scholarly journals Mass Spectrometry-Based Proteomic Investigation of Heterogeneous Biofilms: A Review

2021 ◽  
Vol 18 (2) ◽  
pp. 67-87
Author(s):  
Mohamad Fakhri Yaacob ◽  
◽  
Nur Anisah Johari ◽  
Alya Nur Athirah Kamaruzzaman ◽  
Mohd Fakharul Zaman Raja Yahya ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Large Scale ◽  
Microbial Population ◽  
Developmental Stages ◽  
Wide Spectrum ◽  
High Sensitivity ◽  
High Specificity ◽  
Health Concern ◽  
Proteomics Experiment ◽  
Proteome Expression

Biofilm represents a major public health concern. It is a highly structured and heterogeneous microbial population that is well protected by a hydrated extracellular matrix. In most cases, the difficulties in combating a wide spectrum of biofilm-associated diseases are due to the presence of dormant cells and differential molecular expression. Proteomics is the large-scale and systematic study of cellular proteome expression at any given time by mass spectrometry. It allows high-sensitivity and high-specificity identification of differentially expressed proteins in the biofilms. Over the past few decades, multiple lines of proteomic works have successfully elucidated various aspects of the biofilm including developmental stages, antimicrobial resistance, and survival mechanisms. However, the heterogeneity of biofilms may contribute to inconsistent proteome expression throughout a proteomic experiment. This is due to the fact that the mature biofilm is often associated with the mixture between monolayer and multilayer biofilms, thick microbial population, and chemical gradient of nutrients. This review highlights the biofilm heterogeneities, the principle of mass spectrometry in proteomics, and the possible strategies for quantitative proteomic analysis of heterogeneous biofilms. It is suggested that isolation of monolayer biofilm, laser capture microdissection, flow cytometry, and subtractive proteome profiling may be considered for an accurate and reliable quantitative proteomics experiment.

Direct Analysis of Elastomer Compounds by Soft Ionization, Tandem (MS/MS) and High Resolution (AC-MS) Mass Spectrometry

1995 ◽  
Vol 68 (5) ◽  
pp. 783-793 ◽  
Author(s):  
Robert P. Lattimer
Keyword(s):  
Mass Spectrometry ◽  
Thermoplastic Polyurethane ◽  
High Sensitivity ◽  
High Specificity ◽  
Mass Spectrometric ◽  
Spectrometric Analysis ◽  
Tandem Ms ◽  
Organic Additives ◽  
Soft Ionization ◽  
Modern Mass

Abstract In recent years very effective mass spectrometric methods have been developed for direct polymer compound analysis. The high sensitivity, high specificity, and superb mixture analysis capabilities of modern mass spectrometry make it an invaluable tool in the polymer industry, particularly for qualitative analysis (chemical identification) of organic additives as well as polymeric components. “Survey” mass spectra obtained with soft ionization methods — field ionization (FI-MS) and chemical ionization (CI-MS) — provide diagnostic overviews of chemical composition. The supplemental tandem (MS/MS) and atomic composition (AC-MS) techniques are used to make specific identifications of various organic ingredients. This report describes the direct mass spectrometric analysis of three different elastomer compounds. Organic additives, including curatives, were identified via thermal desorption methods in a commercial EPDM bearing as well as a diene rubber V-belt. The composition of a commercial thermoplastic polyurethane was determined via pyrolysis (Py-CI-MS). These problem-solving examples illustrate the very effective role that mass spectrometry can play in the industrial polymer analysis laboratory.

scholarly journals A Potent Peroxidase from Solid Cell Culture of Ocimum Basilicum with High Sensitivity for Blood Glucose Determination

2021 ◽  
Author(s):  
Parvin Mohammadnejad ◽  
Saeed Soleimani Asl ◽  
Zahra Rasoulian ◽  
Saeed Aminzadeh ◽  
Jaleh Ghashghaie ◽  
...  
Keyword(s):  
Cell Culture ◽  
Large Scale ◽  
Specific Activity ◽  
High Sensitivity ◽  
High Specificity ◽  
Global Market ◽  
Ocimum Basilicum ◽  
Scale Production ◽  
Market Demand ◽  
Glucose Determination

Abstract Objectives Extensive applications of peroxidase (POX) have raised the global market demand at a considerable rate during the forecast period of 2020 - 2025. Nonetheless, the large-scale POX preparation still relies on the extraction from agricultural products, while there is an accumulative driving force toward employing biotechnological processes with agricultural hassle free identity. In pursuit of this trend; Results, A novel heme peroxidase was purified to homogeneity (MW of 40 kD) from the callus culture of Ocimum basilicum L. in darkness on Murashige-Skoog medium supplemented by 2,4-dichlorophenoxyacetic acid (10-6 M) and kinetin (10-5 M). The highest activity of the purified peroxidase (ObPOX) was observed in Tris-base buffer at pH 7.5 and 80 °C. ObPOX showed high stability over pH(s) 5 to 7.5 and temperatures of 15 to 60 °C. ObPOX specific activity was 1245.142 AU mg-1 in the presence of phenol, 4 times higher than that of HRP. ObPOX showed moderate affinity for guaiacol (Km = 21.5 mM), but obtained an exceptionally high specificity constant (kcat/Km = 66743.7 s-1M-1) for GASA (4-[(4-Hydroxy-3-methoxyphenyl) azo]-benzenesulfonic acid), the introduced substrate for determination of blood sugar. Applying ObPOX instead of HRP in glucose measurements of the real samples improved the regression constant of the correlation diagram between the tests and the lab results from 0.958 to 0.981. Conclusion Physicochemical properties of ObPOX as well as the growth rate of basil callus (5.04 g L-1 per day) and the yield of ObPOX production (35 mg per 100 g dry biomass per subculture) designates O. basilicum cell culture for large-scale production of a robust peroxidase.

scholarly journals A new Syber Green real time PCR to detect SARS-CoV-2

2020 ◽  
Author(s):  
D.R. Marinowic ◽  
G. Zanirati ◽  
F.V.F. Rodrigues ◽  
M.V.C. Grahl ◽  
A.M. Alcará ◽  
...  
Keyword(s):  
Diagnostic Test ◽  
Large Scale ◽  
Low Cost ◽  
Phylogenetic Analyses ◽  
High Sensitivity ◽  
High Specificity ◽  
Cost Effective ◽  
Etiologic Agent ◽  
Rt Pcr ◽  
Human Respiratory Tract

Abstract Phylogenetic analyses demonstrated that etiologic agent of pandemic outbreak is a betacoronavirus named SARS-CoV-2. For public health interventions, a diagnostic test with high sensitivity and specificity is required. The gold standard protocol for diagnosis by WHO is the RT-PCR. To detect low viral load and large-scale screening a low-cost diagnostic test becomes necessary. Here we develop a cost-effective test capable of to detect the new coronavirues. We validated an auxiliary protocol for molecular diagnosis with RT-PCR SYBR Green methodology to successfully screen negative cases of SARS-CoV-2. Our results demonstrated that a set of primers with high specificity, and no homology with other viruses from Coronovideae family or human respiratory tract pathogenic viruses. Optimization of annealing temperature and polymerization time led to an high specificity in the PCR products. We have developed a more affordable and swift methodology for negative SARS-CoV-2 screening. This methodology can be applied on large scale populational to soften panic and economic burden through guidance for isolation strategies.

Gold Nanoparticles-Amplified Indirect Competitive Enzyme-Linked Immunosorbent Assay of BDE-121 Using a Monoclonal Antibody

2021 ◽  
Vol 21 (10) ◽  
pp. 5036-5043
Author(s):  
Yan Qiao ◽  
Qing-Yun Cai
Keyword(s):  
Mass Spectrometry ◽  
Monoclonal Antibody ◽  
Gold Nanoparticles ◽  
Immunosorbent Assay ◽  
Inhibitory Concentration ◽  
High Sensitivity ◽  
High Specificity ◽  
Gas Chromatography Mass Spectrometry ◽  
Enzyme Linked Immunosorbent Assay ◽  
Good Recovery

In this study, we developed a monoclonal antibody against 2,3’,4,5’,6-pentabromodiphenylether (BDE-121) using a synthesized hapten, and established an indirect competitive enzyme-linked immunosorbent assay (IC-ELISA), using gold nanoparticles, to amplify the signal. The monoclonal antibody showed high specificity, with a half inhibitory concentration (IC50) value of 2.78 ng/mL, towards BDE-121. The developed IC-ELISA exhibited high sensitivity and stability as well as good recovery. The intra-assay deviation is below 6.8% and the inter-assay deviations range from 6.5% to 8.7%. The assay of the actual samples was found to be consistent with those of gas chromatography/mass spectrometry (GC/MS).

Hyphenated Mass Spectrometry Techniques in the Diagnosis of Amyloidosis

2019 ◽  
Vol 26 (1) ◽  
pp. 104-120 ◽  
Author(s):  
Marta Spodzieja ◽  
Sylwia Rodziewicz-Motowidło ◽  
Aneta Szymanska
Keyword(s):  
Mass Spectrometry ◽  
Biological Samples ◽  
Large Scale ◽  
Amyloid Fibrils ◽  
High Sensitivity ◽  
Diagnostic Procedures ◽  
Comprehensive Information ◽  
Diagnostic Biopsy ◽  
Separation Of Proteins ◽  
Subcutaneous Adipose

Amyloidoses are a group of diseases caused by the extracellular deposition of proteins forming amyloid fibrils. The amyloidosis is classified according to the main protein or peptide that constitutes the amyloid fibrils. The most effective methods for the diagnosis of amyloidosis are based on mass spectrometry. Mass spectrometry enables confirmation of the identity of the protein precursor of amyloid fibrils in biological samples with very high sensitivity and specificity, which is crucial for proper amyloid typing. Due to the fact that biological samples are very complex, mass spectrometry is usually connected with techniques such as liquid chromatography or capillary electrophoresis, which enable the separation of proteins before MS analysis. Therefore mass spectrometry constitutes an important part of the so called “hyphenated techniques” combining, preferentially in-line, different analytical methods to provide comprehensive information about the studied problem. Hyphenated methods are very useful in the discovery of biomarkers in different types of amyloidosis. In systemic forms of amyloidosis, the analysis of aggregated proteins is usually performed based on the tissues obtained during a biopsy of an affected organ or a subcutaneous adipose tissue. In some cases, when the diagnostic biopsy is not possible due to the fact that amyloid fibrils are formed in organs like the brain (Alzheimer’s disease), the study of biomarkers presented in body fluids can be carried out. Currently, large-scale studies are performed to find and validate more effective biomarkers, which can be used in diagnostic procedures. We would like to present the methods connected with mass spectrometry which are used in the diagnosis of amyloidosis based on the analysis of proteins occurring in tissues, blood and cerebrospinal fluid.

Analysis of Components in Rubber Compounds Using Mass Spectrometry

1989 ◽  
Vol 62 (3) ◽  
pp. 548-567 ◽  
Author(s):  
Robert P. Lattimer ◽  
Robert E. Harris
Keyword(s):  
Mass Spectrometry ◽  
High Sensitivity ◽  
High Specificity ◽  
Mass Spectrometric ◽  
Rubber Compound ◽  
Organic Additives ◽  
Mixture Analysis ◽  
Rubber Compounds ◽  
Invaluable Tool ◽  
Modern Mass

Abstract A large number of very successful mass-spectrometric methods have been developed for rubber-compound analysis. The high sensitivity, high specificity, and superb mixture analysis capabilities of modern mass spectrometry make it an invaluable tool in the polymer industry, particularly for qualitative identification of organic additives. In many cases, mass spectrometry can provide unique information that is not available by use of any other technique.

scholarly journals A new SYBR Green real-time PCR to detect SARS-CoV-2

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
D. R. Marinowic ◽  
G. Zanirati ◽  
F. V. F. Rodrigues ◽  
M. V. C. Grahl ◽  
A. M. Alcará ◽  
...  
Keyword(s):  
Diagnostic Test ◽  
Large Scale ◽  
Low Cost ◽  
High Sensitivity ◽  
High Specificity ◽  
Cost Effective ◽  
Etiologic Agent ◽  
Sybr Green ◽  
Rt Pcr ◽  
Human Respiratory Tract

AbstractPhylogenetic analysis has demonstrated that the etiologic agent of the 2020 pandemic outbreak is a betacoronavirus named SARS-CoV-2. For public health interventions, a diagnostic test with high sensitivity and specificity is required. The gold standard protocol for diagnosis by the Word Health Organization (WHO) is RT-PCR. To detect low viral loads and perform large-scale screening, a low-cost diagnostic test is necessary. Here, we developed a cost-effective test capable of detecting SARS-CoV-2. We validated an auxiliary protocol for molecular diagnosis with the SYBR Green RT-PCR methodology to successfully screen negative cases of SARS-CoV-2. Our results revealed a set of primers with high specificity and no homology with other viruses from the Coronovideae family or human respiratory tract pathogenic viruses, presenting with complementarity only for rhinoviruses/enteroviruses and Legionella spp. Optimization of the annealing temperature and polymerization time led to a high specificity in the PCR products. We have developed a more affordable and swift methodology for negative SARS-CoV-2 screening. This methodology can be applied on a large scale to soften panic and economic burden through guidance for isolation strategies.

scholarly journals Steroid metabolites for diagnosing and predicting clinicopathological features in cortisol-producing adrenocortical carcinoma.

2020 ◽  
Author(s):  
Sawako Suzuki ◽  
Tomoki Minamidate ◽  
Akina Shiga ◽  
Yutarou Ruike ◽  
Kazuki Ishiwata ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Steroid Hormone ◽  
High Sensitivity ◽  
High Specificity ◽  
Clinicopathological Features ◽  
Gas Chromatography Mass Spectrometry ◽  
Large Area ◽  
Urine Metabolites ◽  
Positive Correlations ◽  
Adrenocortical Carcinomas

Abstract Background: Approximately 60% of adrenocortical carcinomas (ACC) are functional, and Cushing’s syndrome is the most frequent diagnosis that has been revealed to have a particularly poor prognosis. Since 30% of ACC present steroid hormone-producing disorganization, measurement of steroid metabolites in suspected ACC is recommended. Previous reports demonstrated that steroid hormone precursors or their urine metabolites, which can be assessed using liquid chromatography tandem mass spectrometry (LC-MS/MS) or gas chromatography mass spectrometry (GC-MS) respectively, are useful for distinguishing ACC from cortisol-producing adenomas (CPA); however, despite high precision, LC-MS/MS and GC-MS require a highly trained team, are expensive and have limited capacity. Methods: Here, we examined 12 serum steroid metabolites using an immunoassay, which is a more rapid and less costly method than LC-MS/MS, in cortisol-producing ACC and CPA. Further, the correlation of each steroid metabolite to the classification stage and pathological status in ACC was analyzed. Results: Reflecting disorganized steroidogenesis, the immunoassay revealed that all basal levels of steroid precursors were significantly increased in cortisol-producing ACC compared to CPA; in particular, 17-hydroxypregnenolone (glucocorticoid and androgen precursor) and 11-deoxycorticosterone (mineralocorticoid precursor) showed a large area under the ROC curve with high sensitivity and specificity when setting the cut-off at 1.78 ng/ml and 0.4 mg/ml, respectively. Additionally, a combination of androstenedione and DHEAS also showed high specificity with high accuracy. In cortisol-producing ACC, 11-deoxycortisol (glucocorticoid precursor) showed significant positive correlations with predictive prognostic factors used in ENSAT classification, while testosterone showed significant positive correlations to the Ki67-index in both men and women. Conclusion: Less expensive and more widely available RIA and ECLIA may also biochemically distinguish ACC from CPA and may predict the clinicopathological features of ACC.

scholarly journals Steroid metabolites for diagnosing and predicting clinicopathological features in cortisol-producing adrenocortical carcinoma.

2020 ◽  
Author(s):  
Sawako Suzuki ◽  
Tomoki Minamidate ◽  
Akina Shiga ◽  
Yutarou Ruike ◽  
Kazuki Ishiwata ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Steroid Hormone ◽  
High Sensitivity ◽  
High Specificity ◽  
Clinicopathological Features ◽  
Gas Chromatography Mass Spectrometry ◽  
Large Area ◽  
Urine Metabolites ◽  
Positive Correlations ◽  
Adrenocortical Carcinomas

Abstract Background: Approximately 60% of adrenocortical carcinomas (ACC) are functional, and Cushing’s syndrome is the most frequent diagnosis that has been revealed to have a particularly poor prognosis. Since 30% of ACC present steroid hormone-producing disorganization, measurement of steroid metabolites in suspected ACC is recommended. Previous reports demonstrated that steroid hormone precursors or their urine metabolites, which can be assessed using liquid chromatography tandem mass spectrometry (LC-MS/MS) or gas chromatography mass spectrometry (GC-MS) respectively, are useful for distinguishing ACC from cortisol-producing adenomas (CPA); however, despite high precision, LC-MS/MS and GC-MS require a highly trained team, are expensive and have limited capacity. Methods: Here, we examined 12 serum steroid metabolites using an immunoassay, which is a more rapid and less costly method than LC-MS/MS, in cortisol-producing ACC and CPA. Further, the correlation of each steroid metabolite to the classification stage and pathological status in ACC was analyzed. Results: Reflecting disorganized steroidogenesis, the immunoassay revealed that all basal levels of steroid precursors were significantly increased in cortisol-producing ACC compared to CPA; in particular, 17-hydroxypregnenolone (glucocorticoid and androgen precursor ) and 11-deoxycorticosterone (mineralocorticoid precursor) showed a large area under the ROC curve with high sensitivity and specificity when setting the cut-off at 1.78 ng/ml and 0.4 mg/ml, respectively. Additionally, a combination of androstenedione and DHEAS also showed high specificity with high accuracy. In cortisol-producing ACC, 11-deoxycortisol (glucocorticoid precursor) showed significant positive correlations with predictive prognostic factors used in ENSAT classification, while testosterone showed significant positive correlations to the Ki67-index in both men and women.Conclusion: Less expensive and more widely available RIA and ECLIA may also biochemically distinguish ACC from CPA and may predict the clinicopathological features of ACC.

scholarly journals A Potent Peroxidase from Solid Cell Culture of Ocimum Basilicum with High Sensitivity for Blood Glucose Determination

2021 ◽  
Author(s):  
Parvin Mohammadnejad ◽  
Saeed Soleimani Asl ◽  
Zahra Rasoulian ◽  
Saeed Aminzadeh ◽  
Jaleh Ghashghaie ◽  
...  
Keyword(s):  
Cell Culture ◽  
Large Scale ◽  
Specific Activity ◽  
High Sensitivity ◽  
High Specificity ◽  
Global Market ◽  
Ocimum Basilicum ◽  
Scale Production ◽  
Market Demand ◽  
Glucose Determination

Abstract Extensive applications of peroxidase (POX) have raised the global market demand at a considerable rate during the forecast period of 2020 - 2025. Nonetheless, the large-scale POX preparation still relies on the extraction from agricultural products, while there is an accumulative driving force toward employing biotechnological processes with agricultural hassle free identity. In pursuit of this trend, a novel heme peroxidase was purified to homogeneity (MW of 40 kD) from the callus culture of Ocimum basilicum L. in darkness on Murashige-Skoog medium supplemented by 2,4-dichlorophenoxyacetic acid (10-6 M) and kinetin (10-5 M). The highest activity of the purified peroxidase (ObPOX) was observed in Tris-base buffer at pH 7.5 and 80 °C. ObPOX showed high stability over pH(s) 5 to 7.5 and temperatures of 15 to 60 °C. ObPOX specific activity was 1245.142 AU mg-1 in the presence of phenol, 4 times higher than that of HRP. ObPOX showed moderate affinity for guaiacol (Km = 21.5 mM), but obtained an exceptionally high specificity constant (kcat/Km = 66743.7 s-1M-1) for GASA (4-[(4-Hydroxy-3-methoxyphenyl) azo]-benzenesulfonic acid), the introduced substrate for determination of blood sugar. Applying ObPOX instead of HRP in glucose measurements of the real samples improved the regression constant of the correlation diagram between the tests and the lab results from 0.958 to 0.981. Physicochemical properties of ObPOX as well as the growth rate of basil callus (5.04 g L-1 per day) and the yield of ObPOX production (35 mg per 100 g dry biomass per subculture) designates O. basilicum cell culture for large-scale production of a robust peroxidase.

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