scholarly journals The beauty of being (label)-free: sample preparation methods for SWATH-MS and next-generation targeted proteomics

F1000Research ◽  
2014 ◽  
Vol 2 ◽  
pp. 272 ◽  
Author(s):  
Jakob Vowinckel ◽  
Floriana Capuano ◽  
Kate Campbell ◽  
Michael J. Deery ◽  
Kathryn S. Lilley ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Protein Identification ◽  
Protein Detection ◽  
Cost Effective ◽  
High Signal ◽  
Label Free ◽  
Signal Variation ◽  
Signal Stability ◽  
Advantages And Disadvantages ◽  
Free Data

The combination of qualitative analysis with label-free quantification has greatly facilitated the throughput and flexibility of novel proteomic techniques. However, such methods rely heavily on robust and reproducible sample preparation procedures. Here, we benchmark a selection of in gel, on filter, and in solution digestion workflows for their application in label-free proteomics. Each procedure was associated with differing advantages and disadvantages. The in gel methods interrogated were cost effective, but were limited in throughput and digest efficiency. Filter-aided sample preparations facilitated reasonable processing times and yielded a balanced representation of membrane proteins, but led to a high signal variation in quantification experiments. Two in solution digest protocols, however, gave optimal performance for label-free proteomics. A protocol based on the detergent RapiGest led to the highest number of detected proteins at second-best signal stability, while a protocol based on acetonitrile-digestion, RapidACN, scored best in throughput and signal stability but came second in protein identification. In addition, we compared label-free data dependent (DDA) and data independent (SWATH) acquisition on a TripleTOF 5600 instrument. While largely similar in protein detection, SWATH outperformed DDA in quantification, reducing signal variation and markedly increasing the number of precisely quantified peptides.

scholarly journals The beauty of being (label)-free: sample preparation methods for SWATH-MS and next-generation targeted proteomics

F1000Research ◽  
2013 ◽  
Vol 2 ◽  
pp. 272 ◽  
Author(s):  
Jakob Vowinckel ◽  
Floriana Capuano ◽  
Kate Campbell ◽  
Michael J. Deery ◽  
Kathryn S. Lilley ◽  
...  
Keyword(s):  
Sample Preparation ◽  
Protein Identification ◽  
Protein Detection ◽  
Cost Effective ◽  
High Signal ◽  
Label Free ◽  
Signal Variation ◽  
Signal Stability ◽  
Advantages And Disadvantages ◽  
Free Data

The combination of qualitative analysis with label-free quantification has greatly facilitated the throughput and flexibility of novel proteomic techniques. However, such methods rely heavily on robust and reproducible sample preparation procedures. Here, we benchmark a selection of in gel, on filter, and in solution digestion workflows for their application in label-free proteomics. Each procedure was associated with differing advantages and disadvantages. The in gel methods interrogated were cost effective, but were limited in throughput and digest efficiency. Filter-aided sample preparations facilitated reasonable processing times and yielded a balanced representation of membrane proteins, but led to a high signal variation in quantification experiments. Two in solution digest protocols, however, gave optimal performance for label-free proteomics. A protocol based on the detergent RapiGest led to the highest number of detected proteins at second-best signal stability, while a protocol based on acetonitrile-digestion, RapidACN, scored best in throughput and signal stability but came second in protein identification. In addition, we compared label-free data dependent (DDA) and data independent (SWATH) acquisition. While largely similar in protein detection, SWATH outperformed DDA in quantification, reducing signal variation and markedly increasing the number of precisely quantified peptides.

Nanotextured Electrical Immunoassays for Ultrasensitive Protein Detection

2011 ◽  
Vol 1346 ◽  
Author(s):  
Timothy O. Mertz ◽  
Krishna Vattipalli ◽  
Tom Barrett ◽  
John Carruthers ◽  
Shalini Prasad
Keyword(s):  
Protein Detection ◽  
Cost Effective ◽  
Disease Diagnosis ◽  
Detection Methods ◽  
Enzyme Linked Immunosorbent Assay ◽  
Detection Accuracy ◽  
Label Free ◽  
Electrode Arrays ◽  
Protein Biomarkers ◽  
Hybrid Sensors

ABSTRACTThis paper describes the development of nanomonitors, which are electrical immunoassays for detection of multiple protein biomarkers. These devices are hybrid sensors with micro-fabricated electrode arrays on a silicon substrate, and integrated nanoporous alumina membranes to provide protein confinement and signal amplification. The disease biomarkers C-reactive protein and Myeloperoxidase have been detected by the nanomonitors in ultra-low concentrations. Proteins were detected in pure samples, human serum, and patient blood samples. The detection accuracy and sensitivity of the nanomonitors in patient samples was comparable to the Enzyme Linked Immunosorbent Assay (ELISA) method of protein detection. Nanomonitors provide the additional benefits of being rapid, label-free, sensitive, and cost effective, providing improvements over traditional protein detection methods, and having potential applications in disease diagnosis.

scholarly journals A Single-Substrate Biosensor with Spin-Coated Liquid Crystal Film for Simple, Sensitive and Label-Free Protein Detection

Biosensors ◽  
2021 ◽  
Vol 11 (10) ◽  
pp. 374
Author(s):  
Po-Chang Wu ◽  
Chao-Ping Pai ◽  
Mon-Juan Lee ◽  
Wei Lee
Keyword(s):  
Liquid Crystal ◽  
Film Thickness ◽  
Limit Of Detection ◽  
Protein Detection ◽  
Cost Effective ◽  
Antibody Concentration ◽  
Label Free ◽  
Free Protein ◽  
Single Substrate ◽  
Simplified Procedure

A liquid crystal (LC)-based single-substrate biosensor was developed by spin-coating an LC thin film on a dimethyloctadecyl[3-(trimethoxysilyl)propyl]ammonium chloride (DMOAP)-decorated glass slide. Compared with the conventional sandwiched cell configuration, the simplified procedure for the preparation of an LC film allows the film thickness to be precisely controlled by adjusting the spin rate, thus eliminating personal errors involved in LC cell assembly. The limit of detection (LOD) for bovine serum albumin (BSA) was lowered from 10−5 g/mL with a 4.2-μm-thick sandwiched cell of the commercial LC E7 to 10−7 g/mL with a 4.2-μm-thick spin-coated E7 film and further to 10−8 g/mL by reducing the E7 film thickness to 3.4 μm. Moreover, by exploiting the LC film of the highly birefringent nematic LC HDN in the immunodetection of the cancer biomarker CA125, an LOD comparable to that determined with a sandwiched HDN cell was achieved at 10−8 g/mL CA125 using a capture antibody concentration an order of magnitude lower than that in the LC cell. Our results suggest that employing spin-coated LC film instead of conventional sandwiched LC cell provides a more reliable, reproducible, and cost-effective single-substrate platform, allowing simple fabrication of an LC-based biosensor for sensitive and label-free protein detection and immunoassay.

A Sample Preparation on Decapsulation Methodology for Effective Failure Analysis on Thin Small Leadless (TSLP) Flip Chip Package with Copper Pillar (CuP) Bump Interconnect Technology

2014 ◽  
Author(s):  
Gwee Hoon Yen ◽  
Ng Kiong Kay
Keyword(s):  
Sample Preparation ◽  
Failure Analysis ◽  
Flip Chip ◽  
Fault Localization ◽  
Cost Effective ◽  
Fault Isolation ◽  
Voltage Contrast ◽  
Interconnect Technology

Abstract Today, failure analysis involving flip chip [1] with copper pillar bump packaging technologies would be the major challenges faced by analysts. Most often, handling on the chips after destructive chemical decapsulation is extremely critical as there are several failure analysis steps to be continued such as chip level fault localization, chip micro probing for fault isolation, parallel lapping [2, 3, 4] and passive voltage contrast. Therefore, quality of sample preparation is critical. This paper discussed and demonstrated a quick, reliable and cost effective methodology to decapsulate the thin small leadless (TSLP) flip chip package with copper pillar (CuP) bump interconnect technology.

scholarly journals Utilising Commercially Fabricated Printed Circuit Boards as an Electrochemical Biosensing Platform

Micromachines ◽  
2021 ◽  
Vol 12 (7) ◽  
pp. 793
Author(s):  
Uroš Zupančič ◽  
Joshua Rainbow ◽  
Pedro Estrela ◽  
Despina Moschou
Keyword(s):  
Printed Circuit Boards ◽  
Electrochemical Sensors ◽  
Cost Effective ◽  
Electrochemical Sensing ◽  
Label Free ◽  
Circuit Boards ◽  
Printed Circuit ◽  
Sensing Applications ◽  
Electrochemical Biosensing ◽  
Pre Treatment

Printed circuit boards (PCBs) offer a promising platform for the development of electronics-assisted biomedical diagnostic sensors and microsystems. The long-standing industrial basis offers distinctive advantages for cost-effective, reproducible, and easily integrated sample-in-answer-out diagnostic microsystems. Nonetheless, the commercial techniques used in the fabrication of PCBs produce various contaminants potentially degrading severely their stability and repeatability in electrochemical sensing applications. Herein, we analyse for the first time such critical technological considerations, allowing the exploitation of commercial PCB platforms as reliable electrochemical sensing platforms. The presented electrochemical and physical characterisation data reveal clear evidence of both organic and inorganic sensing electrode surface contaminants, which can be removed using various pre-cleaning techniques. We demonstrate that, following such pre-treatment rules, PCB-based electrodes can be reliably fabricated for sensitive electrochemical biosensors. Herein, we demonstrate the applicability of the methodology both for labelled protein (procalcitonin) and label-free nucleic acid (E. coli-specific DNA) biomarker quantification, with observed limits of detection (LoD) of 2 pM and 110 pM, respectively. The proposed optimisation of surface pre-treatment is critical in the development of robust and sensitive PCB-based electrochemical sensors for both clinical and environmental diagnostics and monitoring applications.

scholarly journals Continuous-Wave THz Imaging for Biomedical Samples

2020 ◽  
Vol 11 (1) ◽  
pp. 71
Author(s):  
Yaya Zhang ◽  
Chuting Wang ◽  
Bingxin Huai ◽  
Shiyu Wang ◽  
Yating Zhang ◽  
...  
Keyword(s):  
Medical Imaging ◽  
Biomedical Imaging ◽  
Continuous Wave ◽  
Biological Tissues ◽  
Thz Spectroscopy ◽  
Label Free ◽  
Thz Imaging ◽  
Malignant Tumours ◽  
Advantages And Disadvantages ◽  
Imaging Application

In the past few decades, the applications of terahertz (THz) spectroscopy and imaging technology have seen significant developments in the fields of biology, medical diagnosis, food safety, and nondestructive testing. Label-free diagnosis of malignant tumours has been obtained and also achieved significant development in THz biomedical imaging. This review mainly presents the research status and prospects of several common continuous-wave (CW) THz medical imaging systems and applications of THz medical imaging in biological tissues. Here, we first introduce the properties of THz waves and how these properties play a role in biomedical imaging. Then, we analyse both the advantages and disadvantages of the CW THz imaging methods and the progress of these methods in THz biomedical imaging in recent ten years. Finally, we summarise the obstacles in the way of the application of THz bio-imaging application technology in clinical detection, which need to be investigated and overcome in the future.

Plasma enhanced label-free immunoassay for alpha-fetoprotein based on a U-bend fiber-optic LSPR biosensor

RSC Advances ◽  
2015 ◽  
Vol 5 (31) ◽  
pp. 23990-23998 ◽  
Author(s):  
Gaoling Liang ◽  
Zhongjun Zhao ◽  
Yin Wei ◽  
Kunping Liu ◽  
Wenqian Hou ◽  
...  
Keyword(s):  
Surface Plasmon Resonance ◽  
Surface Plasmon ◽  
Plasmon Resonance ◽  
Localized Surface Plasmon Resonance ◽  
Fiber Optic ◽  
Cost Effective ◽  
Alpha Fetoprotein ◽  
Localized Surface Plasmon ◽  
Label Free

A simple, label-free and cost-effective localized surface plasmon resonance (LSPR) immunosensing method was developed for detection of alpha-fetoprotein (AFP).

scholarly journals Electrokinetic Formation of “Microbridges” for Protein Biomarkers as Sensors

2007 ◽  
Vol 12 (5) ◽  
pp. 311-317 ◽  
Author(s):  
Vindhya Kunduru ◽  
Shalini Prasad
Keyword(s):  
Electric Fields ◽  
Microelectrode Array ◽  
Protein Detection ◽  
High Specificity ◽  
Detection Methods ◽  
Label Free ◽  
Protein Biomarkers ◽  
Detection Scheme ◽  
Polystyrene Beads ◽  
Conducting Path

We demonstrate a technique to detect protein biomarkers contained in vulnerable coronary plaque using a platform-based microelectrode array (MEA). The detection scheme is based on the property of high specificity binding between antibody and antigen similar to most immunoassay techniques. Rapid clinical diagnosis can be achieved by detecting the amount of protein in blood by analyzing the protein's electrical signature. Polystyrene beads which act as transportation agents for the immobile proteins (antigen) are electrically aligned by application of homogenous electric fields. The principle of electrophoresis is used to produce calculated electrokinetic movement among the anti-C-reactive protein (CRP), or in other words antibody funtionalized polystyrene beads. The electrophoretic movement of antibody-functionalized polystyrene beads results in the formation of “Microbridges” between the two electrodes of interest which aid in the amplification of the antigen—antibody binding event. Sensitive electrical equipment is used for capturing the amplified signal from the “Microbridge” which essentially behaves as a conducting path between the two electrodes. The technique circumvents the disadvantages of conventional protein detection methods by being rapid, noninvasive, label-free, repeatable, and inexpensive. The same principle of detection can be applied for any receptor—ligand-based system because the technique is based only on the volume of the analyte of interest. Detection of the inflammatory coronary disease biomarker CRP is achieved at concentration levels spanning over the lower microgram/milliliter to higher order nanogram/milliliter ranges.

scholarly journals Label-Free Electrochemical Biosensor Based on Au@MoS₂–PANI for Escherichia coli Detection

Chemosensors ◽  
2021 ◽  
Vol 9 (3) ◽  
pp. 49
Author(s):  
Pushap Raj ◽  
Man Hwan Oh ◽  
Kyudong Han ◽  
Tae Yoon Lee
Keyword(s):  
Escherichia Coli ◽  
Bacterial Infections ◽  
Limit Of Detection ◽  
Bacterial Pathogens ◽  
Cost Effective ◽  
Covalent Immobilization ◽  
Label Free ◽  
Self Assembled Monolayer ◽  
Detection Range ◽  
Linear Detection

Bacterial infections have become a significant challenge in terms of public health, the food industry, and the environment. Therefore, it is necessary to address these challenges by developing a rapid, cost-effective, and easy-to-use biosensor for early diagnosis of bacterial pathogens. Herein, we developed a simple, label-free, and highly sensitive immunosensor based on electrochemical detection using the Au@MoS₂–PANI nanocomposite. The conductivity of the glassy carbon electrode is greatly enhanced using the Au@MoS₂–PANI nanocomposite and a self-assembled monolayer of mercaptopropionic acid on the gold nanoparticle surface was employed for the covalent immobilization of antibodies to minimize the nonspecific adsorption of bacterial pathogens on the electrode surface. The biosensor established a high selectivity and sensitivity with a low limit of detection of 10 CFU/mL, and detected Escherichia coli within 30 min. Moreover, the developed biosensor demonstrated a good linear detection range, practical utility in urine samples, and electrode regenerative studies.

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