Personalizing patient risk of a life-altering event: An application of machine learning to hemiarch surgery.

OBJECTIVE: The study objective was to assess a machine learning model's ability to predict the occurrence of life-altering events in hemiarch surgery and determine contributing patient characteristics and intraoperative factors. METHODS: In total, 602 patients who underwent hemiarch replacement at a high-volume aortic center from 2009 to 2022 were included. Patients were randomly divided into training (80%) and testing (20%) sets with various eXtreme gradient boosting candidate models constructed to predict the risk of experiencing life-altering events, including stroke, mortality, or new renal replacement therapy requirement. A total of 64 input parameters from the index hospitalization were identified, including 24 demographic characteristics as well as 8 preoperative and 32 intraoperative variables. A SHapley Additive exPlanation beeswarm plot was generated to identify and interpret the impact of individual features on the predictions of the final model. RESULTS: A life-altering event was noted in 15% (90/602) of patients who underwent hemiarch replacement, including urgent/emergency cases and dissections. The final eXtreme Gradient Boosting model demonstrated a cross-validation accuracy of 88% on the testing set and was well calibrated as evidenced by a low Brier score of 0.12. The best performing model achieved an area under the receiver operating characteristic curve of 0.76 and an area under the precision recall curve of 0.55. The SHapley Additive exPlanation beeswarm plot provided insights into key features that significantly influenced model prediction. CONCLUSIONS: Machine learning demonstrated superior accuracy in predicting hemiarch patients who would experience a life-altering event. This model may help to guide patients and clinicians in stratifying risk on an individual basis, which may in turn influence clinical decision-making.

Identification of three unexpected new psychoactive substances at an Australian drug checking service.

Drug checking is a harm reduction measure that provides people with the opportunity to confirm the identity and purity of substances before consumption. The CanTEST Health and Drug Checking Service is Australia's first fixed-site drug checking service, where clients can learn about the contents of the samples they provide while receiving tailored harm reduction and health advice. Three samples were recently presented to the service with the expectation of 4-fluoromethylphenidate (4F-MPH) 1, methoxetamine (MXE) 2 and 3-methylmethcathinone (3-MMC) 3. The identity of all three samples did not meet these expectations and remained unknown on-site, as no high confidence identifications were obtained. However, further analysis by nuclear magnetic resonance spectroscopy, high resolution gas chromatography-electron ionisation-mass spectrometry and liquid chromatography-electrospray ionisation-mass spectrometry at the nearby Australian National University allowed for the structure elucidation of the three samples as 4-fluoro-α-pyrrolidinoisohexanophenone (4F-α-PiHP) 4, 1-(4-fluorobenzyl)-4-methylpiperazine (4F-MBZP) 5 and N-propyl-1,2-diphenylethylamine (propylphenidine) 6, respectively. Given all three samples were not of the expected identity and have not yet been described as new psychoactive substances in the literature, this study presents a full characterisation of each compound. As exemplified by this rapid identification of three unexpected new psychoactive substances, drug checking can be used as an effective method to monitor the unregulated drug market.

Spinal cord protection for thoracoabdominal aortic aneurysm repair: from bench to bedside.

This keynote lecture and corresponding presentation discuss the anatomy and pathophysiology surrounding spinal cord injury in aortic surgery. This article will discuss risk factors and mechanisms for spinal cord injury, including loss of direct and collateral spinal cord perfusion and ischemia-reperfusion injury. This review will examine these elements in both the laboratory and clinical setting, in addition to other neuroprotective strategies applied in clinical practice. Addressing spinal cord injury requires an integrated and considerate approach to simultaneously optimize spinal cord blood flow, promote collateralization and improve ischemic tolerance. Given the catastrophic clinical consequences for both the patient and their caregivers, continuing to investigate and examine spinal cord injury is of the utmost importance.

Frozen Elephant Trunk for Acute Type A Dissection: Is Risk from Procedure or Patient Characteristics?

BACKGROUND: The initial goal of acute Type A aortic dissection (ATAAD) repair remains to get the patient off the table safely. More extensive repair is being pushed at the index operation with the frozen elephant trunk (FET) operation, but outcomes are suggested to be worse. However, we hypothesize that the risk associated with the FET in ATAAD is from the patient presenting factors rather than the operation itself. METHODS: A retrospective review of a single institution prospective database from 2015 to 2021 was performed. Two cohorts were created based on the indication for FET: evidence of radiographic malperfusion (n = 44) or clinical malperfusion (n = 31). Data were analyzed for preoperative characteristics, intraoperative characteristics, and postoperative outcomes. Statistical univariate analysis was performed with chi-square analysis and t-tests with significance determined at an alpha level of 0.05. RESULTS: Preoperative characteristics were similar in each group, independent of malperfusion markers. The intraoperative characteristics were similar, except the clinical malperfusion group had more packed red blood cells and cryoprecipitate given. The clinical malperfusion group had longer intensive care unit length of stay (p < 0.001), more postoperative strokes (p < 0.001), more reoperations (p <0.0001), and higher mortality rate (p = 0.0003). CONCLUSION: These data suggest that clinical malperfusion increases the risk of major complications and death. However, full arch replacement with FET in the absence of clinical malperfusion does not appear to add risk to the operation for ATAAD. Patients with increased risk of distal degeneration should be considered for more aggressive replacement to avoid subsequent arch replacement.

Aortic shape variation after frozen elephant trunk procedure predicts aortic events: Principal component analysis study.

Objective: The frozen elephant trunk procedure is a well-established technique for the repair of type A ascending aortic dissection and complex aortic arch pathology. The ultimate shape created by the repair may have consequences in long-term complications. The purpose of this study was to apply a machine learning technique to comprehensively describe 3-dimensional aortic shape variations after the frozen elephant trunk procedure and associate these variations with aortic events. Methods: Computed tomography angiography acquired before discharge of patients (n = 93) who underwent the frozen elephant trunk procedure for type A ascending aortic dissection or ascending aortic arch aneurysm was preprocessed to yield patient-specific aortic models and centerlines. Aortic centerlines were subjected to principal component analysis to describe principal components and aortic shape modulators. Patient-specific shape scores were correlated with outcomes defined by composite aortic event, including aortic rupture, aortic root dissection or pseudoaneurysm, new type B dissection, new thoracic or thoracoabdominal pathologies, residual descending aortic dissection with residual false lumen flow, or thoracic endovascular aortic repair complications. Results: The first 3 principal components accounted for 36.4%, 26.4%, and 11.6% of aortic shape variance, respectively, and cumulatively for 74.5% of the total shape variation in all patients. The first principal component described variation in arch height-to-length ratio, the second principal component described angle at the isthmus, and the third principal component described variation in anterior-to-posterior arch tilt. Twenty-one aortic events (22.6%) were encountered. The degree of aortic angle at the isthmus described by the second principal component was associated with aortic events in logistic regression (hazard ratio, 0.98; 95% confidence interval, 0.97-0.99; P = .046). Conclusions: The second principal component, describing angulation at the region of the aortic isthmus, was associated with adverse aortic events. Observed shape variation should be evaluated in the context of aortic biomechanical properties and flow hemodynamics.

Conditional Survival in Lung Transplantation: An Organ Procurement and Transplantation Network Database Analysis.

Lung transplantation survival estimates are traditionally reported as fixed 1-, 5-, and 10-year mortality rates. Alternatively, this study aims to demonstrate how conditional survival models can provide useful prognostic information tailored to the time a recipient has already survived from the date of transplantation. Recipient data was obtained from the Organ Procurement and Transplantation Network database. Data from 24,820 adult recipients over age 18 who received a lung transplant between 2002 and 2017 were included in the study. Five-year observed conditional survival estimates were calculated by recipient age, sex, race, transplant indication, transplant type ( i.e. , single or double), and renal function at the time of transplantation. Significant variability exists in conditional survival following lung transplantation. Each specific recipient characteristic significantly impacted conditional survival during at least one time point in the first 5 years. Younger age and double lung transplantation were the two most positive predictors of improved conditional survival consistently throughout the 5-year study period. Conditional survival in lung transplantation recipients changes over time and across recipient characteristics. Hazards of mortality are not fixed and need to be dynamically evaluated as a function of time. Conditional survival calculations can provide more accurate prognostic predictions than unconditional survival estimates.

Comparison Study of Iliac Branch Endoprosthesis when Used on and off Label.

BACKGROUND: The aim of this study is to compare how instructions for use (IFU) affected perioperative and intermediate term outcomes for common iliac artery aneurysms (CIAA) treated with the Gore Excluder iliac branch endoprosthesis (IBE). METHODS: A retrospective analysis was performed of all patients treated at two affiliated academic centers from September 2016 to May 2020. Outcomes were compared between IFU and nonIFU IBE cases. Criteria for nonIFU included: (1) use with a nonGore aortic endoprosthesis (n = 10), (2) isolated IBE (n = 3), and (3) requiring nondedicated covered stents for additional extension into a more suitable landing zone in the ipsilateral internal iliac artery or one of its branches (n = 11). Perioperative and intermediate term data were collected for both groups. The primary end points were free from the major adverse event (MAE) at 30 days and primary effectiveness at 1 year. RESULTS: A total of 51 CIAA (39 patients) were treated with an IBE. Overall, 15 patients were treated under IFU and 24 under nonIFU. The IFU group mean age was older (72 vs. 67 years, P = 0.03), and males (97%) were primarily treated. Comorbidities were similar except nonIFU had more patients with previous endovascular abdominal aortic aneurysm repair on presentation (0 vs. 4 cases, P = 0.04). Procedure (178 vs. 264 min, P = 0.02) and fluoroscopy (52 vs. 74 min, P = 0.04) times were longer in the nonIFU group. Technical success was 100% for both groups, and there was no difference in device related reintervention at 30 days (0 vs. 1, P = 0.44). There was no MAE in either group at 30 days. Intervention for any endoleak was similar between the groups (2 vs. 3, P = 0.94). Percent CIAA sac regression was similar between the groups (19% vs. 18%, P = 0.21). There was no difference for primary effectiveness at 1 year (93% vs. 92%, P = 0.85). There was one death per group at one year not related to an aortic or iliac cause. CONCLUSIONS: In properly selected patients with complex anatomy, IBE can be used with nondedicated aortic and internal iliac components with good early term outcomes.

Lagovirus Non-structural Protein p23: A Putative Viroporin That Interacts With Heat Shock Proteins and Uses a Disulfide Bond for Dimerization.

The exact function(s) of the lagovirus non-structural protein p23 is unknown as robust cell culture systems for the Rabbit haemorrhagic disease virus (RHDV) and other lagoviruses have not been established. Instead, a range of in vitro and in silico models have been used to study p23, revealing that p23 oligomerizes, accumulates in the cytoplasm, and possesses a conserved C-terminal region with two amphipathic helices. Furthermore, the positional homologs of p23 in other caliciviruses have been shown to possess viroporin activity. Here, we report on the mechanistic details of p23 oligomerization. Site-directed mutagenesis revealed the importance of an N-terminal cysteine for dimerization. Furthermore, we identified cellular interactors of p23 using stable isotope labeling with amino acids in cell culture (SILAC)-based proteomics; heat shock proteins Hsp70 and 110 interact with p23 in transfected cells, suggesting that they 'chaperone' p23 proteins before their integration into cellular membranes. We investigated changes to the global transcriptome and proteome that occurred in infected rabbit liver tissue and observed changes to the misfolded protein response, calcium signaling, and the regulation of the endoplasmic reticulum (ER) network. Finally, flow cytometry studies indicate slightly elevated calcium concentrations in the cytoplasm of p23-transfected cells. Taken together, accumulating evidence suggests that p23 is a viroporin that might form calcium-conducting channels in the ER membranes.

Profiling Urinary Sulfate Metabolites With Mass Spectrometry.

The study of urinary phase II sulfate metabolites is central to understanding the role and fate of endogenous and exogenous compounds in biological systems. This study describes a new workflow for the untargeted metabolic profiling of sulfated metabolites in a urine matrix. Analysis was performed using ultra-high-performance liquid chromatography-high resolution tandem mass spectrometry (UHPLC-HRMS/MS) with data dependent acquisition (DDA) coupled to an automated script-based data processing pipeline and differential metabolite level analysis. Sulfates were identified through k-means clustering analysis of sulfate ester derived MS/MS fragmentation intensities. The utility of the method was highlighted in two applications. Firstly, the urinary metabolome of a thoroughbred horse was examined before and after administration of the anabolic androgenic steroid (AAS) testosterone propionate. The analysis detected elevated levels of ten sulfated steroid metabolites, three of which were identified and confirmed by comparison with synthesised reference materials. This included 5α-androstane-3ß,17α-diol 3-sulfate, a previously unreported equine metabolite of testosterone propionate. Secondly, the hydrolytic activity of four sulfatase enzymes on pooled human urine was examined. This revealed that Pseudomonas aeruginosa arylsulfatases (PaS) enzymes possessed higher selectivity for the hydrolysis of sulfated metabolites than the commercially available Helix pomatia arylsulfatase (HpS). This novel method provides a rapid tool for the systematic, untargeted metabolic profiling of sulfated metabolites in a urinary matrix.

Survival following lung transplantation: A population-based nested case-control study.

BACKGROUND: Lung transplantation is the mainstay of treatment for patients with end-stage respiratory failure. This study sought to evaluate survival following transplantation compared to the general population and quantify standardized mortality ratios (SMRs) using a nested case-control study design. METHODS: Control subjects were nonhospitalized inhabitants of the United States identified through the National Longitudinal Mortality Study. Case subjects were adults who underwent lung transplantation between 1990 and 2007 and identified through the Organ Procurement and Transplantation Network. Propensity-matching (5:1, nearest neighbor, caliper = 0.1) was utilized to identify suitable control subjects based on age, sex, race, and location of residency. The primary study endpoint was 10-year survival. RESULTS: About 14,977 lung transplant recipients were matched to 74,885 nonhospitalized US residents. The 10-year survival rate of lung transplant recipients was 28% (95% confidence interval [CI] = 27%-29%). The population expected mortality rate was 19 deaths/100 person-years while the observed ratio was 104 deaths/100 person-years (SMR = 5.39, 95% CI = 5.35-5.43). The largest discrepancies between observed and expected mortality rates were in females (SMR = 5.97), Hispanic (SMR = 10.70), and single lung recipients (SMR = 5.92). SMRs declined over time (1990-1995 = 5.79, 1996-2000 = 5.64, and 2001-2007 = 5.10). Standardized mortality peaks in the first year after transplant and decreases steadily over time. CONCLUSIONS: Lung transplant recipients experience a fivefold higher SMR compared to the nonhospitalized population. Long-term mortality rates have experienced consistent decline over time.

Appraisal of Donation After Circulatory Death: How Far Could We Expand the Heart Donor Pool?

BACKGROUND: the incidence of organ donation after circulatory death (DCD) is increasing; however, heart use has lagged behind other solid organs. Ex vivo perfusion devices are under United States Food and Drug Administration review for use in DCD heart recovery. This study sought to measure the potential increase in the donor pool if DCD heart donation becomes widely adopted. METHODS: DCD donor data were obtained from Organ Procurement and Transplantation Network database. Selection criteria included donor age 18 to 49 years, donors meeting Maastricht III criteria, warm ischemia time ≤30 minutes, and donation between 2015 and 2020. Exclusion criteria were coronary disease, prior myocardial infarction, ejection fraction <0.50, significant valve disease, bacteremia, pulmonary capillary wedge pressure >15 mm Hg, and history of HIV/hepatitis C virus infections. RESULTS: There were 12 813 DCD donors during this period, of which 3528 met study criteria, and 70 hearts (2%) were transplanted. The use of DCD hearts would represent an additional 48 heart transplants per month, which corresponds to a 21% (3458 of 16 521) increase across the country. Median warm ischemia was 23 minutes, with no difference between hearts that were or were not transplanted (23 vs 22.5 minutes, P = .97). The frequency with which other organs were successfully transplanted was kidney, 92%; liver, 44%; lung, 7%; intestine, 0%; and pancreas, 2%. CONCLUSIONS: Wide adoption of DCD heart transplantation could yield a substantial increase in the donor pool size, with approximately 580 additional organs being available each year across the United States. This would represent the largest increase in the donor pool in the modern era of heart transplantation.

Measuring the effect of the COVID-19 pandemic on solid organ transplantation.

BACKGROUND: The COVID-19 pandemic has uniquely affected the United States. We hypothesize that transplantation would be uniquely affected. METHODS: In this population-based cohort study, adult transplantation data were examined as time series data. Autoregressive-integrated-moving-average models of transplantation rates were developed using data from 1990 to 2019 to forecast the 2020 expected rates in a theoretical scenario if the pandemic did not occur to generate observed-to-expected (O/E) ratios. RESULTS: 32,594 transplants were expected in 2020, and only 30,566 occurred (O/E 0.94, CI 0.88-0.99). 58,152 waitlist registrations were expected and 50,241 occurred (O/E 0.86, CI 0.80-0.94). O/E ratios of transplants were kidney 0.92 (0.86-0.98), liver 0.96 (0.89-1.04), heart 1.05 (0.91-1.23), and lung 0.92 (0.82-1.04). O/E ratios of registrations were kidney 0.84 (0.77-0.93), liver 0.95 (0.86-1.06), heart 0.99 (0.85-1.18), and lung 0.80 (0.70-0.94). CONCLUSIONS: The COVID-19 pandemic was associated with a significant deficit in transplantation. The impact was strongest in kidney transplantation and waitlist registration.

Decision Support Needs for Patients with Severe Symptomatic Aortic Stenosis.

Social workers in healthcare settings often support patient decision-making processes for complex medical decisions. The objective of this study was to examine decision support needs for patients considering aortic valve replacement (AVR) for aortic stenosis. Seventeen qualitative interviews were conducted to explore treatment decision experiences of patients who accepted AVR. Analysis was conducted using a mixed inductive-deductive approach. Fear was a prevalent response for most participants in the face of AVR. Two general paths of decision making emerged: an "active" information seeking approach, or a "passive" simplicity seeking approach. Patients with unique clinical presentations felt alienated by the decision-making process. Acknowledging fear while understanding different decision-making styles provide opportunities for social workers and other members of multidisciplinary teams to support complex patient decisions.

An Experimentally Verified LC-MS Protocol toward an Economical, Reliable, and Quantitative Isotopic Analysis in Nitrogen Reduction Reactions.

To substitute the energy-intensive Haber-Bosch process for the synthesis of ammonia, some labile techniques, such as photocatalysis, electrocatalysis, photoelectrocatalysis, and photothermocatalysis, have emerged and attracted intense research interest. However, the contamination of the reaction system is one of the major concerns on how to reliably and accurately evaluate the performance of these catalysts, which is why various control studies are involved. Isotopic labeling studies are one of the most reliable control strategies in nitrogen fixation experiments, to ensure that N2 is exclusively the source of the generated ammonia. As a convenient, sensitive and accurate technique distinguished with a quantitative atomic mass resolution, liquid chromatography-mass spectrometry (LC-MS) has been extensively employed for the detection of ammonia in aqueous electrolyte systems. However, the previous work protocols for 15 N2 isotopic analysis using LC-MS either involved hazardous procedures which could potentially damage the instrument, or lacked in their experimental verification using real samples. Herein, a safe, reproducible and economical protocol for the detection of ammonia using LC-MS is presented, exhibiting an exponentially steep progressive detectivity of 15 N abundance, well verified with a series of experimental results for nitrogen reduction reactions. This is expected to provide a prudent cost-effective and sustainable gateway into isotopic analysis.

Application of the Double-Mutant Cycle Strategy to Protein Aggregation Reveals Transient Interactions in Amyloid-ß Oligomers.

Transient oligomeric intermediates in the peptide or protein aggregation pathway are suspected to be the key toxic species in many amyloid diseases, but deciphering their molecular nature has remained a challenge. Here we show that the strategy of "double-mutant cycles", used effectively in probing protein-folding intermediates, can reveal transient interactions during protein aggregation. It does so by comparing the changes in thermodynamic parameters between the wild type, and single and double mutants. We demonstrate the strategy by probing the possible transient salt bridge partner of lysine 28 (K28) in the oligomeric states of amyloid ß-40 (Aß40), the putative toxic species in Alzheimer's disease. In mature fibrils, the binding partner is aspartate 23. This interaction differentiates Aß40 from the more toxic Aß42, where K28's binding partner is the C-terminal carboxylate. We selectively acetylated K28 and amidated the C-terminus of Aß40, creating four distinct variants. Spectroscopic measurements of the kinetics and thermodynamics of aggregation show that K28 and the C-terminus interact transiently in the early phases of the Aß40 aggregation pathway. Hydrogen-deuterium exchange mass spectrometry (using a simple analysis method that we introduce here that takes into account the isotopic mass distribution) supports this interpretation. It is also supported by cellular toxicity measurements, suggesting possible similarities in the mechanisms of toxicity of Aß40 oligomers (which are more toxic than Aß40 fibrils) and Aß42. Our results show that double-mutant cycles can be a powerful tool for probing transient interactions during protein aggregation.

Poor mobilization of autologous CD34+ peripheral blood stem cells in haematology patients undergoing autologous stem cell transplantation is associated with the presence of variants in genes implicated in clonal haematopoiesis of indeterminant potential.

Clonal haematopoiesis of indeterminant potential (CHIP) increases in frequency with age. The effect of CHIP on the mobilization of autologous CD34+ peripheral blood stem cells (PBSC) has not been reported. This study uses a DNA-based targeted candidate gene approach to identify the presence of somatic mutations in ASXL1, DNMT3A, JAK2, SF3B1, TET2 and TP53 in CD34+ haematopoietic progenitor cell-apheresis products of 96 patients who undergo PBSC mobilization for autologous stem cell transplantation (ASCT). Variants were identified in a significantly greater proportion of patients who experience poor CD34+ PBSC mobilization. A DNA-based targeted candidate gene array is able to predict poor CD34+ PBSC mobilization and may be deployed pre-emptively to minimize mobilization and graft failures.

Paraneoplastic bullous pemphigoid - A sign of clear cell renal carcinoma.

Paraneoplastic bullous pemphigoid is a rare paraneoplastic syndrome. Rash is pruritic, with erythematous eruption of large subepidermal bullae over skin and often mucosal surfaces. We present an 84y woman with a three week history of erythematous rash, and 48hrs of bullae. A left clear cell renal cell carcinoma was identified on CT imaging during the presentation. Subsequent removal of the tumour resulted in resolution of bullous pemphigoid symptoms. This first-of-kind case and successful result strengthens the association between renal cell carcinoma and paraneoplastic bullous pemphigoid, arguing for a high degree of clinical suspicion in unexplained presentations of bullous pemphigoid.

Protein synthesis increases with photosynthesis via the stimulation of translation initiation.

Leaf protein synthesis is an essential process at the heart of plant nitrogen (N) homeostasis and turnover that preferentially takes place in the light, that is, when N and CO2 fixation occur. The carbon allocation to protein synthesis in illuminated leaves generally accounts for ca. 1 % of net photosynthesis. It is likely that protein synthesis activity varies with photosynthetic conditions (CO2/O2 atmosphere composition) since changes in photorespiration and carbon provision should in principle impact on amino acid supply as well as metabolic regulation via leaf sugar content. However, possible changes in protein synthesis and translation activity when gaseous conditions vary are virtually unknown. Here, we address this question using metabolomics, isotopic techniques, phosphoproteomics and polysome quantitation, under different photosynthetic conditions that were varied with atmospheric CO2 and O2 mole fraction, using illuminated Arabidopsis rosettes under controlled gas exchange conditions. We show that carbon allocation to proteins is within 1-2.5 % of net photosynthesis, increases with photosynthesis rate and is unrelated to total amino acid content. In addition, photosynthesis correlates to polysome abundance and phosphorylation of ribosomal proteins and translation initiation factors. Our results demonstrate that translation activity follows photosynthetic activity, showing the considerable impact of metabolism (carboxylation-oxygenation balance) on protein synthesis.

Improving the Identification and Coverage of Plant Transmembrane Proteins in Medicago Using Bottom-Up Proteomics.

Plant transmembrane proteins (TMPs) are essential for normal cellular homeostasis, nutrient exchange, and responses to environmental cues. Commonly used bottom-up proteomic approaches fail to identify a broad coverage of peptide fragments derived from TMPs. Here, we used mass spectrometry (MS) to compare the effectiveness of two solubilization and protein cleavage methods to identify shoot-derived TMPs from the legume Medicago. We compared a urea solubilization, trypsin Lys-C (UR-TLC) cleavage method to a formic acid solubilization, cyanogen bromide and trypsin Lys-C (FA-CTLC) cleavage method. We assessed the effectiveness of these methods by (i) comparing total protein identifications, (ii) determining how many TMPs were identified, and (iii) defining how many peptides incorporate all, or part, of transmembrane domains (TMD) sequences. The results show that the FA-CTLC method identified nine-fold more TMDs, and enriched more hydrophobic TMPs than the UR-TLC method. FA-CTLC identified more TMPs, particularly transporters, whereas UR-TLC preferentially identified TMPs with one TMD, particularly signaling proteins. The results suggest that combining plant membrane purification techniques with both the FA-CTLC and UR-TLC methods will achieve a more complete identification and coverage of TMPs.