Quenching Trypsin Is Unnecessary in Filter-Based Bottom-Up Proteomics.

Quenching digestions in proteomics prior to analysis is routine in order to eliminate residual protease activity. Residual activity leads to overdigestion, nonspecific star-activity, and back-exchange in isotopic 18O quantitation. Chemical and isobaric labeling (e.g., TMT/iTRAQ) of proteins or peptides for mass spectrometry-based proteomics is generally incompatible with ubiquitous postdigestion acidification. This necessitates buffer exchange and pH adjustments. We demonstrate that quenching is unnecessary with peptides generated from protein filter-traps, as trypsin activity and intact trypsin are negligible in the eluate from these preparations. Labeling can be directly performed on enzymatic digests from these methods, improving recovery, throughput, and ease of automation.

The financial and environmental impact of unopened medical supplies discarded in the emergency department.

BACKGROUND: Inefficient supply chain management within the US healthcare industry results in significant financial and environmental impact. Unopened medical supplies may routinely be discarded in the Emergency Department (ED), contributing as a source of unnecessary medical waste. OBJECTIVES: Quantify the financial and environmental impact of unopened medical supplies that are routinely discarded in two EDs. METHODS: The study utilized a waste audit of collection bins targeting unopened medical supplies that would have otherwise been discarded. Associated financial cost was calculated using data from the purchasing department and from an online search. End-of-life (EOL) environmental impact was calculated using the M+ Wastecare calculator. A lifecycle analysis was performed on a supplier-packaged intubation kit, which the study identified as a significant source of waste. RESULTS: High volumes of unused, unopened supplies (143.48 kg) were collected during the study period with a yearly extrapolated value of 1337 kg. Purchasing costs over 44 days at Hospital A and 37 days at Hospital B for these items amounted to $16,159.71 across both sites with a yearly extrapolated value of $150,631.73. Yearly extrapolated EOL impact yielded 5.79 tons per year of CO2eq. Components from supplier-packaged intubation kits were found to contribute to 45.2% of collected items at one site which purchased them. Lifecycle analysis of an intubation kit yields 23.6 kg of CO2eq. CONCLUSION: This study demonstrates that the disposal of unopened medical supplies contributes a significant source of financial and environmental waste in the ED setting. The results continue to support the trend of procedure kits generating significant environmental and financial waste.

Evidence-Based Practices for Interviewing Graduate Medical Education Applicants: A Systematic Review.

Background Although the selection interview is a standard admission practice for graduate medical education (GME) programs in the United States, there is a dearth of recent reviews on optimizing the trainee interview process, which has low reliability, high cost, and major risk of bias. Objective To investigate the evidence base for different selection interview practices in GME. Methods We searched 4 literature databases from inception through September 2022. Two investigators independently conducted title/abstract screening, full-text review, data extraction, and quality assessment. Disagreements were mediated by discussion. We used backward reference searching of included articles to identify additional studies. We included studies of different interview methods and excluded literature reviews, non-GME related publications, and studies comparing different applicant populations. We examined study characteristics, applicant and interviewer preferences, and interview format. We evaluated study quality using the Medical Education Research Study Quality Instrument (MERSQI). Results Of 2192 studies, 39 (2%) met our inclusion criteria. The evidence base was rated as moderately low quality using MERSQI criteria. Applicants reported preferences for several one-on-one interviews lasting 15 to 20 minutes, interviews by current trainees, and interviews including social events with only trainees. Applicants had mixed perceptions of virtual versus in-person interviews and reported that virtual interviews saved costs. The multiple mini interview (MMI) required more applicant and interviewer time than individual interviews but demonstrated construct and predictive validity and was preferred by applicants and interviewers. Conclusions Based on moderately low-quality evidence, using the MMI, training interviewers, and providing applicants with basic program information in advance should be considered for GME selection interviews.

N - acetyl-transferases required for iron uptake and aminoglycoside resistance promote virulence lipid production in M. marinum.

Phagosomal lysis is a key aspect of mycobacterial infection of host macrophages. Acetylation is a protein modification mediated enzymatically by N-acetyltransferases (NATs) that impacts bacterial pathogenesis and physiology. To identify NATs required for lytic activity, we leveraged Mycobacterium marinum, a nontubercular pathogen and an established model for M. tuberculosis. M. marinum hemolysis is a proxy for phagolytic activity. We generated M. marinum strains with deletions in conserved NAT genes and screened for hemolytic activity. Several conserved lysine acetyltransferases (KATs) contributed to hemolysis. Hemolysis is mediated by the ESX-1 secretion system and by phthiocerol dimycocerosate (PDIM), a virulence lipid. For several strains, the hemolytic activity was restored by the addition of second copy of the ESX-1 locus. Using thin-layer chromatography (TLC), we found a single NAT required for PDIM and phenolic glycolipid (PGL) production. MbtK is a conserved KAT required for mycobactin siderophore synthesis and virulence. Mycobactin J exogenously complemented PDIM/PGL production in the Δ mbtK strain. The Δ mbtK M. marinum strain was attenuated in macrophage and Galleria mellonella infection models. Constitutive expression of either eis or papA5, which encode a KAT required for aminoglycoside resistance and a PDIM/PGL biosynthetic enzyme, rescued PDIM/PGL production and virulence of the Δ mbtK strain. Eis N-terminally acetylated PapA5 in vitro , supporting a mechanism for restored lipid production. Overall, our study establishes connections between the MbtK and Eis NATs, and between iron uptake and PDIM and PGL synthesis in M. marinum . Our findings underscore the multifunctional nature of mycobacterial NATs and their connection to key virulence pathways. Significance Statement: Acetylation is a modification of protein N-termini, lysine residues, antibiotics and lipids. Many of the enzymes that promote acetylation belong to the GNAT family of proteins. M. marinum is a well-established as a model to understand how M. tuberculosis causes tuberculosis. In this study we sought to identify conserved GNAT proteins required for early stages of mycobacterial infection. Using M. marinum, we determined that several GNAT proteins are required for the lytic activity of M. marinum. We uncovered previously unknown connections between acetyl-transferases required for iron uptake and antimicrobial resistance, and the production of the unique mycobacterial lipids, PDIM and PGLOur data support that acetyl-transferases from the GNAT family are interconnected, and have activities beyond those previously reported.

Lytic transglycosylase Slt of Pseudomonas aeruginosa as a periplasmic hub protein.

Peptidoglycan is a major constituent of the bacterial cell wall. Its integrity as a polymeric edifice is critical for bacterial survival and, as such, it is a preeminent target for antibiotics. The peptidoglycan is a dynamic crosslinked polymer that undergoes constant biosynthesis and turnover. The soluble lytic transglycosylase (Slt) of Pseudomonas aeruginosa is a periplasmic enzyme involved in this dynamic turnover. Using amber-codon-suppression methodology in live bacteria, we incorporated a fluorescent chromophore into the structure of Slt. Fluorescent microscopy shows that Slt populates the length of the periplasmic space and concentrates at the sites of septation in daughter cells. This concentration persists after separation of the cells. Amber-codon-suppression methodology was also used to incorporate a photoaffinity amino acid for the capture of partner proteins. Mass-spectrometry-based proteomics identified 12 partners for Slt in vivo. These proteomics experiments were complemented with in vitro pulldown analyses. Twenty additional partners were identified. We cloned the genes and purified to homogeneity 22 identified partners. Biophysical characterization confirmed all as bona fide Slt binders. The identities of the protein partners of Slt span disparate periplasmic protein families, inclusive of several proteins known to be present in the divisome. Notable periplasmic partners (KD < 0.5 µM) include PBPs (PBP1a, KD = 0.07 µM; PBP5 = 0.4 µM); other lytic transglycosylases (SltB2, KD = 0.09 µM; RlpA, KD = 0.4 µM); a type VI secretion system effector (Tse5, KD = 0.3 µM); and a regulatory protease for alginate biosynthesis (AlgO, KD < 0.4 µM). In light of the functional breadth of its interactome, Slt is conceptualized as a hub protein within the periplasm.

The loss of the PDIM/PGL virulence lipids causes differential secretion of ESX-1 substrates in Mycobacterium marinum.

The mycobacterial cell envelope is a major virulence determinant in pathogenic mycobacteria. Specific outer lipids play roles in pathogenesis, modulating the immune system and promoting the secretion of virulence factors. ESX-1 (ESAT-6 system-1) is a conserved protein secretion system required for mycobacterial pathogenesis. Previous studies revealed that mycobacterial strains lacking the outer lipid PDIM have impaired ESX-1 function during laboratory growth and infection. The mechanisms underlying changes in ESX-1 function are unknown. We used a proteo-genetic approach to measure phthiocerol dimycocerosate (PDIM)- and phenolic glycolipid (PGL)-dependent protein secretion in M. marinum, a non-tubercular mycobacterial pathogen that causes tuberculosis-like disease in ectothermic animals. Importantly, M. marinum is a well-established model for mycobacterial pathogenesis. Our findings showed that M. marinum strains without PDIM and PGL showed specific, significant reductions in protein secretion compared to the WT and complemented strains. We recently established a hierarchy for the secretion of ESX-1 substrates in four (I-IV) groups. Loss of PDIM differentially impacted secretion of Group III and IV ESX-1 substrates, which are likely the effectors of pathogenesis. Our data suggest that the altered secretion of specific ESX-1 substrates is responsible for the observed ESX-1-related effects in PDIM-deficient strains.IMPORTANCEMycobacterium tuberculosis, the cause of human tuberculosis, killed an estimated 1.3 million people in 2022. Non-tubercular mycobacterial species cause acute and chronic human infections. Understanding how these bacteria cause disease is critical. Lipids in the cell envelope are essential for mycobacteria to interact with the host and promote disease. Strains lacking outer lipids are attenuated for infection, but the reasons are unclear. Our research aims to identify a mechanism for attenuation of mycobacterial strains without the PDIM and PGL outer lipids in M. marinum. These findings will enhance our understanding of the importance of lipids in pathogenesis and how these lipids contribute to other established virulence mechanisms.

Gender Diversity and Research Productivity of Journal Editorial and Professional Society Board Members in Medical Education.

Purpose: To describe gender diversity and research productivity among medical education boards. Methods: We examined gender, training status, and research productivity of board members of Journal Citation Reports-listed medical education journals and affiliated professional societies. We determined gender using gendered pronouns and-if unavailable-software. We evaluated differences using χ2 and t-tests. Results: Overall, half of board members but 44% of editors-in-chief and 20% of society leaders were female. Female-led journals and societies had higher female representation than their non-female-led counterparts; trainee board members were more likely to be female. Conclusions: Gender disparities exist among executives on journal and affiliated professional society boards in medical education.

N-terminal proteomics of Mycobacterium marinum using bottom-up label-free quantitative analysis in data-dependent acquisition mode on a timsTOF Pro mass spectrometer.

N-terminal acetylation in Mycobacterium tuberculosis is correlated with pathogenic activity. We used genomics and bottom-up proteomics to identify protein Emp1 as the sole acetyltransferase responsible for acetylation of EsxA, a known virulence factor. Using custom data analysis, we screened the proteome to identify 22 additional putative substrates of Emp1.

Single Surgeon Comparison of Midline Versus Overlapping Locoregional Flap Closure Following Spinal Instrumentation.

BACKGROUND: Two techniques for paraspinous muscle flap closure of spine surgeries have been described: one with tension-free mobilization of the muscle flaps approximated at the midline and one with perforators more aggressively dissected to allow for overlapping of the flaps. We seek to compare the surgical outcomes in patients who underwent either type of complex spinal closure as no investigation has yet evaluated a superior technique. METHODS: An institutional review board (IRB)-approved retrospective analysis was conducted on all patients who underwent spine surgery followed by locoregional muscle flap complex closure performed by a single plastic surgeon between January 2016 and July 2021. Patients were divided into 2 groups based on which closure method was employed. Odds ratios (ORs) and 95% confidence intervals (CIs) were computed by multivariable logistic regression with Firth's correction. RESULTS: One hundred and 10 patients with similar baseline demographics were included. There were significantly more smokers (15% vs. 0%, P = 0.02) and a significantly greater rate of postoperative radiation (40% vs. 17%, P = 0.009) in the overlapping group. After controlling for smoking and postoperative radiation, the incidence of surgical site infection, skin necrosis, dehiscence, hematoma, and seroma did not differ between the groups. The procedure length per centimeter of closure was shorter in the midline approximation group, although this data fell just short of significance (3.2 vs. 3.8 minutes/cm, P = 0.08). CONCLUSIONS: The present study demonstrates that both the overlapping and midline approximation of muscle flaps are equally safe and effective strategies for locoregional closure of spinal wounds.

The loss of the PDIM/PGL virulence lipids causes differential secretion of ESX-1 substrates in Mycobacterium marinum.

The mycobacterial cell envelope is a major virulence determinant in pathogenic mycobacteria. Specific outer lipids play roles in pathogenesis, modulating the immune system and promoting the secretion of virulence factors. ESX-1 (ESAT-6 system-1) is a conserved protein secretion system required for mycobacterial pathogenesis (1, 2). Previous studies revealed that mycobacterial strains lacking the outer lipid PDIM have impaired ESX-1 function during laboratory growth and infection (3-5). The mechanisms underlying changes in ESX-1 function are unknown. We used a proteo-genetic approach to measure PDIM and PGL-dependent protein secretion in M. marinum , a non-tubercular mycobacterial pathogen that causes tuberculosis-like disease in ectothermic animals (6, 7). Importantly, M. marinum is a well-established model for mycobacterial pathogenesis (8, 9). Our findings showed that M. marinum strains without PDIM and PGL showed specific, significant reductions in protein secretion compared to the WT and complemented strains. We recently established a hierarchy for the secretion of ESX-1 substrates in four (I-IV) groups (10). Loss of PDIM differentially impacted secretion of Groups III and IV ESX-1 substrates, which are likely the effectors of pathogenesis. Our data suggests that the altered secretion of specific ESX-1 substrates is responsible for the observed ESX-1-related effects in PDIM-deficient strains.

Algorithmic assessment of shoulder function using smartphone video capture and machine learning.

Tears within the stabilizing muscles of the shoulder, known as the rotator cuff (RC), are the most common cause of shoulder pain-often presenting in older patients and requiring expensive advanced imaging for diagnosis. Despite the high prevalence of RC tears within the elderly population, there is no previously published work examining shoulder kinematics using markerless motion capture in the context of shoulder injury. Here we show that a simple string pulling behavior task, where subjects pull a string using hand-over-hand motions, provides a reliable readout of shoulder mobility across animals and humans. We find that both mice and humans with RC tears exhibit decreased movement amplitude, prolonged movement time, and quantitative changes in waveform shape during string pulling task performance. In rodents, we further note the degradation of low dimensional, temporally coordinated movements after injury. Furthermore, a logistic regression model built on our biomarker ensemble succeeds in classifying human patients as having a RC tear with > 90% accuracy. Our results demonstrate how a combined framework bridging animal models, motion capture, convolutional neural networks, and algorithmic assessment of movement quality enables future research into the development of smartphone-based, at-home diagnostic tests for shoulder injury.

Amber-codon suppression for spatial localization and in vivo photoaffinity capture of the interactome of the Pseudomonas aeruginosa rare lipoprotein A lytic transglycosylase.

The 11 lytic transglycosylases of Pseudomonas aeruginosa have overlapping activities in the turnover of the cell-wall peptidoglycan. Rare lipoprotein A (RlpA) is distinct among the 11 by its use of only peptidoglycan lacking peptide stems. The spatial localization of RlpA and its interactome within P. aeruginosa are unknown. We employed suppression of introduced amber codons at sites in the rlpA gene for the introduction of the unnatural-amino-acids Νζ -[(2-azidoethoxy)carbonyl]-l-lysine (compound 1) and Nζ -[[[3-(3-methyl-3H-diazirin-3-yl)propyl]amino]carbonyl]-l-lysine (compound 2). In live P. aeruginosa, full-length RlpA incorporating compound 1 into its sequence was fluorescently tagged using strained-promoted alkyne-azide cycloaddition and examined by fluorescence microscopy. RlpA is present at low levels along the sidewall length of the bacterium, and at higher levels at the nascent septa of replicating bacteria. In intact P. aeruginosa, UV photolysis of full-length RlpA having compound 2 within its sequence generated a transient reactive carbene, which engaged in photoaffinity capture of neighboring proteins. Thirteen proteins were identified. Three of these proteins-PBP1a, PBP5, and MreB-are members of the bacterial divisome. The use of the complementary methodologies of non-canonical amino-acid incorporation, photoaffinity proximity analysis, and fluorescent microscopy confirm a dominant septal location for the RlpA enzyme of P. aeruginosa, as a divisome-associated activity. This accomplishment adds to the emerging recognition of the value of these methodologies for identification of the intracellular localization of bacterial proteins.

An N-acetyltransferase required for ESAT-6 N-terminal acetylation and virulence in Mycobacterium marinum.

IMPORTANCE: N-terminal acetylation is a protein modification that broadly impacts basic cellular function and disease in higher organisms. Although bacterial proteins are N-terminally acetylated, little is understood how N-terminal acetylation impacts bacterial physiology and pathogenesis. Mycobacterial pathogens cause acute and chronic disease in humans and in animals. Approximately 15% of mycobacterial proteins are N-terminally acetylated, but the responsible enzymes are largely unknown. We identified a conserved mycobacterial protein required for the N-terminal acetylation of 23 mycobacterial proteins including the EsxA virulence factor. Loss of this enzyme from M. marinum reduced macrophage killing and spread of M. marinum to new host cells. Defining the acetyltransferases responsible for the N-terminal protein acetylation of essential virulence factors could lead to new targets for therapeutics against mycobacteria.

Seizures Following Self-Medication With Colloidal Silver: A Case Report.

Silver-containing products have been used for medicinal purposes since antiquity. Throughout the ages and indeed up until the present time, silver has been employed with the hopes of treating a myriad of diseases including the common cold, skin problems, infections, and even cancer. However, silver has no known biological role in human physiology, and taking silver may lead to adverse reactions. The better-known adverse reactions of silver include argyria, or a gray-blue cutaneous discoloration, which is a known effect of silver accumulation. Additionally renal or hepatic injury may also be experienced. Reports of neurological adverse reactions are rare, however, and the extant medical literature contains very few descriptions of such cases. We report herein a case of a 70 year old man who presented with seizures as the sole manifestation of silver toxicity after self-medicating with colloidal silver.

Immune-Related Long Non-Coding RNA Signatures for Tongue Squamous Cell Carcinoma.

BACKGROUND: Tongue squamous cell carcinoma (TSCC) represents one of the major subsets of head and neck cancer, which is characterized by unfavorable prognosis, frequent lymph node metastasis, and high mortality rate. The molecular events regulating tongue tumorigenesis remain elusive. In this study, we aimed to identify and evaluate immune-related long non-coding RNAs (lncRNAs) as prognostic biomarkers in TSCC. METHODS: The lncRNA expression data for TSCC were obtained from The Cancer Genome Atlas (TCGA) and the immune-related genes were downloaded from the Immunology Database and Analysis Portal (ImmPort). Pearson correlation analysis was performed to identify immune-related lncRNAs. The TCGA TSCC patient cohort was randomly divided into training and testing cohorts. In the training cohort, univariate and multivariate Cox regression analyses were used to determining key immune-related lncRNAs, which were then validated through Cox regression analysis, principal component analysis (PCA), and receiver operating characteristic (ROC) analysis in the testing cohort. RESULTS: Six immune-related signature lncRNAs (MIR4713HG, AC104088.1, LINC00534, NAALADL2-AS2, AC083967.1, FNDC1-IT1) were found to have prognostic value in TSCC. Multivariate and univariate cox regression analyses showed that the risk score based on our six-lncRNA model, when compared to other clinicopathological factors (age, gender, stage, N, T), was an important indicator of survival rate. In addition, Kaplan-Meier survival analysis demonstrated significantly higher overall survival in the low-risk patient group than the high-risk patient group within both training and testing cohorts. The ROC analysis indicated that the AUCs for 5-year overall survival were 0.790, 0.691, and 0.721, respectively, for training, testing, and entire cohorts. Finally, PCA analysis demonstrated that the high-risk and low-risk patient groups presented significant deviation regarding their immune status. CONCLUSIONS: A prognostic model based on six immune-related signature lncRNAs was established. This six-lncRNA prognostic model has clinical significance and may be helpful in the development of personalized immunotherapy strategies.

A Tool for Low-Cost, Quantitative Assessment of Shoulder Function Using Machine Learning.

Tears within the stabilizing muscles of the shoulder, known as the rotator cuff (RC), are the most common cause of shoulder pain-often presenting in older patients and requiring expensive, advanced imaging for diagnosis1-4. Despite the high prevalence of RC tears within the elderly population, there are no accessible and low-cost methods to assess shoulder function which can eschew the barrier of an in-person physical exam or imaging study. Here we show that a simple string pulling behavior task, where subjects pull a string using hand-over-hand motions, provides a reliable readout of shoulder health across animals and humans. We find that both mice and humans with RC tears exhibit decreased movement amplitude, prolonged movement time, and quantitative changes in waveform shape during string pulling task performance. In rodents, we further note the degradation of low dimensional, temporally coordinated movements after injury. Furthermore, a predictive model built on our biomarker ensemble succeeds in classifying human patients as having a RC tear with >90% accuracy. Our results demonstrate how a combined framework bridging task kinematics, machine learning, and algorithmic assessment of movement quality enables future development of smartphone-based, at-home diagnostic tests for shoulder injury.

An N-acetyltransferase required for EsxA N-terminal protein acetylation and virulence in Mycobacterium marinum.

N-terminal protein acetylation is a ubiquitous post-translational modification that broadly impacts diverse cellular processes in higher organisms. Bacterial proteins are also N-terminally acetylated, but the mechanisms and consequences of this modification in bacteria are poorly understood. We previously quantified widespread N-terminal protein acetylation in pathogenic mycobacteria (C. R. Thompson, M. M. Champion, and P.A. Champion, J Proteome Res 17(9): 3246-3258, 2018, https:// doi: 10.1021/acs.jproteome.8b00373). The major virulence factor EsxA (ESAT-6, Early secreted antigen, 6kDa) was one of the first N-terminally acetylated proteins identified in bacteria. EsxA is conserved in mycobacterial pathogens, including Mycobacterium tuberculosis and Mycobacterium marinum, a non-tubercular mycobacterial species that causes tuberculosis-like disease in ectotherms. However, enzyme responsible for EsxA N-terminal acetylation has been elusive. Here, we used genetics, molecular biology, and mass-spectroscopy based proteomics to demonstrate that MMAR_1839 (renamed Emp1, ESX-1 modifying protein, 1) is the putative N-acetyl transferase (NAT) solely responsible for EsxA acetylation in Mycobacterium marinum. We demonstrated that ERD_3144, the orthologous gene in M. tuberculosis Erdman, is functionally equivalent to Emp1. We identified at least 22 additional proteins that require Emp1 for acetylation, demonstrating that this putative NAT is not dedicated to EsxA. Finally, we showed that loss of emp1 resulted in a significant reduction in the ability of M. marinum to cause macrophage cytolysis. Collectively, this study identified a NAT required for N-terminal acetylation in Mycobacterium and provided insight into the requirement of N-terminal acetylation of EsxA and other proteins in mycobacterial virulence in the macrophage.

Refining Treatment Strategies in Patients With Fingertip Wounds and End-Stage Renal Disease.

BACKGROUND: Individuals with end-stage renal disease (ESRD) and fingertip wounds are at high risk of poor wound healing, ultimately requiring amputations. Optimal performance of upper extremity amputation (UEA) in patients with ESRD is important to decrease complications and minimize total operative procedures needed. This study evaluated outcomes of UEA in patients with ESRD and described risk factors predisposing patients to complications. METHODS: A retrospective analysis of patients receiving nontraumatic UEA for fingertip wounds was conducted, stratified by patients with and without ESRD. Demographics, comorbidities, complications, and hospital course were analyzed between groups for differences. Subanalysis of patients with ESRD was conducted to characterize operative course and predictors of complications. RESULTS: A total of 132 patients were included, 106 controls and 26 with ESRD. Compared with controls, patients with ESRD required more amputations (P < .001) and total operations (P < .001) to achieve wound healing. Patients with ESRD experienced higher rates of postoperative complications (P < .001). Predictors for complications in patients with ESRD were comorbid diabetes (odds ratio [OR]: 45; 95% confidence interval [CI], 1.7-1226.9), vascular disease (OR: 30; 95% CI, 2-441.8), arterial calcification (OR: 18; 95% CI, 1.56-207.5), and presence of a hemodialysis shunt in the affected arm (OR: 18; 95% CI, 1.56-207.5). Within patients with ESRD, initial amputation at, or proximal to, the metacarpophalangeal joint (MCPJ) led to fewer amputations (1.2 vs 2.19, P = .04) and fewer total operative procedures (4.1 vs 6.6, P = .03), compared with initial amputation distal to the MCPJ. CONCLUSION: In nontraumatic fingertip wounds, patients with ESRD had worse operative outcomes than patients without ESRD. More aggressive management of fingertip wounds using earlier and more proximal initial amputations may expedite wound healing in certain high-risk patients with ESRD.

Miniprep assisted proteomics (MAP) for rapid proteomics sample preparation.

Complete enzymatic digestion of proteins for bottom-up proteomics is substantially improved by use of detergents for denaturation and solubilization. Detergents however, are incompatible with many proteases and highly detrimental to LC-MS/MS. Recently; filter-based methods have seen wide use due to their capacity to remove detergents and harmful reagents prior to digestion and mass spectrometric analysis. We hypothesized that non-specific protein binding to negatively charged silica-based filters would be enhanced by addition of lyotropic salts, similar to DNA purification. We sought to exploit these interactions and investigate if low-cost DNA purification spin-filters, 'Minipreps,' efficiently and reproducibly bind proteins for digestion and LC-MS/MS analysis. We propose a new method, Miniprep Assisted Proteomics (MAP), for sample preparation. We demonstrate binding capacity, performance, recovery and identification rates for proteins and whole-cell lysates using MAP. MAP recovered equivalent or greater protein yields from 0.5-50 µg analyses benchmarked against commercial trapping preparations. Nano UHPLC-MS/MS proteome profiling of lysates of Escherichia coli had 99.3% overlap vs. existing approaches and reproducibility of replicate minipreps was 98.8% at the 1% FDR protein level. Label Free Quantitative proteomics was performed and 91.2% of quantified proteins had a %CV <20% (2044/2241). Miniprep Assisted Proteomics can be performed in minutes, shows low variability, high recovery and proteome depth. This suggests a significant role for adventitious binding in developing new proteomics sample preparation techniques. MAP represents an efficient, ultra-low-cost alternative for sample preparation in a commercially obtainable device that costs ∼$0.50 (USD) per miniprep.