Surface-enhanced Laser Desorption And Ionization Research Articles

Comparison of algorithms for pre-processing of SELDI-TOF mass spectrometry data

Surface-enhanced laser desorption and ionization (SELDI) time of flight (TOF) is a mass spectrometry technology. The key features in a mass spectrum are its peaks. In order to locate the peaks and quantify their intensities, several pre-processing steps are required. Though different approaches to perform pre-processing have been proposed, there is no systematic study that compares their performance. In this article, we present the results of a systematic comparison of various popular packages for pre-processing of SELDI-TOF data. We evaluate their performance in terms of two of their primary functions: peak detection and peak quantification. Regarding peak quantification, the performance of the algorithms is measured in terms of reproducibility. For peak detection, the comparison is based on sensitivity and false discovery rate. Our results show that for spectra generated with low laser intensity, the software developed by Ciphergen Biosystems (ProteinChip Software 3.1 with the additional tool Biomarker Wizard) produces relatively good results for both peak quantification and detection. On the other hand, for the data produced with either medium or high laser intensity, none of the methods show uniformly better performances under both criteria. Our analysis suggests that an advantageous combination is the use of the packages MassSpecWavelet and PROcess, the former for peak detection and the latter for peak quantification.

Cover Picture: Proteomics – Clinical Applications 7‐8/2008

AbstractIn this issue of Proteomics – Clinical Applications you will find the following highlighted articles:Hi ho SELDI, away!That closing cry of the Lone Ranger meant that everything was under control. The bad guy was either dead or in jail, the good guy had his job or his ranch back. Here it means that O'Riordan et al. have begun to unearth some of the urinary proteome changes that result in chronic allograft nephropathy (CAN). Based on studies of 75 renal transplant patients and 20 healthy controls using surface enhanced laser desorption and ionization (SELDI), patients could be classified by stages. Advanced CAN associated proteins included α1‐microglobulin, β2 microglobulin, prealbumin, and endorepellin (the anti‐angiogenic C‐terminus of perlecan). Statistical analysis suggests that urinary endorepellin serves as a biomarker for advanced CAN.O'Riordan, E. et al., Proteomics 2008, 8, 1025–1035.Contrast response prediction, detection: “First, do no harm” reduxIt is sometimes difficult to deal with the tradeoffs faced in medical practice…we may do harm while doing good. In this case, using a radio‐opaque contrast agent to determine the state of a child's heart in preparation for surgery may lead to acute kidney injury (AKI), requiring an extended stay, dialysis, or increased chance of death for the next 6 months to 5 years. Contrast‐induced nephropathy (CIN) is the third most frequent cause of hospital acquired AKI. Bennett et al. used proteomic tools (SELDI‐TOF‐MS) to examine urine for biomarkers that would predict possible difficulties and found two good candidates: a down‐regulated 41 amino acid variant of beta‐defensin‐1 and an up‐regulated protein of 4631 Da of unknown origin. Both predicted CIN before the procedure. At present, less toxic contrast agents are also less effective.Bennett, M. R. et al., Proteomics 2008, 8, 1058–1064.Sugar, daddy!Regulation of glucose levels is a collaborative effort of the pancreas and kidneys. Excessive levels of the sugar lead to a variety of pathological conditions, including high levels of reactive oxygen species. So et al. have examined the changes that occur in protein expression in cultured kidney cell line HK‐2. They found high glucose caused twelve differential changes in HK‐2 cells, five of which were significant. Enolase 1 and triosephosphate isomerase are the two up‐regulated enzymes, while glyceraldehyde phosphate dehydrogenase, pyruvate dehydrogenase, and lactate dehydrogenase are down‐regulated. Mitochondrial ATP synthase subunit d is also affected by glucose levels. Knowing the names of some of the players in the diabetes game is a help, but much remains to be explored and explained before the “Case Closed” stamp is called for.So, E.‐J. et al., Proteomics 2008, 8, 1118–1126.

In this issue: Proteomics – Clinical Applications 7‐8/2008

AbstractIn this issue of Proteomics – Clinical Applications you will find the following highlighted articles:Hi ho SELDI, away!That closing cry of the Lone Ranger meant that everything was under control. The bad guy was either dead or in jail, the good guy had his job or his ranch back. Here it means that O'Riordan et al. have begun to unearth some of the urinary proteome changes that result in chronic allograft nephropathy (CAN). Based on studies of 75 renal transplant patients and 20 healthy controls using surface enhanced laser desorption and ionization (SELDI), patients could be classified by stages. Advanced CAN associated proteins included α1‐microglobulin, β2 microglobulin, prealbumin, and endorepellin (the anti‐angiogenic C‐terminus of perlecan). Statistical analysis suggests that urinary endorepellin serves as a biomarker for advanced CAN.O'Riordan, E. et al., Proteomics 2008, 8, 1025–1035.Contrast response prediction, detection: “First, do no harm” reduxIt is sometimes difficult to deal with the tradeoffs faced in medical practice…we may do harm while doing good. In this case, using a radio‐opaque contrast agent to determine the state of a child's heart in preparation for surgery may lead to acute kidney injury (AKI), requiring an extended stay, dialysis, or increased chance of death for the next 6 months to 5 years. Contrast‐induced nephropathy (CIN) is the third most frequent cause of hospital acquired AKI. Bennett et al. used proteomic tools (SELDI‐TOF‐MS) to examine urine for biomarkers that would predict possible difficulties and found two good candidates: a down‐regulated 41 amino acid variant of beta‐defensin‐1 and an up‐regulated protein of 4631 Da of unknown origin. Both predicted CIN before the procedure. At present, less toxic contrast agents are also less effective.Bennett, M. R. et al., Proteomics 2008, 8, 1058–1064.Sugar, daddy!Regulation of glucose levels is a collaborative effort of the pancreas and kidneys. Excessive levels of the sugar lead to a variety of pathological conditions, including high levels of reactive oxygen species. So et al. have examined the changes that occur in protein expression in cultured kidney cell line HK‐2. They found high glucose caused twelve differential changes in HK‐2 cells, five of which were significant. Enolase 1 and triosephosphate isomerase are the two up‐regulated enzymes, while glyceraldehyde phosphate dehydrogenase, pyruvate dehydrogenase, and lactate dehydrogenase are down‐regulated. Mitochondrial ATP synthase subunit d is also affected by glucose levels. Knowing the names of some of the players in the diabetes game is a help, but much remains to be explored and explained before the “Case Closed” stamp is called for.So, E.‐J. et al., Proteomics 2008, 8, 1118–1126.

Putting the “Bio” back into Biomarkers: Orienting Proteomic Discovery toward Biology and away from the Measurement Platform

The discipline of mass spectrometry (MS)-based serum biomarker profiling is a relatively young field that was launched in a flourish of scientific hope and the clinical promise of a better cancer test (1)(2)(3)(4)(5). Nevertheless, the field has already undergone a rollercoaster cycle of optimism and disappointment and renewed enthusiasm during its brief 7-year history (1)(2)(3)(4)(5)(6)(7)(8). Before 1999 very little effort was made to directly use MS (MALDI-TOF or ES) as a means to discover new blood biomarkers. Many scientists thought that serum was too complex for MS analysis, and dominated by contaminants, rendering these samples unacceptable for direct introduction into expensive and sophisticated MS research instruments. All of these arguments seemed to be overturned in 1998 when a new class of MALDI-TOF, surface-enhanced laser desorption and ionization (SELDI) was commercialized. Scientists with no formal training in MS saw SELDI as an opportunity to explore the application of MALDI MS to biomarker discovery. Although this method had relatively low resolution, it appeared to provide a fresh, one-step approach to the search for ion signatures of hundreds of candidate biomarkers. Importantly, SELDI-TOF offered a means to support a critical new hypothesis that was emerging in the protein biomarker field. This hypothesis abandoned the assumption that a single specific tumor cell–derived cancer biomarker existed. Instead, cascades of biomarkers were generated from the tumor tissue microenvironment through interactions between the tumor cells and the host cells (e.g., endothelial cells, stroma cells, and immune cells) (1)(5). The tissue-microenvironment hypothesis predicted that a panel of biomarkers could achieve a sensitivity and specificity superior to previous failed cancer biomarker searches (1)(5)(6)(7)(8). In 2002 SELDI provided …

Investigation of human embryo-conditioned IVF culture medium through the use of surface enhanced laser desorption and ionization (SELDI) mass spectrometry

Investigation of human embryo-conditioned IVF culture medium through the use of surface enhanced laser desorption and ionization (SELDI) mass spectrometry

Diurnal protein expression in blood revealed by high throughput mass spectrometry proteomics and implications for translational medicine and body time of day

Molecular gene cycling is useful for determining body time of day (BTOD) with important applications in personalized medicine, including cardiovascular disease and cancer, our leading causes of death. However, it impractically requires repetitive invasive tissue sampling that is obviously not applicable for humans. Here we characterize diurnal protein cycling in blood using high-throughput proteomics; blood proteins are easily accessible, minimally invasive, and can importantly serve as surrogates for what is happening elsewhere in the body in health and disease. As proof of the concept, we used normal C57BL/6 mice maintained under regular 24-h light and dark cycles. First, we demonstrated fingerprint patterns in 24-h plasma, revealed using surface-enhanced laser desorption and ionization (SELDI). Second, we characterized diurnal cycling proteins in blood using chromatography and tandem electrospray ionization mass spectrometry. Importantly, we noted little association between the cycling blood proteome and tissue transcriptome, delineating the necessity to identify de novo cycling proteins in blood for measuring BTOD. Furthermore, we explored known interaction networks to identify putative functional pathways regulating protein expression patterns in blood, thus shedding new light on our understanding of integrative physiology. These studies have profound clinical significance in translating the concept of BTOD to the practical realm for molecular diagnostics and open new opportunities for clinically relevant discoveries when applied to ELISA-based molecular testing and/or point-of-care devices.

1141644472 Surface Enhanced Laser Desorption and Ionization (SELDI) mass spectrometry for investigation of human embryo conditioned IVF culture medium

Introduction: The human pre‐implantation embryo metabolism is still widely unknown. Mass spectrometry is a very sensitive proteomic method to detect the entire spectrum of molecules in samples below 1 μL. Surface Enhanced Laser Desorption Ionisation – Time Of Flight (SELDI – TOF) mass spectrometry is a further advanced method to reduce overlapping signals from different substances.Material and Methods: A total of 22 different pre‐implantation embryo supernatants (10 μL each) from 16 different single‐cultured embryos were analyzed. From 6 embryos, supernatants were obtained both at day 3 and 5. Embryo stage was defined by microscopy. Different stages including arrested embryos, 4‐ or 8‐cell stage, early and expanded blastocysts were investigated. SELDI combines a chip‐based separation technology with mass spectrometry. All samples were fractioned by applying six different pH milieus and then exposed to chips with six differently coated surfaces. All fractions were analyzed on a mass spectrometer type ProteinChip System, Series 4000 (Ciphergen Biosystems, Inc.).Results: By comparing the protein profiles of the analyzed embryos the implemented bioinformatic software was able to classify blinded samples correctly into groups of embryos as defined by microscopy. The use of IMAC‐chips (copper‐coated) and CM10‐chips (cation‐exchange) led to highest resolution of peaks. Although the masses of numerous molecules in culture media are assessed by this method, the exact definition of substances needs application of further techniques.Conclusions: These first SELDI mass spectrometric analyses of embryo culture media demonstrate the capability of discriminating different stages of human pre‐implantation embryos by their protein expression profiles.

Toward functional proteomics of alveolar macrophages

Alveolar macrophages (AM) belong to a phenotype of macrophages with distinct biological functions and important pathophysiological roles in lung health and disease. The molecular details determining AM differentiation from blood monocytes and AM roles in lung homeostasis are largely unknown. With the use of different technological platforms, advances in the field of proteomics have made it possible to search for differences in protein expression between AM and their precursor monocytes. Proteome features of each cell type provide new clues into understanding mononuclear phagocyte biology. In-depth analyses using subproteomics and subcellular proteomics offer additional information by providing greater protein resolution and detection sensitivity. With the use of proteomic techniques, large-scale mapping of phosphorylation differences between the cell types have become possible. Furthermore, two-dimensional gel proteomics can detect germline protein variants and evaluate the impact of protein polymorphisms on an individual's susceptibility to disease. Finally, surface-enhanced laser desorption and ionization (SELDI) time-of-flight mass spectrometry offers an alternative method to recognizing differences in protein patterns between AM and monocytes or between AM under different pathological conditions. This review details the current status of this field and outlines future directions in functional proteomic analyses of AM and monocytes. Furthermore, this review presents viewpoints of integrating proteomics with translational topics in lung diseases to define the mechanisms of disease and to uncover new diagnostic and therapeutic targets.

Serum proteomics profiling—a young technology begins to mature

During the three years since the US National Cancer Institute–Food and Drug Administration (NCI-FDA) proteomics group published their seminal1 (but flawed2–4) study using mass spectrometry to profile the serum proteome of ovarian cancer patients, more than 60 published studies have applied similar technology to a wide range of cancers and other diseases. Although some studies have looked directly at tumor tissue, most have targeted easily accessible fluids such as serum. A few studies used highresolution instruments with a matrix-assisted laser desorption and ionization (MALDI) ion source and a time-of-flight (TOF) ion detector. The majority, however, have relied on lower resolution instruments and surface-enhanced laser desorption and ionization (SELDI), a MALDI variant that uses commercially prepared chromatographic surfaces (ProteinChips; Ciphergen, Fremont, CA, USA) to separate proteins. These studies have a common goal: to identify biomarker patterns in the proteome that can be used for diagnosis, prognosis or monitoring of disease. The preponderance of evidence from these studies seems to suggest that proteomic profiling is able to detect anonymous protein peaks that are expressed differently in cancer patients than in healthy individuals5–7. The studies also provide evidence that we may be able to discover patterns that are useful for prediction of the presence or absence of disease8–11. For serum proteomics to realize its full potential, however, several problems in sensitivity and reproducibility remain unresolved. This article outlines some possible improvements in experimental design and data analysis that might help in this regard.

Serum diagnosis of pancreatic adenocarcinoma using surface-enhanced laser desorption and ionization mass spectrometry.

Each year in the United States, approximately 30,000 people die from pancreatic cancer. Fewer than 5% of patients survive >5 years after diagnosis, because most patients present with advanced disease. Early diagnosis may improve the prognosis of patients with pancreatic cancer. In an attempt to improve on current approaches to the serological diagnosis of pancreatic cancer, we analyzed serum samples from patients with and without pancreatic cancer using surface-enhanced laser desorption and ionization (SELDI) protein chip mass spectrometry. Using a case-control study design, serum samples from 60 patients with resectable pancreatic adenocarcinoma were compared with samples from 60 age- and sex-matched patients with nonmalignant pancreatic diseases, as well as 60 age- and sex-matched healthy controls. To increase the number of proteins potentially identifiable, serum was fractionated using anion exchange and profiled on two ProteinChip surfaces (metal affinity capture and weak cation exchange). We determined a minimum set of protein peaks able to discriminate between patient groups and used the unified maximum separability algorithm to compare the performance of the individual marker panels alone or in conjunction with CA19-9. Among the peaks identified by SELDI profiling that had the ability to distinguish between patient groups, the 2 most discriminating protein peaks could differentiate patients with pancreatic cancer from healthy controls with a sensitivity of 78% and specificity of 97%. These 2 markers performed significantly better than the current standard serum marker, CA19-9 (P < 0.05). The diagnostic accuracy of the 2 markers was improved by using them in combination with CA 19-9. Similarly, a combination of 3 SELDI markers and CA19-9 was superior to CA19-9 alone in distinguishing individuals with pancreatic cancer from the combined pancreatic disease controls and healthy subject groups (P = 0.078). SELDI markers were also better than CA19-9 in distinguishing patients with pancreatic cancer from those with pancreatitis. SELDI profiling of serum can be used to accurately differentiate patients with pancreatic cancer from those with other pancreatic diseases and from healthy controls.

Differential protein profile in the ear-punched tissue of regeneration and non-regeneration strains of mice: a novel approach to explore the candidate genes for soft-tissue regeneration

Wound repair/regeneration is a genetically controlled, complex process. In order to identify candidate genes regulating fast wound repair/regeneration in soft-tissue, the temporal protein profile of the soft-tissue healing process was analyzed in the ear-punched tissue of regeneration strain MRL/MpJ-Fas lpr (MRL) mice and non-regeneration strain C57BL/6J(B6) mice using surface-enhanced laser desorption and ionization (SELDI) ProteinChip technology. Five candidate proteins were identified in which responses of MRL to the ear punch were 2–4-fold different compared to that of B6. Their corresponding genes were predicted using an antigen–antibody assay validated mass-based approach. Most of the predicted genes are known to play a role or are likely to play a role in the wound repair/regeneration. Of the five candidate proteins, the amount of the 23 560 Da protein in the ear-punched tissue was significantly correlated with the rate of ear healing in six representative strains of mice, making it a good candidate for fast wound repair/regeneration. We speculate that the increased concentration of the 23 560 Da protein in the wound tissue could stimulate the expression of various growth-promoting proteins and consequently speed up the wound repair/regeneration processes. Here, we have shown that examination of protein expression profile using SELDI technology, coupled with database search, is an alternative approach to search for candidate genes for wound repair/regeneration. This novel approach can be implemented in a variety of biological applications.