Day 3 :
- Track 9: Clinical chemistry, Biomarkers and Diagnostics assays
Track 10: Complementary separation techniques
Track 11: Green Technologies, Future Challenges and Trends
Session Introduction
Juan G. Santiago
Stanford University, USA
Title: Life in the shock wave: Accelerating DNA reactions with isotachophoresis
Biography:
Juan Santiago received his MS and PhD in Mechanical Engineering from the University of Illinois at Urbana-Champaign in 1992 and 1995. His research includes the development of microsystems for on-chip chemical analysis, drug delivery, sample preparation methods, and desalination of water. Applications of this work include molecular medical diagnostics, drug discovery, environmental monitoring, and the production of drinking water. He is a Fellow of the American Physical Society, a Fellow of the American Society of Mechanical Engineering, an Associate Editor of the journal Microfluidics and Nanofluidics, co-founder of several companies in the microfluidics area, co-inventor of micron-resolution particle image velocimetry, and director of the Stanford Microfluidics Laboratory. He served as Associate Editor of Lab on a Chip '08-'13. He has given 26 keynote and named lectures and more than 100 additional invited lectures. His work is cited about 1000 times per year. He has graduated 26 PhD students and advised eight postdoctoral researchers. 16 of his former advisees are now professors at major universities. He has authored and co-authored over 150 archival publications and 200 conference papers, and holds 36 patents (16 of which are licensed).
Abstract:
We use isotachophoresis (ITP) to achieve fast and specific analyses of molecular targets in complex mixtures. We use ITP to pre-concentrate and purify; molecular recognition for specificity; and ITP to control and increase the rate of chemical reactions between molecular probes and target macromolecules. ITP is an electrophoresis technique that uses two buffers which include a high mobility leading electrolyte (LE) and a low mobility trailing electrolyte (TE). Sample species with mobilities bracketed by those of the LE and TE focus into the TE-to-LE interface. For trace sample concentrations, multiple species focus in so-called peak mode wherein multiple analytes mix and strongly overlap within an order of 10 µm wide ionic concentration shock wave. This co-focusing mixes target species and pre-concentrates them to accelerate reactions. We have integrated DNA and RNA extraction with sequence-specific quantitation using a variety of mobile and immobile cDNA probes. We pre-concentrate target and probe molecules by >10,000x and achieve in 30 sec reactions which would normally take 4 days. We have shown specific and sensitive detection of target sequences in order 5 minutes with little or no off-chip sample preparation, and without target amplification.
Takayuki Kawai
RIKEN, Japan
Title: Ultra-sensitive capillary electrophoresis for single cell analysis
Biography:
Takayuki Kawai received MSc and PhD in School of Engineering, Kyoto University, for his research on highly sensitive micro-scale electrophoresis. He then became a Postdoc in AIST and University of Illinois to study micro-fluidics and capillary electrophoresis for high-performance bio-analytical chemistry. In 2014, he was promoted to Research Scientist in RIKEN where he started the current research on integrated single-cell analysis. He also has the position of a PRESTO Researcher in Japan Science and Technology Agency, which is awarded to very few outstanding young scientists in Japan. He has published 13 qualified papers to date.
Abstract:
Recently, single-cell analysis is becoming more and more important to understand cell-to-cell heterogeneity in the complicated life system. Although large molecules within the single cell can easily be visualized via immune-histochemical staining, simultaneous analysis of numerous compounds still remains one of the most challenging issues. Among several analytical techniques, Capillary Electrophoresis (CE) coupled to Laser-Induced Fluorescence (LIF) or Mass Spectrometer (MS) is an effective method due to its high separation efficiency, low sample requirements, and high detectability. In this presentation, ultra-sensitive CE-LIF/MS system is introduced that uses online sample pre-concentration techniques like Large-Volume Sample Stacking (LVSS) for the single cell analysis. So far, pM detectability has been achieved in the analysis of biogenic compounds such as amino acids and oligosaccharides. Successful demonstration of LVSS-CE-LIF chiral analysis of a single cell will finally be introduced.
Stanislav L. Karsten
NeuroInDx, Inc., USA
Title: KuiqpicKTM and UnipicKTM: versatile instruments for single cell acquisition and complex heterogeneous tissue microdissection
Time : 09:30-09:50
Biography:
Dr. Karsten has completed his PhD in Medical Genetics and Pathology from Uppsala University, Sweden in 2000 and postdoctoral studies in Functional Neurogenomics from UCLA in 2004. Currently, Dr. Karsten is a President and CEO of NeuroInDx, Inc., a California based company focused on the development of instrumentation for cell and tissue specific sample acquisition and analysis in biomedical research. Prior to his position at NeuroInDx, Dr. Karsten was an Assistant Professor of Neurology where his laboratory conducted research in neuroscience. Dr. Karsten’s research work was published in over 30 peer-reviewed journals including Nature, Neuron and PNAS and resulted in several patents
Abstract:
Cell specific studies are imperative for sound research in molecular biology and biomedical research. Tissue heterogeneity (e.g., brain) poses significant challenge for retrieval of cell and region specific information. Moreover, recent progress in single cell research creates strong demand for rapid acquisition of individual cells from both tissues cell cultures. However, existing approaches of single cell acquisition, such as flow sorting and laser assisted microdissection are costly, sophisticated and often methodologically limited. We have developed a line of versatile instruments (e.g., KuiqpicKTM, UnipicKTM) which are compatible with any inverted microscope and have a wide range of cell and tissue acquisition parameters. Both instruments permit rapid collection of individual cells from various cell cultures and tissues for a range of downstream applications including function single cell studies, high quality protein, RNA and DNA isolation and sequence analysis. Collection of cells may be performed directly from native tissues and cell cultures with low impact on cellular viability. Native, fresh frozen, and sucrose treated tissues could be used for microdissection. Here, the recent progress in the instrument’s automation is presented further widening potential areas of application
Asokan C
Sokoto State University. Nigeria
Title: Role Of α-B Crystallin On Serum Amyloid A Fibrils And Effect Of β-Amyloid With Systemic Amyloidotic Mice Brain
Biography:
DR. Asokan C has completed his PhD at the age of 27 years from University of Madras and postdoctoral studies from Columbia University. NY. USA. He is the Associate Professor, Department of Biochemistry, Sokoto State University, Sokoto. Nigeria. He has published more than 36 papers in reputed journals and has been serving as an editorial board member of repute
Abstract:
Enhanced expression of amyloid β-peptide (Aβ) and deposition is the main causative factor in Alzheimer’s disease (AD). Factors that lead to the genesis of accumulation and toxicity of Aβs are yet to be identified. While studying the effect of systemic amyloid on the functions of the mice brain, it was accidentally found that the mice brains contain accumulated Aβs, which are extractable with hexafluroisopropanol (HFIP) solvent. By purifying with semi preparative HPLC on HFIP extracts, two major fractions containing mixture of Aβs with variable composition were observed. We have characterized these mixtures by electron microscopic and spectroscopic methods. Our results indicate that, the accumulated Aβ fibrils have similar morphological and conformational characteristics as that of Aβs of AD brains.
Peter Mikuš
University in Bratislava, Slovakia
Title: Two-dimensional capillary electrophoresis separation and on-line tandem mass spectrometry detection of antigripal drugs and their metabolites in urine matrices
Biography:
Peter Mikuš has completed his PhD at the age of 30 years from Comenius University (Slovakia). He is Researcher, University Teacher, Associated Professor, and Director of the Toxicological and Antidoping Center at the Faculty of Pharmacy Comenius University in Bratislava (FPCU) as well as Head of the Department of Pharmaceutical Analysis and Nuclear Pharmacy FPCU. A research team of P.M. is focused on the development, validation and application of advanced hyphenated analytical methods, based on a combination of 2D-separation and spectral (UV-VIS, MS/MS) techniques, for pharmaceutical and biomedical research. He has published more than 60 papers in reputed CC journals.
Abstract:
The advanced two dimensional isotachophoresis (ITP) – capillary zone electrophoresis (CZE) hyphenated with tandem mass spectrometry (MS/MS, here triple quadrupole, QqQ) was developed in the present work to demonstrate analytical potentialities of this approach in the analysis of antigripal drugs in multicomponent ionic matrices. This was demonstrated on the determination of pg.mL-1 levels of pheniramine, its desmethyl metabolite and phenylephrine in directly injected (unpretreated) urine samples, suitable for the monitoring of concentration vs. time dependences of these species. The main benefits of ITP-CZE-ESI-QqQ in comparison with CZE-ESI-QqQ are considerably higher sensitivity / lower LODs obtainable and more favorable parameters of precision and recovery/accuracy when performing analyzes at lower concentration levels. The separation selectivity of ITP-CZE-ESI-QqQ towards the targeted analytes is enhanced via the heart cut effect while CZE-ESI-QqQ has the advantage of utilization of the EOF effect for the monitoring additional (e.g. neutral) species related to the targeted analytes. Anyway, the on-line ITP sample preparation provides, besides pre-concentration and separation selectivity, also the maximum reduction of ionic sample matrix constituents entering MS. In this way, MS can be effectively protected (long-term working life is ensured), potential detection interferences are minimized and signal to noise ratio of the detection response is maximized. Obviously, ITP-CZE-ESI-QqQ is superior than ITP-CZE-UV regarding sensitivity/lower LOD, selectivity and identification reliability but disadvantageous regarding cost of instrumentation/analysis and the scale (number) of CE electrolytes usable for MS. The ITP-CZE-ESI-QqQ method has a great potential to be routinely applied for monitoring various targeted ultra-trace ionic drugs in biomedical/clinical laboratories.
Mercy Ekwere
Department of Biochemistry, Nigeria
Title: Comparative sex hormonal indices of male and female rats exposed to uppercott (mixture of organophosphate/ pyrethroid) pesticide.
Biography:
The effect of oral administration of a mixture of cypermethrin and dimethoate (organophosphate and pyrethroid groups of pesticide, commonly called uppercott) on serum sex hormonal profile of male and female rats was assessed. 10 mg/kg and 20 mg/kg body weight of uppercott were given to separated male (M) and female (F) rats grouped as F2, M2 andF3, M3 respectively. Rats in groups F1, M1 served as control. After 28days, the animals were sacrificed; sera collected and assayed for follicle stimulating hormones (FSH), Luteinizing hormones (LH), testosterone, estradiol and progesterone. Results showed that all serum sex hormones, for both male and female rats, were significantly altered as doses increased from control via 10 mg/kg to 20 mg/kg. FSH and LH were significantly reduced (p<0.05) in both male and female rats (M1FSH, M2FSH and M3FSH were 3.36±0.32 mIU/ml, 2.32±0.31mIU/ml and 1.9±0.15mIU/ml and F1FSH-3.32±015, F2FSH- 2.48±0.17 and F3FSH, 1.96±0.13. Estradiol and progesterone in the female rats; and testosterone for male rats decreased significantly (p<0.05). Toxic effects of uppercott on testosterone and progesterone male and female rats respectively, were not significantly different (p>0.05) from each other. Uppercott significantly reduced all sex hormonal levels especially at higher dose. Constant exposure to high concentration of organophosphate and pyrethroid pesticides could have toxic effects on male and female fertility and reduce reproductive integrity. A limit should be placed on the use of such toxic pesticides/insecticides.
Abstract:
The effect of oral administration of a mixture of cypermethrin and dimethoate (organophosphate and pyrethroid groups of pesticide, commonly called uppercott) on serum sex hormonal profile of male and female rats were assessed. 10mg/kg and 20mg/kg body weight of uppercott were given to separated male (M) and female (F) rats grouped as F2, M2 and F3, M3 respectively. Rats in groups F1, M1 served as control. After 28days, the animals were sacrificed; sera collected and assayed for follicle stimulating hormones (FSH), Luteinizing hormones (LH), testosterone, estradiol and progesterone. Results showed that all serum sex hormones, for both male and female rats, were significantly altered as doses increased from control via 10mg/kg to 20mg/kg. FSH and LH were significantly reduced (p<0.05) in both male and female rats (M1FSH, M2FSH and M3FSH were 3.36±0.32mIU/ml, 2.32±0.31mIU/ml and 1.9± 0.15mIU/ml and F1FSH-3.32±015, F2FSH- 2.48±0.17 and F3FSH, 1.96±0.13. Estradiol and progesterone in the female rats; and testosterone for male rats decreased significantly (p<0.05). Toxic effects of uppercott on testosterone and progesterone male and female rats respectively, were not significantly different (p>0.05) from each other. Uppercott significantly reduced all sex hormonal levels especially at higher dose. Constant exposure to high concentration of organophosphate and pyrethroid pesticides could have toxic effects on male and female fertility and reduce reproductive intergrity. A limit should be placed on the use of such toxic pesticides/insecticides.
Asokan C
Department of Biochemistry, Nigeria
Title: Characteristics of Fibrillar form of β-Amyloid Peptide Fractions from Mice Brain affected by Systemic Amyloidosis.
Biography:
DR. Asokan C has completed his PhD at the age of 27 years from University of Madras and postdoctoral studies from Columbia University. NY. USA. He is the Associate Professor, Department of Biochemistry, Sokoto State University, Sokoto. Nigeria. He has published more than 36 papers in reputed journals and has been serving as an editorial board member of repute.
Abstract:
Enhanced expression of amyloid β-peptide (Aβ) and deposition is the main causative factor in Alzheimer’s disease (AD). Factors that lead to the genesis of accumulation and toxicity of Aβs are yet to be identified. While studying the effect of systemic amyloid on the functions of the mice brain, it was accidentally found that the mice brains contain accumulated Aβs, which are extractable with hexafluroisopropanol (HFIP) solvent. By purifying with semi preparative HPLC on HFIP extracts, two major fractions containing mixture of Aβs with variable composition were observed. We have characterized these mixtures by electron microscopic and spectroscopic methods. Our results indicate that, the accumulated Aβ fibrils have similar morphological and conformational characteristics as that of Aβs of AD brains.
Radim Vespalec
Academy of Sciences of the Czech Republic,Czech Republic
Title: Analytical task stemming from therapeutical prospects of electron deficient boron cluster compounds
Biography:
Radim Vespalec has received equivalent to PhD from the Institute of Physical Chemistry, Academy of Sciences of the Czechoslovak Republic, Praha, in 27. The scientific degree Dr.Sc. he received from Technical University of Pardubice in 57, the pedagogical Asoc. Prof. degree he received from Masaryk University Brno in 58. He is senior scientist in the Institute of Biophysics. Web of Science reports his 80 scientific articles He contributed to 3 monographs.
Abstract:
The existence, reactions, structures and properties of compounds occurring in nature, and their synthetic analogues are explainable by the idea of two-center two-electron bond. These families contain either electron exact or electron rich building blocks from the viewpoint of electron structure. Electron deficient building blocks have never been found in nature, and exist only in synthetic species. Boron cluster compounds (BCCs) create the most intensely investigated family of species with electron deficient cluster. Their existence has been explained by the accumulation of unique electron deficient bonds, which bind together three boron atoms or, sometimes, their substitutes, in clusters. Pronouncedly electron deficient clusters either determine or substantially affect properties of BCCs, and their prospects. Therapeutical prospects attract the highest attention now, and many compounds with boron clusters are synthesized as candidates for therapeutical uses. These compounds must pass through mandatory studies and checks, which require variety of chemical analyses, identically with other compounds. However, analytical methods do not exist for analyses of compounds with boron clusters. The pieces of knowledge from chiral separation of BCCs prove the dissimilarity of some analytical properties of species with and without boron clusters, and indicate the absence of criteria for the a priori estimation of different analytical properties for compounds with and without clusters. Thus, missing analytical methods cannot be derived from existing knowledge. Analytical research of BCCs motivated by their medical prospects is the best way to preventive elimination some obstacles, which may hamper medical uses of compounds with boron clusters.
Doo Soo Chung
Seoul National University, Korea
Title: Sensitive Speciation of Arsenic Compounds with Capillary Electrophoresis
Biography:
Doo Soo Chung has completed his PhD from Harvard University and postdoctoral studies from MIT and Iowa State University. He has published more than 100 papers in reputed journals.
Abstract:
Arsenic is naturally abundant in the crust of the earth and introduced into the aquatic system through dissolution and weathering of minerals. Chronic ingestion of arsenic in water may cause various diseases, including cancer and keratosis. A guideline for arsenic in drinking water has been set at 10 ppb of total arsenic by the World Health Organization (WHO). However, inorganic forms of arsenic, such as arsenites [As (III)] and arsenates [As (V)], are much more toxic than the organic forms as the mono methylarsonic acid (MMA) and dimethylarsinic acid (DMA). Hence the quantitation of specific arsenic species may be more meaningful than the total arsenic determination for the evaluation of the health risks from arsenic-contaminated drinking water. We report a highly sensitive way of arsenic speciation using a commercial Capillary Electrophoresis (CE) instrument equipped with a UV absorbance detector. We used a counter-flow electrokinetic supercharging technique to enhance the detection sensitivity. Electrokinetic supercharging is one of the most powerful sample stacking methods that combine field amplified sample injection and transient isotachophoresis. In counter flow electrokinetic supercharging, a constant counter pressure is applied during sample injection in order to counterbalance the movement of the injected sample zone, obtaining a pronounced increase in the amount of sample injected and the portion of the capillary available for electrophoresis.
Fabiano Freire Costa
Federal University of Juiz de Fora, Brazil
Title: Microfluidic Chip Electrophoresis Investigation of Major Milk Proteins: Study of Buffers Effects and Quantitative Approaches
Biography:
Dr. Fabiano completed his post-doctorate in advanced milk chemistry with the theoretical and practical combination of microfluidic chip electrophoresis, scanning electron microscopy, transmission electron microscopy, differential scanning calorimeter and static light scattering at the Embrapa Dairy Cattle National Research, Brazil in 2013. He joined at the Federal University of Juiz de Fora/Governador Valadares, Minas Gerais, Brazil in 2013 as a Professor. This led to the development of new methodology to analyze milk. Dr. Fabiano is member of the International Dairy Federation acting in the working groups: Analytical Methods for Additives & Contaminants (SCAMAC); Analytical Methods for Composition (SCAMC); Food Additives (SCFA) and Inventory of Methods and Procedures to Monitor the Integrity Supplier's Milk (TF-ISM-01).
Abstract:
The separation and quantification of major milk proteins are fundamental in dairy research. Therefore, accurate and rapid methods are profoundly important. The microfluidic chip technique is faster, and uses considerably fewer chemicals and materials than traditional techniques. The objective of this study was to improve experimental methods for separating and quantifying major milk proteins using the microfluidic chip technique. Deionized water, a total protein solubilization buffer (TPS buffer) and a separating milk protein buffer (SEP buffer) were added to the treatment of milk samples and their effects were evaluated. The results showed an excellent separation of whey proteins with α-lactalbumin migrating first, followed by β-lactoglobulin in the presence of both buffers. However, better results for major case in separation were achieved when the SEP buffer was added. The order of the migration time was: β-casein first, followed by αs-casein and κ-casein. The quantitative analysis showed significant differences among the percentages of protein fractions from both buffers. The results using microfluidic chip technology using the SEP buffer solution were comparable to those obtained by SDS-PAGE for these proteins and with the data reported in the literature.
Z. Mansoori
Amirkabir University of Technology, Iran
Title: Numerical investigation of gas- solid separation by liquid layer in horizontal pipes
Biography:
Zohreh Mansoori completed her PhD in Mechanical Engineering from Amirkabir University of Technology (Polythechnic), Tehran, Iran. She is associate professor and the head of Energy Research center in Amirkabir University of Technology. She has published more than 35 papers in reputed journals and has been serving as an editorial board member of repute.
Abstract:
Gas- solid separation is studied considering a liquid layer in the lower part of horizontal pipe. The particles falling in liquid layer could not go back in gas flow. The presence of solid particles on stratified wavy gas- liquid flows has been studied. In present paper, the numerical four- way simulation of solid particles in gas- liquid wavy stratified flow has been used. The computational model is shown to be able to evaluate the effect of the liquid layer on particles separation.