What LC mode do you use? (answer all that applies)

Mostly Reversed Phase mode

Ion Exchange Chromatography

Mostly Normal Phase mode

Size Exclusion Chtomatography (SEC or GPC)

Chiral chromatography

Affinity Chromatography

Hydropilic Interaction Chromatography (HILIC)

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Do you use U-HPLC (UPLC) in your daily work

Yes, I use mostly U-HPLC (UPLC)

Yes, but only here and there

Yes, very rarely, I still prefer to use HPLC

No, but I would like to use it if I have access to

No, I think it is just a hype and have no future

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General Information & Resources	Articles & Reviews	MS Technology Tutorials and Overviews	API ionization links
BIO Tools	Utilities and General Background material	Waters Website	Old Micromass' Support
UPLC	Special Applications	Clinical MS/MS	HPLC Operation
Sample preparation	Metabonomics	Method Development	Gel Electrophoresis

GENERAL INFORMATION & RESOURCES אינפורמציה כללית ומשאבים

 

    Get the laboratory ready for liquid chromatography mass spectrometry work

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Articles, Posters & Reviews מאמרים,  פוסטרים וסקירות

• Troubleshooting LC-MS problems 5 mb (register first)
• Principles of MS quantitation
• Principles of Quadrupole and ion Trap
• LC-MS primer
  

 

Atmospheric Pressure Ionization links יוניזציה בלחץ אטמוספרי

 

• Charge state and mass assignment

• Determining the Charge States of Multiply-Charged Ions

• Example of mass spectra at various ionization modes

• Electrospray ionization - lecture (pdf)

MS Technology Tutorials & Overviews מדריכים וסקירות על הטכנולוגיה

 

• Reversed Phase: (Very good for beginners)
• Handouts of my LC-MS Lecture at Forum Scientific (My courses' attendees: ask for the password)
• Introduction to Mass Spectrometry at Scripps Center for Mass Spectrometry
• Introduction to Mass Spectrometry 
• Introduction to Ion Trap Mass Spectrometer
•  Introduction to Quadrupole Time-of-Life
• Basics of LC-ESI-MS-MS of Peptides
• Theoretical Review of Mass Spectroscopic Techniques - Cambridge University
• The Chemistry Hypermedia Project: MS Tutorial
• GC-MS and MS Course with test
  

 

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Proteomic Tutorials and lectures מדריכים בפרוטאומיקה

• Peptide Sequencing with Electrospray LC/MS Part 1: Ion Types and Nomenclature by Frederick Klink 
• Labeled and Label Free Proteomics 
  
• Introduction to Bioinformatics - by Suresh Kumar
  
• What is bioinformatics? 
  
• MASCOT Searches - Help Topic Index and background
  
• Peptide fragmentation
  
• Integration with the human genome of peptide sequences obtained by high-throughput mass
• spectrometry 
  
• Introduction to Proteomics Part 1  by B. Tilg, A. Graber und K. Weinberger 
  
• Amino Acid Structures
  
• Amino Acids chemistry

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BIOINFORMATICS Tools כלים בביואינפורמטיקה

• SUMMARY of links on searching Bio databases
• MASCOT searches 
• Search for peptides and proteins  in ExPASy
• Swiss Prot general search
• BLAST NCBI  Search for peptides and protein IDHELP PAGE FOR BLAST
• GlycoMod - Oligosaccharide structures from mass spectroscopy
• GlycanMass - Oligosaccharide mass from structure
• FindMod - Post-translational modifications by peptide mass
• Pacific Nothwest National Laboratories USA: Proteomics Software Tools

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Utilities & Background Material חומר שימושי

• Help Desk of Mass Spectrometry
  
• Download Spectra of Pesticides
  
• Search for Delta Mass - A Database of Protein Post Translational Modifications
  
• ChemCalc- calculate MW 
  
• Fragment Finder 
  
• AA to Peptide MW calculator
  
• Periodic Table utility
  
• Protein characterization and identification tools - Summary
  
• Common Postranslational Modifications masses
  
• Post translational modifications masses
  
• More Delta masses of modifications
  
• EMBL

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Mass Spectroscopy - Sample Preparation הכנת דוגמאות

 

• General Guidelines for Preparing Samples for MS Analysis - UTexes
  Sample preparation references for MALDI
  Sample preparation of Proteins for MS (PharmaGenomics November/December 2003)
  

 

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2D GEL ELECTROPHORESIS LINKS קישורים לאלקטרופורזת ג'ל

• 2D GEL ELECTROPHORESIS FOR PROTEOMICS-TUTORIAL  Arjan van Rossum A Basic Primer to Protein by Roger Biringer

Metabonomics Tutorial and Tools

Chemometrics of metabonomics

LC-MS Method Development פיתוח שיטות

• Eliminate TFA and Improve Sensitivity of Peptide Analyses by LC/MS
  
• Narrow bore, micro and nano LC columns in LC-MS- J. MS
  
• Determination of Carryover and Contamination for Mass Spectrometry – Based Chromatographic
• Assays  
  
• Bioanalytical Method development of Therapeutic Peptides   
  
• FDA Guidance to the industry on validation of bioanalytical methods 
  
• Key Elements of Bioanalytical Method Validation for Small Molecules
• Mass Spectrometry Adduct Calculator 
• "Ghost" peaks: Background contamination
  

Special Applications יישומים מיוחדים

Clinical  LC-MS/MS - for Newborn disorders יישום רפואי

• Tandem Mass Spectrometry in Newborn Screening A Primer for Neonatal and Perinatal Nurses 
  
• Use of  Tandem Mass Spectrometry  Donald H. Chace,* Theodore A. Kalas, and Edwin W. Naylor 
  
• A Primer on Newborn Metabolic Screening
  
• Expanded Newborn Screening for Inborn Errors of Metabolism by Electrospray Ionization
  
• Screening for Inborn (MS/MS) Errors of Metabolism 
  
• The Expanding Role of Mass Spectrometry in Metabolite Profiling and Characterization
      Elizabeth J. Want, Benjamin F. Cravatt, and Gary Siuzdak
  
• Newborn screening - CDC
  
• Utah Newborn Screening Training program  
  
• DIAGNOSIS OF INBORN ERRORS OF METABOLISM BY TANDEM MASS SPECTROMETRY Pinar T. Ozand
  
• New York-guide for health professionals 
  
• ANALYSIS OF CARNITINE AND ACYLCARNITINES IN BIOLOGICAL FLUIDS AND APPLICATION TO A CLINICAL STUDY 
  
• Neonatal screening  
  
• Neonatal screening for inborn errors of metabolism
  
• New developments in neonatal screening 
  
  Reagents for  Acylcarnitines and Amino acids analysis in blood spots ריאגנטים עבור אנליזה לתינוקות
  
  
• Bulanolic HCl - Regis Technologies
  
• Cambridge Isotopes Laboratory
  
• Materials and Methods for  acylcarnitine analysis by MS/MS
  
• Metabolic Laboratory: Standards for metabolic research
  
• Experimental: Article-Clin Chem  51:4 745–752 (2005)   
• Protocol (ppt)
  

Introduction

The liquid chromatograph is very complex and it consists of sophisticated technology and sometimes delicate and total ignorance in the manner it works might sabotage its parts which are very expensive.  In addition, there are chemical processes of distribution between the two phases, the mobile and stationary, while the analysis goes on, and there is a need for a minimum understanding of their theoretical basis to operate the system correctly.  Also, a minimum understanding of the method, in which the system performs data analysis, is needed so that no false data or results will be obtained in the analysis.

LISTS OF STEPS NEEDED BEFORE ANY RUN BY HPLC  רשימת צעדים מקדימים לפני הרצה :

 

• Filter the solvents with membranes with cutoff of 0.22-0.45 mM
• Use clean and transparent reservoirs through which precipitates and colloids can be distinguished.
• Make sure that the solvents will be easily mixed with the previous solvents in the same inlets (A, B, C, D).  For example methanol or water should not be placed instead of hexane directly, or any organic solvent should not be placed directly instead of a buffer reservoir.
• Degass the solvents and purge all the tubing that lead to the pump.
• Connect the column carefully according to the flow direction marked on it (do not connect directly to the detector).
• Flow the appropriate solvents through the column at a low flow-rate (0.1-0.5 ml/min) or reach the composition gradually using the appropriate gradient.
•  Select the appropriate wavelength (or other type of setting) in the detector and wait for stable baseline
• Prepare the set of methods in the workstation: Instrument method for the control on the system, Processing method for the data processing and the Report method for the report of final results.
• When the system and the methods are ready a blank run should be performed to test the system and verify that it is clean from interferences.

Solvent delivery system המשאבה

The column הקולונה

Detector  הגלאי

Injector   המזרק

Gradient and isocratic  גרדיינט או איזוקרטי

Data Processing  עיבוד נתונים

Copyright (C)1997 Dr. Shulamit Levin