed in gel filtration analysis. Re-chromatography of the monomer peak showed the same cluster distribution. The tendency to associate was detected also in MALDI-TOF MS measuring regular adducts up to 200 kDa. Conclusions/Significance: PHP forms multimers consisting of up to more than 35 protein molecules. In MALDI-TOF MS it generates adduct ions every 16 kDa. The protein can be produced with high quality so that its use as calibration compoun

Longitudinal Professional Identity Development Amongst Medical Students

Abstract Title: Longitudinal Professional Identity Development Amongst Medical Students Background: Professional development is a core competency for medical student education. A standardized model for assessment of student longitudinal professional identity development will allow medical schools to better implement interventions. Methods: To assess professional development at a large, Midwest, allopathic medical school, a survey with seven statements regarding professional development was created. The statements encompassed domains of mentorship, communication skills, professionalism, and innovation and asked students to rank each statement from 1-5 (1 - highly deficient, 5 - highly proficient). The online, anonymous survey was emailed to all students (n = 1154) over a 2 month time period. Results: 319 (27.6%) surveys were completed. Responses between year 1-2 and year 3-4 showed a unanimous increase in average proficiency across all 7 statements. Year 3-4 had a significant increase in overall proficiency (p Conclusion: Although professional identity development follows an overall upward trend, year 3 is a vulnerable period for professional identity development. While increased accessibility to advising is needed in all four years, it is even more necessary in year 3. The power of the study is limited by the number of responses

Engineered reporter phages for detection of Escherichia coli, Enterococcus, and Klebsiella in urine

The rapid detection and species-level differentiation of bacterial pathogens facilitates antibiotic stewardship and improves disease management. Here, we develop a rapid bacteriophage-based diagnostic assay to detect the most prevalent pathogens causing urinary tract infections: Escherichia coli, Enterococcus spp., and Klebsiella spp. For each uropathogen, two virulent phages were genetically engineered to express a nanoluciferase reporter gene upon host infection. Using 206 patient urine samples, reporter phage-induced bioluminescence was quantified to identify bacteriuria and the assay was benchmarked against conventional urinalysis. Overall, E. coli, Enterococcus spp., and Klebsiella spp. were each detected with high sensitivity (68%, 78%, 87%), specificity (99%, 99%, 99%), and accuracy (90%, 94%, 98%) at a resolution of ≥10$^{3}$ CFU/ml within 5 h. We further demonstrate how bioluminescence in urine can be used to predict phage antibacterial activity, demonstrating the future potential of reporter phages as companion diagnostics that guide patient-phage matching prior to therapeutic phage application