Surface-enhanced Laser Desorption Research Articles

Dynamic Monitoring of Serum Protein in Acute Respiratory Distress Syndrome Based on Artificial Neural Network.

Acute respiratory distress syndrome (ARDS) is one of the more serious diseases in human lung disease. Reducing its incidence rate is an important task in current clinical research. Dynamic monitoring of serum protein in patients will help to achieve the early diagnosis and treatment of ARDS. In this study, a protein monitoring model based on artificial neural network is proposed. First, surface enhanced laser desorption ionization time-of-flight mass spectrometry is used for protein detection, and then BP neural network is used for protein classification and content analysis. In the experimental analysis, serum samples from patients with acute respiratory distress syndrome in our hospital from November 2020 to August 2021 were selected for experimental testing. The experimental results show that the serum protein monitoring model based on BP neural network has low error and high convergence ability and can monitor individual protein in protein monitoring, and the area under the ROC curve in diagnostic performance reaches 0.854. The above results show that the artificial neural network has a good effect on the dynamic monitoring of serum protein in acute respiratory distress syndrome, and the diagnostic performance evaluation can reach 0.854, which has the ability to significantly improve the clinical diagnosis and treatment of acute respiratory distress syndrome.

Comparative Studies on a Branded Recombinant Factor VIIa and Its Generic Version to Demonstrate Bioequivalence

Background: Recombinant factor VIIa (NovoSeven® Copenhagen, Denmark) is used in the management of bleeding in patients with hemophilia. A generic biosimilar version of NovoSeven® is also developed for the same indication (AryoSeven™, AryoGen, Tehran-Iran). These generic versions of rFVIIa are claimed to be biosimilar to the Novoseven. The purpose of this study is to compare several batches of generic versions of rFVIIa namely, Aryoseven with multiple commercially available batches of branded Novoseven to determine bioavailability.Methods: Four commercially available random lots of Novoseven were obtained through US and European sources and compared to 5 clinical batches of Aryoseven obtained from Aryogen (Tehran, Iran). These drugs were diluted with saline to make a stock solution of 1mg/ml. Further dilutions were made to obtain working concentrations of 100, 10, 1 and 0.1 µg/ml. Molecular profile using surface enhanced laser desorption ionization (SELDI), gel electrophoretic profile (GEP), immunoblotting studies, and procoagulant activity as measured by clot based method were carried out. Thrombin generation studies were carried out after supplementing with NHP at 0-1.0 ug/ml. The pharmacokinetic profile of one representative batch of Novoseven and Aryoseven was studied in primates after a 50 ug/ml bolus IV in terms of factor VIIa levels.Results: The SELDI mass spectrometric profile of all rFVIIa preparations were comparable. The GEP and immunoblotting studies of the two groups of agents showed a comparable profile with distinct peaks at 50 KDa and 29 KDa. In the clot based functional assays performed in the citrated whole blood and retrieved plasma the two batches of FVIIa produced comparable procoagulant effects in the PT and aPTT assays. In the 0.25 - 1.0 µg/ml range in comparison to the control with saline supplementation, the aPTT did not exhibit any significant shortening whereas there was a sharpening of PT values in the whole blood which was identical in both the AryoSeven™ and NovoSeven® preparations. In the retrieved plasma both products produced comparable effect on the PT and aPTT assays. In platelet rich plasma (PRP) and platelet poor plasma (PPP) AryoSeven and NovoSeven at graded concentration of 0.125 to 1 µg/ml did not produce any measurable change in both PT and aPTT assays. The correction studies carried out in the FVII deficient plasma at a concentration range of 1 to 0.01 µg/ml did not reveal any differences between 2 groups of drugs. The relative correction was comparable in all concentrations for the 2 tissue factor reagents namely Innovin and Recomboplastin 2G. The results were indistinguishable for the 2 groups of drugs. The effect of NovoSeven® and AryoSeven™ on the reversal of anticoagulation in orally anticoagulated patients' were similar in relative correction of the plasma INR for the 2 groups of FVIIa preparations in the range of 0.03 to 1 µg/ml. In the thrombin generation studies both the Novoseven and Aryoseven produced similar effects as measured by increased thrombin generation. In the primate pharmacokinetic studies both groups exhibited similar pharmacokinetic profile.Conclusions: These results demonstrate that four batches of Novoseven and 5 individual clinical batches of Aryoseven were found to be biosimilar. The pharmacokinetic profile of the Novoseven and Aryoseven were studied in primates measuring the factor VIIa level for 6 hours and were found to be comparable. Thus, the generic Aryoseven is biosimilar to branded Novoseven and warrants further validation studies. DisclosuresNo relevant conflicts of interest to declare.

New technologies for diagnosing active TB: the VANTDET diagnostic accuracy study

Background Tuberculosis (TB) is a devastating disease for which new diagnostic tests are desperately needed. Objective To validate promising new technologies [namely whole-blood transcriptomics, proteomics, flow cytometry and quantitative reverse transcription-polymerase chain reaction (qRT-PCR)] and existing signatures for the detection of active TB in samples obtained from individuals with suspected active TB. Design Four substudies, each of which used samples from the biobank collected as part of the interferon gamma release assay (IGRA) in the Diagnostic Evaluation of Active TB study, which was a prospective cohort of patients recruited with suspected TB. Setting Secondary care. Participants Adults aged ≥ 16 years presenting as inpatients or outpatients at 12 NHS hospital trusts in London, Slough, Oxford, Leicester and Birmingham, with suspected active TB. Interventions New tests using genome-wide gene expression microarray (transcriptomics), surface-enhanced laser desorption ionisation time-of-flight mass spectrometry/liquid chromatography–mass spectrometry (proteomics), flow cytometry or qRT-PCR. Main outcome measures Area under the curve (AUC), sensitivity and specificity were calculated to determine diagnostic accuracy. Positive and negative predictive values were calculated in some cases. A decision tree model was developed to calculate the incremental costs and quality-adjusted life-years of changing from current practice to using the novels tests. Results The project, and four substudies that assessed the previously published signatures, measured each of the new technologies and performed a health economic analysis in which the best-performing tests were evaluated for cost-effectiveness. The diagnostic accuracy of the transcriptomic tests ranged from an AUC of 0.81 to 0.84 for detecting all TB in our cohort. The performance for detecting culture-confirmed TB or pulmonary TB was better than for highly probable TB or extrapulmonary tuberculosis (EPTB), but was not high enough to be clinically useful. None of the previously described serum proteomic signatures for active TB provided good diagnostic accuracy, nor did the candidate rule-out tests. Four out of six previously described cellular immune signatures provided a reasonable level of diagnostic accuracy (AUC = 0.78–0.92) for discriminating all TB from those with other disease and latent TB infection in human immunodeficiency virus-negative TB suspects. Two of these assays may be useful in the IGRA-positive population and can provide high positive predictive value. None of the new tests for TB can be considered cost-effective. Limitations The diagnostic performance of new tests among the HIV-positive population was either underpowered or not sufficiently achieved in each substudy. Conclusions Overall, the diagnostic performance of all previously identified ‘signatures’ of TB was lower than previously reported. This probably reflects the nature of the cohort we used, which includes the harder to diagnose groups, such as culture-unconfirmed TB or EPTB, which were under-represented in previous cohorts. Future work We are yet to achieve our secondary objective of deriving novel signatures of TB using our data sets. This was beyond the scope of this report. We recommend that future studies using these technologies target specific subtypes of TB, specifically those groups for which new diagnostic tests are required. Funding This project was funded by the Efficacy and Mechanism Evaluation (EME) programme, a MRC and NIHR partnership.

Changes of protein levels in human urine reflect the dysregulation of signaling pathways of chronic kidney disease and its complications

The increasing prevalence of chronic kidney disease (CKD) seriously is threatening human health and overall quality of life. The discovery of biomarkers of pathogenesis of CKD and the associated complications are very important for CDK diagnosis and treatment. In this paper, urine protein biomarkers were investigated because urine sample collection is convenient and non-invasive. We analyzed the protein concentrations in the urine of CKD patients and extracted abnormal protein signals comparing with the healthy control groups. The enriched signaling pathways that may characterize CKD pathology were identified from these proteins. We applied surface-enhanced laser desorption and ionization time of flight mass spectrometry technology to detect different protein peaks in urine samples from patients with CKD and healthy controls. We searched the proteins corresponding to protein peaks through the UniProt database and identified the signaling pathways of CKD and its complications by using the NIH DAVID database. 42 low abundance proteins and 46 high abundance proteins in the urine samples from CKD patients were found by comparing with healthy controls. Seven KEGG pathways related to CKD and its complications were identified from the regulated proteins. These pathways included chemokine signaling pathway, cytokine–cytokine receptor interaction, oxidative phosphorylation, cardiac muscle contraction, Alzheimer’s disease, Parkinson's disease, and salivary secretion. In CKD stages 2, 3, 4, and 5, five proteins showed significantly differential abundances. The differential protein signals and regulated signaling pathways will provide new insight for the pathogenesis of CKD and its complications. These altered proteins may also be used as novel biomarkers for the noninvasive and convenient diagnosis methods of CKD and its complications through urine testing in the future.

Development and Characterization of an Avirulent Leishmania major Strain.

Leishmania major causes cutaneous leishmaniasis. An antileishmanial vaccine for humans is unavailable. In this study, we report development of two attenuated L. major strains-5ASKH-HP and LV39-HP-by continuous culture (high passage) of the corresponding virulent strains (low passage). Both avirulent strains showed similar changes in proteome profiles when analyzed by surface-enhanced laser desorption ionization mass spectrometry. Liquid chromatography-mass spectrometry and microarray characterization of 5ASKH strains revealed substantially altered gene and protein expression profiles, respectively. Both virulent and avirulent L. major strains grew comparably in culture, but the avirulent strain survived significantly less in BALB/c-derived peritoneal macrophages. Both attenuated strains failed to infect BALB/c mice and elicited IFN-γ, but not IL-4 and IL-10, responses. 5ASKH-HP parasites failed to induce significant infection even in severely immunocompromised- SCID or inducible NO synthase-, CD40-, or IL-12-deficient mice, indicating attenuation. The avirulent strain induced less IL-10, but higher IL-12, in macrophages. The avirulent strain failed to reduce CD40 relocation to the detergent-resistant membrane domain and to inhibit CD40-induced phosphorylation of the kinases Lyn and protein kinase C-β and MAPKs MKK-3/6 and p38MAPK or to upregulate MEK-1/2 and ERK-1/2 in BALB/c-derived peritoneal macrophages. The virulent and the avirulent strains reciprocally modulated CD40-induced Ras-mediated signaling through PI-3K and Raf-1. Avirulent 5ASKH-primed BALB/c mice were protected against virulent L. major challenge infection. The loss of virulence accompanied by substantially altered proteome profiles and the elicitation of host-protective immune responses indicate plausibly irreversible attenuation of the L. major strain and its potential use as a vaccine strain.

THE ROLE OF NEW TECHNOLOGIES IN DEFINING SALIVARY PROTEIN COMPOSITION FOLLOWING PLACEMENT OF FIXED ORTHODONTIC APPLIANCES - BREAKTHROUGH IN THE DEVELOPMENT OF NOVEL DIAGNOSTIC AND THERAPEUTIC PROCEDURES.

SUMMARYHuman saliva is rich in proteins of variable functions (e.g., enzymes, immunoglobulins, cytokines) and origin (blood plasma, salivary glands, or oral microflora). Circadian dynamics, volume and composition (electrolytes, pH, protein, etc.) of secreted saliva vary with local and systemic physiological and pathophysiological conditions. Therefore, the composition of saliva, protein in particular, has been intensively investigated to identify the potential markers and/or mechanisms of systemic and local diseases. Proteomic techniques used for the analysis of biological fluids have enabled great advances in salivary protein stabilization (as the main precondition for their analysis) and detection of those found in saliva in very low concentrations, including small proteins and peptides. This review brings the main characteristics of current proteomic techniques such as liquid chromatography-mass spectrometry, two-dimensional electrophoresis-mass spectrometry, and surface-enhanced laser desorption ionization/time of flight/mass spectrometry. These techniques enable simultaneous identification of hundreds and thousands of protein molecules, as well as identifying those of a potential biological value in particular states. This literature review is focused on the state-of-the-art and possibilities offered by proteomic techniques in analyzing the effects of orthodontic appliances on salivary protein composition and searching for potential markers of therapeutic success/failure or for the molecules by which therapeutic effects are achieved.

Screening and identification of non-inflammatory specific protein markers in Wilms’ tumor tissues

Wilms' tumor is one of the most common malignancies in children, and early diagnosis is critical for its subsequent treatment and prognosis. Our previous study employed proteomics to investigate protein markers in the serum of Wilms' tumor children. The present study aimed to identify specific protein markers in Wilms' tumor. Proteomic comparison of Wilms' tumor with normal kidney tissues and the sera of systemic inflammatory response syndrome (SIRS) controls was performed. Surface-enhanced laser desorption ionization time-of-flight (SELDI-TOF-MS) identified a protein with m/z 8350 as specific to Wilms' tumor. The target protein was purified using sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and identified as profilin-1 by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight (MALDI-TOF/TOF). Its expression was validated using real-time polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA). Our data identify profilin-1 as a potential protein marker for Wilms’ tumor and demonstrate the feasibility of the above procedures for screening and identification of tumor-specific protein markers.

Magnetic nanocomposite-based SELDI probe for extraction and detection of drugs, amino acids and fatty acids.

The use of carbon black-Fe3O4 magnetic nanocomposite (CB-Fe3O4) as a probe for surface-enhanced laser desorption ionization mass spectrometry (SELDI-MS) with a high extraction efficiency and sensitive detection is described. The magnetic nanocomposite was synthesized and fully characterized using X-ray photoelectron spectroscopy, X-ray diffraction, Raman spectroscopy, Fourier transform infrared spectroscopy, Ultraviolet-Visible spectroscopy, transmission electron microscopy and nitrogen sorption. The feasibility of the SELDI probe to extract and detect three classes of drugs (labetalol, metoprolol, doxepin, desipramine, triprolidine and methapyrilene) spiked in wine is demonstrated. All the drugs were successfully and reproducibly extracted and detected with high efficiency and with limits of detection (LOD) between 1 and 1000pgmL-1. The adsorption capacity of the nanocomposite for the drugs was evaluated by UV-Vis spectroscopy. The results showed that 27.8-36.1% of the drugs were adsorbed on the magnetic probe within 3min. The nanocomposite was also applied for efficient analysis of amino acids and fatty acids. Both types of analytes can be extracted within a few minutes and then successfully quantified by SELDI-MS. Graphical abstract A schematic presentation of carbon black-Fe3O4 magnetic probe for SELDI analysis of small molecules. The probe containing the analyte(s) is collected with the aid of a magnet and deposited on the target plate for mass spectrometry analysis.

Application of protein chip combined with SELDI-TOF-MS detection to investigate serum protein expression in patients with silicosis fibrosis.

The present study aimed to observe the identification of biomarkers of silicosis based on the differentially expressed serum proteins between normal healthy individuals and patients with silicosis fibrosis. A total number of 20 patients with clinically diagnosed silicosis were screened, which were designated as the foundation treatment group. In addition, 20 age-matched healthy patients attending a check-up at the physical examination department were selected. Serum samples were obtained and a combined protein chip with surface-enhanced laser desorption ionization flight mass spectrometry was applied to perform serum analysis. Data preprocessing, screening differences in peak, hierarchical cluster analysis, Principal Component Analysis, construction of a decision tree model, and prediction based on the differences between peaks corresponding to proteins were performed to analyze the data. The results revealed differences in the proteins in serum between the normal group and the group prior to foundation treatment prediction. The corresponding names of the protein peak, predicted protein, and gene name were as follows: M1948_00, complement c3 frag, C3; M2017_02, amyloid-βa4 protein, APP; and M2879_56, hepcidin, HAMP. Differentially expressed serum proteins in the normal group and the basis treatment group were predicted, including M2017_02, amyloid-βa4 protein, APP; M2879_56, hepcidin, HAMP; and M3224_97, fibrinogen-α chain frags, FGA. The differentially expressed serum proteins in the group prior to basis treatment and the group following basis treatment were predicted, including M2001_69, amyloid-βa4 protein, APP; M2017_02, amyloid-βa4 protein, APP, M4144_81, plasma protease c1 inhibitor frag, and SERPING1. In conclusion, there were differences in the proteins in serum between the patients with silicosis fibrosis and healthy individuals.

Experimental Design, Randomization, and Validation.

Featured Article: Baggerly KA, Morris JS, Coombes KR. Reproducibility of SELDI-TOF protein patterns in serum: comparing datasets from different experiments. Bioinformatics 2004;20:777–85.2 Before our article discussed here appeared in 2004, one might have thought a golden age in proteomic diagnostics was at hand. The good news began in 2002 (1), when National Cancer Institute (NCI)3/Food and Drug Administration (FDA) researchers claimed to have processed high-throughput measurements of easy-to-get samples with machine learning algorithms to extract previously elusive diagnoses. The high-throughput measurements were from mass spectrometry, specifically surface-enhanced laser desorption and ionization (SELDI) assays. The samples were minimally processed serum. The machine learning algorithms were, broadly, black boxes taking peak intensities (nominally peptide abundances) and producing categorical calls: “disease,” “no disease,” or “something else.” The diagnosis was whether a woman had ovarian cancer. The numbers looked impressive. Starting with 216 samples (100 women with ovarian cancer, 100 healthy controls, and 16 women with …

Peptidyl-prolyl cis-trans isomerase A – A novel biomarker of multi-episodic (recurrent) ocular toxoplasmosis

Ocular toxoplasmosis (OT) is the most common etiology of posterior uveitis. The high incidence of macular scarring associated with OT is a leading cause of visual morbidity. Serum biomarkers of the disease would aid in its diagnosis. This study sought, for the first time, to elucidate serum biomarkers for OT by mass spectrometry. Blood samples were collected from four groups of nine patients each; toxoplasmosis IgG-with no history of uveitis, non-toxoplasmosis uveitis, first episode OT, and symptomatic recurrent OT. Serum was isolated and subjected to proteomics analysis using 2-dimensional gel electrophoresis (2D-GE) and surface-enhanced laser desorption ionization mass spectrometry (SELDI-MS). Selected proteins were further separated by SDS-PAGE and sequenced using tandem MS. Results were cross-validated with a T. gondii outbreak biomarker database that occurred in Brazil. Fifty markers of OT and 46 markers of recurrent disease were discovered by SELDI-MS of which 30 and 15, respectively, were cross-validated. 2D-GE analysis yielded 57 bands, selected based on the intensity of the bands, leading to the identification of 20 proteins. Eleven of those identified candidates were also found by SELDI-MS. Four candidates were chosen for immunoblotting. One serum protein, peptidyl-prolyl cis-trans isomerase A (PPIA), was confirmed as a biomarker of multi-episodic OT by immunoblotting in patients. PPIA can identify the patient with active recurrent OT from acute OT, other forms of uveitis and other parasitic infections. A validated PPIA assay may have a role in the diagnosis of the atypical OT patient before more invasive anterior chamber or vitreous tap is performed for PCR analysis or for Goldmann-Witner coefficient calculations. Base-line PPIA levels need to be studied to understand its possible use when deciding for prophylactic antibiotic use in the immunosuppressed sero-positive patient.

Proteomic fingerprinting in HIV/HCV co-infection reveals serum biomarkers for the diagnosis of fibrosis staging.

BackgroundHepatic complications of hepatitis C virus (HCV), including fibrosis and cirrhosis are accelerated in human immunodeficiency virus (HIV)-infected individuals. Although, liver biopsy remains the gold standard for staging HCV-associated liver disease, this test can result in serious complications and is subject to sampling errors. These challenges have prompted a search for non-invasive methods for liver fibrosis staging. To this end, we compared serum proteome profiles at different stages of fibrosis in HIV/HCV co- and HCV mono-infected patients using surface-enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF MS).MethodsSera from 83 HIV/HCV co- and 68 HCV mono-infected subjects in 4 stages of fibrosis were tested. Sera were fractionated, randomly applied to protein chip arrays (IMAC, CM10 and H50) and spectra were generated at low and high laser intensities.ResultsSixteen biomarkers achieved a p value < 0.01 (ROC values > 0.75 or < 0.25) predictive of fibrosis status in co-infected individuals and 14 in mono infected subjects. Five of these candidate biomarkers contributed to both mono- and co-infected subjects. Candidate diagnostic algorithms were created to distinguish between non-fibrotic and fibrotic individuals using a panel of 4 biomarker peaks.ConclusionThese data suggest that SELDI MS profiling can identify diagnostic serum biomarkers for fibrosis that are both common and distinct in HIV/HCV co-infected and HCV mono-infected individuals.

MALDI-TOF DAN SELDI-TOF MASS SPECTROMETRY DENGAN THROUGHPUT TINGGI UNTUK ANALISIS PROTEOMIK PROFIL PROTEIN DARI PETANDA BIOLOGIS

In recent years, proteomic approach has been widely used for diagnosing the diseases and matrix assisted laser desorption /ionization- time of the flight (MALDI-TOF) mass spectrometry and its modification, surface enhanced laser desorption /ionization-timeof flight (SELDI-TOF) mass spectrometry have became very promising diagnostic tools. High throughputs and relative simplicity of thesetechnologies attracted the researchers to know the biomarkers of specific diseases by analyzing specimens of serum/plasma and otherbody fluids. Analyzing plasma specimens by MALDI/SELDI TOF mass spectrometry provides new information especially about smallprotein and peptides in high abundance. Protein profilings, resulting by these technologies provide higher sensitivity and specificity valuesthan current biomarkers. Knowing how these principle tools work and its promising application to early detection of specific diseases isthe aim of writing this paper.

Identification of Novel Hemostatic Biomarkers of Adverse Clinical Events in Patients Implanted With a Continuous-Flow Left Ventricular Assist Device.

Heart failure affects over 5 million people in the United States. Its rising prevalence and the limited supply of donor hearts is increasing the use of mechanical cardiac support with the implantation of continuous-flow ventricular assist devices (CF-VAD). Patients with CF-VAD implants are at risk of complications, specifically adverse hemostatic events such as nonsurgical bleeding and thrombosis. Development of a pump thrombus requires clinical intervention and/or surgical replacement significantly increasing the risk of patient morbidity and mortality. Identification of biomarkers for these events could improve current risk assessment models, subsequent treatment, and quality of life prognoses for VAD-implanted patients. The standard means for identifying thrombus in VAD patients is currently limited to monitoring levels of lactate dehydrogenase (>2× upper limit of normal), which is incapable of predicting a future event, but describes the risk of a present thrombus. Surface-enhanced laser desorption ionization time-of-flight mass spectrometry is a technique used to identify biomarkers. In this study, 3 groups of unique peaks were identified in plasma from patients with left ventricular assist devices: 8.1-kDa, 11.7-kDa, and a 15.2-/16.1-kDa pair. Unique correlations were found for each peak, respectively, with microparticles (MPs) and MP procoagulant activity, C-reactive protein, and MP-tissue factor. Furthermore, the use of 8.1-kDa peaks may be able to differentiate thrombotic events from other hemostatic events.

An efficient biomarker panel for diagnosis of breast cancer using surface-enhanced laser desorption ionization time-of-flight mass spectrometry.

Breast cancer (BC) is the most frequently diagnosed cancer that affects women worldwide. Early detection of BC is important to improve survival rates and decrease mortality. The aim of the present study was to investigate serum biomarkers using surface-enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF-MS) to distinguish patients with BC from the healthy population and patients with benign breast diseases (BBDs). A total of 62 patients with invasive ductal carcinoma, as confirmed by histopathology, and 47 non-cancerous individuals (NCIs) [16 healthy controls (HCs) and 31 patients with BBD] were enrolled in the present study. Serum protein profiles were determined by SELDI-TOF-MS using an immobilized metal affinity capture array. Serum from patients with BC were compared with that from the HC group using univariate and multivariate statistical analyses. A total of 118 clusters were generated from the individual serum. Univariate analysis revealed that 5 peaks were significantly downregulated (m/z 1,452, 2,670, 3,972, 5,354 and 5,523; P<0.001) and 4 were upregulated (m/z 6,850, 7,926, 8,115 and 8,143; P<0.001) in patients with BC compared with the HC group. A comparison of patients with BC and patients with BBD revealed an additional 9 protein peaks. Among these, 3 peaks (m/z 3,972, 5,336 and 11,185) were significantly downregulated and 6 peaks (m/z 4,062, 4,071, 4,609, 6,850, 8,115 and 8,133) were significantly upregulated. A total of 3 peaks [mass-to-change ratio (m/z) 3,972, 6,850 and 8,115 (BC2)] were common in both sets. The results of the present study suggest that a 4 protein peak set [m/z 3,972, 6,850 and 8,115 (BC2) and 8,949 (BC3)] could be used to distinguish patients with BC from NCI.

Anaphylatoxin C3a: A potential biomarker for esophageal cancer diagnosis.

Esophageal carcinoma is a common malignancy worldwide, with a low 5-year survival rate. As the majority of cases are diagnosed at an advanced stage, there is an urgent need for an effective biomarker for early diagnosis of esophageal cancer patients. Surface-enhanced laser desorption ionization time-of-flight mass spectrometry (SELDI-TOF-MS) was applied to detect the serum protein expression in esophageal cancer patients using ProteinChip software, and the results were analyzed and screened using Biomarker Patterns and SPSS16.0 software. The ELISA method was conducted to determine the concentration of anaphylatoxin C3a, which is one of the complement proteins, in the serum of esophageal cancer patients and non-esophageal cancer participants. A total of 144 effective differential expression protein peaks in the window of 1-10 kDa were obtained (P<0.05). M/Z 8,926.478 (P<10-6) protein peak was employed as the diagnostic biomarker for esophageal carcinoma. This established diagnostic biomarker has a sensitivity of 95% (19/20) and an accuracy of 100% (19/19) for positive prediction. The results suggested that anaphylatoxin C3a may be a promising biomarker in the diagnosis of esophageal carcinoma, and may play a key role in promoting esophageal carcinogenesis.

A Classification Model to Predict the Rate of Decline of Kidney Function.

The African American Study of Kidney Disease and Hypertension (AASK), a randomized double-blinded treatment trial, was motivated by the high rate of hypertension-related renal disease in the African-American population and the scarcity of effective therapies. This study describes a pattern-based classification approach to predict the rate of decline of kidney function using surface-enhanced laser desorption ionization/time of flight proteomic data from rapid and slow progressors classified by rate of change in glomerular filtration rate. An accurate classification model consisting of 7 out of 5,751 serum proteomic features is constructed by applying the logical analysis of data (LAD) methodology. On cross-validation by 10-folding, the model was shown to have an accuracy of 80.6 ± 0.11%, sensitivity of 78.4 ± 0.17%, and specificity of 78.5 ± 0.16%. The LAD discriminant is used to identify the patients in different risk groups. The LAD risk scores assigned to 116 AASK patients generated a receiver operating curves curve with AUC 0.899 (CI 0.845–0.953) and outperforms the risk scores assigned by proteinuria, one of the best predictors of chronic kidney disease progression.

Danger-recognizing proteins, β-defensin-128 and histatin-3, as potential biomarkers of recurrent coronary events.

Conventional risk factors have limited ability to predict recurrent events in subjects with first-time coronary artery disease(CAD). This aim of this study was to identify novel biomarkers using comparative global proteome analysis to improve the risk assessment for recurrent coronary events. We used samples from phase-I of the Indian Atherosclerosis Research Study(IARS), consisting of 2,332subjects, of whom 772were CAD-affected subjects, including 152with recurrent events identified during a 5-year follow-up period. Global proteome analysis was performed on serum samples of 85subjects with recurrent coronary events and 85age- and gender-matched subjects with first-time CAD using surface-enhanced laser desorption ionization time-of-flight mass spectrometry with CM10 arrays. TagIdent was used for protein identification followed by validation by western blot analysis and ELISA. Data were analyzed by logistic analysis, Cox-regression, hazards ratio, C-statistics and combined-marker risk score using SPSS version-17 and R-package version-2.13.0 software. We identified 16significantly differentially expressed protein peaks. Of these, 2peaks corresponding to m/z8588 and1864 were identified as β-defensin-128 and histatin-3, belonging to the danger-recognizing peptide family, which exhibited a significant independent association with recurrent events(odds ratios of7.49 and1.4, respectively). C-statistics improved significantly from 0.677 for conventional risk factors alone to 0.800(p-value=0.001) in combination with β-defensin-128 and histatin-3 with a hazards ratio of1.833. A combined risk score of β-defensin-128 and histatin-3 could reclassify 112out of the 170subjects into intermediate- and high-risk groups. On the whole, our data indicate that β-defensin-128 and histatin-3 may be potential biomarkers whch may be used to improve risk the stratification for recurrent coronary events.

Serum Protein Profile in Women With Pregnancy Morbidity Associated With Antiphospholipid Syndrome.

Context:Antiphospholipid antibodies (aPL) are related with a high risk of pregnancy morbidity (PM) and also of vascular thrombosis. On the basis of recent studies, we expect that in women with PM associated with antiphospholipid syndrome (APS), further factors may be deregulated and involved in pathophysiology of the disease. Such factors may have the potential to become novel biomarkers for APS and its stages.Settings and Design:Descriptive study from a recurrent pregnancy loss program.Aims:To study the protein expression in sera from women with PM with or without aPL.Materials and Methods:Protein profiles were determined by surface enhanced laser desorption and ionization − time of flight mass spectrometry (SELDI-TOF MS) in the serum samples from women with PM, 10 of them with aPL and 12 without aPL. On the basis of the mass-to-charge ratio (m/z) of the protein, signals differentially expressed between the two groups were compared with data banks to approach candidate proteins.Statistical Analysis Used:To determine the differential expression of each protein, a no paired t-test was performed using Ciphergen Express Client 3.1 software.Results:SELDI-TOF analysis makes it possible to discriminate between several proteins in women with PM with and without aPL, although it does not allow protein identification. Nine proteins were found in significantly higher levels in aPL-positive women.Conclusion:The results underline that further factors beyond autoantibodies are involved in PM associated with APS and might lead to the development of new biomarkers.

Identification & Characterization of Novel Hemostatic Biomarkers of Adverse Clinical Events in Patients with Continuous Flow Left Ventricular Assist Device Implants

Introduction: Heart failure affects over 5 million people in the US. Its rising prevalence coupled with a lack of donor hearts has led to an increase in the use of continuous flow left ventricular assist devices (CF-VAD). However, patients with CF-VAD implants are at risk for complications such as non-surgical bleeding and thrombosis, which significantly increase the risk of patient morbidity and mortality. Identification of novel biomarkers for these events could improve current risk assessment models, treatment, and thus, quality of life for VAD implanted patients.Materials and Methods: 71 blood samples were collected peri-operatively and at varying time points post-operatively from 16 patients implanted with a Thoratec HeartMate II CF-VAD. Plasma was prepared and stored at -70°C until analysis. Plasma samples collected from normal individuals were used as controls. Samples were analyzed for annexin V-positive microparticles and microparticles exposing tissue factor, as well as plasma levels of annexin V and C-reactive protein (CRP) by ELISA. Additionally, surface-enhanced laser desorption ionization - mass spectrometry (SELDI-MS) analysis was used to identify changes in biomarker expression. The occurrence of adverse clinical events was determined using an internal RedCap database. CF-VAD thrombosis was characterized as: cerebrovascular accident/transient ischemic attack diagnosed by a neurologist, rise in LDH or plasma free hemoglobin, hemolysis, evidence of pump dysfunction consistent with thrombus identified by pump parameters, echocardiographic or computed tomographic evidence of clot, or surgical pump exchange for thrombus. CF-VAD bleeding events were characterized as: anemia and bleeding determined by a cardiologist.Results: Annexin V plasma levels were increased approximately 3-fold compared to normal plasma (9.23 ±0.85 vs. 2.76 ±0.2 nM). CRP plasma levels were significantly elevated in CF-VAD patients compared to normals (14.5 ±2.7 vs. 2.2 ± 0.6 µg/ml; p=0.034). Although the level of annexin V-positive microparticles were the same in CF-VAD patients and normals, the amount of microparticles exposing tissue factor was significantly higher in CF-VAD patients (3.7 ± 1.1 vs. 0.5 ± 0.1 pg/ml; p=0.038). SELDI-MS analysis indicated three distinct peaks that were present in CF-VAD patients, but absent from normals (8.1, 11.7 and 15.2/16.2 kDa). During the follow-up period, patients experienced 12 CF-VAD-related thrombotic events, 8 hemorrhagic events and 2 septic events. The 8.1 kDa biomarker was present in 10/12 patients with a thrombotic event, 2/6 patients with a hemorrhagic event, 0/2 patients experiencing sepsis and 0/2 patients without an adverse event. Positive correlations with the SELDI-MS peaks and measured parameters were identified: the 8.1 kDa peak was associated with elevated plasma levels of annexin V (p=0.01) and the presence of annexin V-positive microparticles (p=0.005); the 11.7 kDa peak was associated with elevated plasma levels of CRP (p=0.01); the 15.2/16.2 kDa peaks were associated with the presence of microparticle exposing tissue factor (p=0.002), annexin V-positive microparticles (p<0.001) and elevated plasma levels of annexin V (p=0.03).Conclusions: Despite treatment with low-dose aspirin and warfarin, CF-VAD patients exhibit signs of hemostatic activation. SELDI-MS may be useful in the identification of novel biomarkers of CF-VAD-associated adverse events. This data justifies further evaluation of the utility of the SELDI-MS to readily identify biomarkers related to adverse clinical events in a larger CF-VAD population. DisclosuresNo relevant conflicts of interest to declare.