BS313 T1 Maldi Supp Reading - UV MALDI-Evolution and Principle

8/4/2019 BS313 T1 Maldi Supp Reading - UV MALDI-Evolution and Principle

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ORGANIC MASS SPECTROMETRY, VOL. 27, 653-659 (1992)ACCOUNTSMatrix-assisted Ultraviolet Laser Desorption:Evolution and PrinciplesRonald C. BeavisDepartment of Physics, Memorial University of Newfound land, St. Johns, Newfoundland A1B 3x7,Canada

INTRODUCTIONMy first introduction to laser desorption was when Ijoined the Institute of Physical and Theoretical Chem-istry at the Technical University of Munich as a NATOPostdoctoral Fellow in August 1987. I began workingon the problems associated with the use of infrared laserdesorption as a source of gas-phase macromolecules forentrainment by a supersonic jet. I had just finished mydoctoral degree with Ken Standing at the University ofManitoba, working on problems associated with ana-lyzing peptides by static secondary ion mass spectrom-etry (static SIMS) using time-of-flight massspectrometers. Soon after beginning this work, I had theopportunity to attend the Fourth Ion Formation fromOrganic Solids (IFOS IV) conference held at the Uni-versity of Munster, September 21-23, 1987. This con-ference had been organized on a roughly biennial basisby Alfred Benninghoven, the discoverer of organic staticSIMS. The conference was a meeting place for peoplewho were interested in the problem of producing intact,gas-phase ions from organic molecules that do not havean equilibrium partial pressure at any temperature. Themost memorable part of the conference was one sessionwhen Professor Benninghoven broke into the pro-ceedings to announce some results that he had justreceived from a group in Japan. Dr Koichi Tanaka ofShimadzu had sent a reprint of a conference pro-ceedings abstract to Professor Benninghoven showingthe mass spectra of proteins with high molecular masses(2nd Japan-China Joint Symposium on Mass Spec-trometry, September 15-18, 1987). These spectra hadbetter signal-to-noise ratios than could be obtained byplasma desorption mass spectrometry and the ionsignals were very broad. The ion source used wasmysterious: the details of the method of ion productionwere not described and did not come out until muchlater. What was known about the ion source was that itused a pulsed ultraviolet (UV) laser and a new methodof sample preparation. The sample preparation methodwas described as the mixture of the protein with a liquidmatrix and a finely divided metal powder of some type.Discussion of these results dominated private conversa-tions for the rest of the conference, along with specula-tions about the role of the metal powder.Also attending the conference were Franz Hill-enkamp and Micheal Karas. Professor Hillenkamp had

recently moved his group from Frankfurt to Munsterand his group was establishing their laboratory at thetime of the conference. This group had been studyinglaser desorption using pulsed UV lasers and had devel-oped the laser microprobe mass analysis (LAMMA)instrument. They had been pursuing the idea of using amatrix to enhance the yields of high-mass molecularions for some time, although these investigations werenot discussed at the IFOS IV conference.After the conference, there was no further word onthe discovery of Dr Tanaka. There was also no word onthe progress being made by Professor Hillenkampsgroup. This silence was broken at the InternationalMass Spectrometry conference held at Bordeaux,France, in August, 1988. I was present at the sessionwhen Franz Hillenkamp announced their excitingresu1ts.j Using a solid, UV-absorbing matrix (nicotinicacid), they had managed to produce large signals ofprotein ions by irradiating the surface with a short pulseof 266 nm laser radiation. The molecular mass of theprotein did not seem a limitation. Equally exciting wasthe sensitivity that this new technique promised. Thetotal amount of protein loaded into the mass spectrom-eter was roughly 1 pmole. The name given to this newmethod of ion production was matrix-assisted laserdesorption. This term was meant to indicate that thisnew technique was a special case of the old method ofordinary laser desorption. The announcement was fol-lowed closely by the publication of these results inseveral journals. Interestingly, the results of Tanaka etal. were published in an internationally availablejournal4 almost simultaneously with those of theMunster group.Following the announcement in Bordeaux, I wasoffered a position at Rockefeller University in the labor-atory of Brian Chait to investigate matrix-assisted laserdesorption as a possible analytical technique for pro-teins important for biomedical research. I accepted theposition, but before leaving Germany I spent 3 days atMunster, with Micheal Karas and Franz Hillenkamp.They gave me demonstrations of the method and gaveme whatever guidance they could with regard to whatthey felt were the important experimental parameters.Taking this information with me to New York, BrianChait and I set up a time-of-flight mass spectrometer atRockefeller that provided the first reproduction ofKaras and Hillenkamps discovery in another labor-atory.6 Several other laboratories also announced con-

0030-493X/92/060653-07 $08.500 992 by John Wiley & Sons, Ltd.


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654 R. C. BEAVISfirmation of the matrix-assisted laser desorption effectand they have added substantially to the current under-standin g and a pplication of the technique..*SAMPLE PREPARATION :TANAKA AND HILLENKAMPThe attempt to produce gas-phase ions from involatileorganic molecules has been dominated by the develop-ment of new sample preparation methods. The primacyof sample preparation has not been generally recog-nized in nomenclature: most techniques are named afterand classified by the method of energy deposition. Thediscovery of plasma desorption mass spectrometry byMacfarlane and Torgesson9 was turned into a practicalanalytical technique by using nitrocellulose membranesas sample substrates. Static SIM S was made viable byetched silver substrates. Dynam ic SIMS , rebor n asfast atom bombardment (FAB), became useful forfragile organic molecules with the introdu ction of liquidmatrices. In their first paper on FAB, Barber et a/.stated that to produce long-lasting ion signals it isnecessary to make judicious use of solvent and supportsystems, paying particular attention to the viscosity andvolatility of the medium from which the sample isdeposited on the stage. It was not until the followingyear that this statement was clarified to mean the use ofa glycerol rnatrix.j Laser desorption has also gonethrough many different types of sample preparation, butuntil 1987 it was not rega rded by the analytical com-munity as a viable technique for examining high molec-ular mass compounds.The discovery of Tanaka et aI? changed the percep-tion that laser desorption had an upper mass limit of- 000 u. The mode of energy deposition was the sameas that used by many investigators: a pulsed ultravioletlaser (a nitrogen gas laser, A = 337 nm). The samplepreparation method used was the key to the newresults. It involved dissolving the polypeptide of interestin glycerol and mixing the glycerol solution with afinely divided metal powder. A dr op of this suspensionwas placed on a probe tip and the products of laserirradiation were analyzed with a reflectron mass spec-trometer. The use of a low vapor pressure matrixmaterial was not new: i t had already been used indynamic SIMS (FAB). The glycerol was totally trans-parent to 337 nm radiation. The laser light passedthrough the glycerol without any interaction with it. Toproduce an effect, it was necessary to introduce absorb-ing centres into the solution, hence the use of the m etalpowder. A metal particle that has dimensions smallertha n the wavelength of the irrad iating laser is heated bycurrents induced by the rapidly varying (and spatiallycoherent) electric field of the laser light. Because of theirability to produce thermal excitation in response tolaser irradiation, the metal particles are the couplingbetween the light and the liquid m atrix. Th e mechanismthat connects the heating of the metal particles with theejection of intact protein ions has n ot been established.The method of sample preparation chosen by Karasand Hillenkamp was conceptually different. Their longexperience with laser desorption led them to believethat often it was the substrate under a layer of organic

material on a surface that absorbed the laser light, notthe organic molecules themselves. In 1985, Hillenkampand co-workers began to publish paper^'^.'^ containingthe hypothesis that large molecule ions could be pro-duced by mixing the analyte with a matrix material thatwas chosen for its ability to absorb the laser light. Thismatrix material would absorb the light, resulting in itsablation and hopefully to a coupled ablation of analytemolecule ions. Th e intimate role of the m atrix in drivingthe desorption process led to the name proposed:matrix-assisted laser desorption. A variety of matriceswere tried, including tryptophan, nicotinic acid, 3-nitrobenzyl alcohol and 2-nitrophenyl octyl ether, andthe results published were promising, but not spectacu-lar.The breakthrough for matrix-assisted laser desorp-tion as applied to polypeptides came with the discoverytha t nicotinic acid had special properties for large poly-p e p t i d e a n a l ~ t e s . ~ , ~he laser being used was thefourth harmonic of a Q-switched, neodymium-doped,yttrium aluminium garnet laser [Nd: YAG (4)], with awavelength of 266 nm. Nicotinic acid is a solid at roomtemperature, with a low vapor pressure (but a tendencyto sublime) and a high absor ption coefficient at 266 nm.When aqueous solutions of a protein and nicotinic acidwere mixed and a droplet of this mixture was dried, thedeposit formed had the properties they had sought.Large signals of protein molecule ions were observed ina reflectron time-of-flight mass spectrometer when thesample was irradiated.The major difference between the results of theTanaka method of sample preparation and theHillenkampKaras method was sensitivity. Tanakasmethod required nanomoles of protein to prepare asample for analysis. The HillenkampKaras methodrequired picomoles of protein, i.e. it was 1000 timesmore sensitive. The signals obtained from the lattermethod were also more intense and had a higher signal-to-noise ratio. These factors indicated that for an ana-lytical method, the nicotinic acid matrix was perceivedto be more promising. The consequence of this percep-tion is that the H illen kam pK ara s method has becomethe focus of international research, while the Tanakamethod has not received as much attention .Both methods suffered from very wide peaks in themass spectra corresponding to the polypeptide ions.The mass resolution for an ion of m/z = 25000 was-20 for Tanakas method and 40 or Hillenkamp andKarass method. Neither of the original papers4q5attempted to assign an accurate molecular mass to theions in the mass spectra, rather they quoted the calcu-lated mass for the protein sequence. No attempt wasmad e in either case to identify the ion species presentedby the peaks in the spectra, other than to say that theyrepresented the intact polypeptide molecular mass.A SAMPLE MASS SPECTRUM:HONEYBEE PHOSPHOLIPASE A,Before discussing the details of matrix-assisted laserdesorption, an example of the type of mass spectraobserved is in order. Figure 1 shows a portion of themass spectrum of the protein phospholipase A,,


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Figure 1. The singly charged mo lecu le-ion region of a matrix-assisted laser desorption mass spectrum of honeybee (Apis mellifera) venomphospholipaseA,. The sample loaded on the probe tip was 0.5 pI of matrix solution co ntaining 2.2 pmol of phospholipaseA, and 1 pmol ofbovine ribonuclease A (used as the calibrant). The matrix solution used was 50 mM sinapic acid, 0.1 Yo (w/w) trifluoracetic acid and 1 mMbarium acetate (used to precipitate phosphate anions) in water-acetonitrile (2 :1 , v/v). The laser irradiance used was - 1 MW /crn- and theacceleration voltage was +30 kV. The data were recorded in 5 ns time bins. The spectrum shown is the sum of 100 consecutive laser shotswithout selection and has been compressed into 20 ns time bins. The identity and molecular masses of the labelled peaks are discussed inthe text.obtained from honeybee venom. This spectrum wasobtained using the linear time-of-flight mass spectrom-eter constructed at Rockefeller University.6 The condi-tions, materials and spectrometer parameters are givenin the caption. All of the peaks shown are singlycharged, i.e. they are c onsidered to represent [M + H]cationic species. The peaks r and r + m are a result ofdoping the sample with bovine ribonuclease A to obtaina good mass calibration (see Ref. 17 for details of thisprocedure). The most intense peaks corresponding tohoneybee phospholipase A, are labeled a (M , = 15 247),b (M, = 15980), c (M , = 16174) and d (M, = 16293).These masses allow the assignment of peak (a) as theintact polypeptide chain of the molecule without anycarbohydrate. The other peaks are glycoforms of theprote in (see Ref. 18 for a discussion of glycoforms), eachcontaining tw o N-acetylglucosamines a nd two (b), three(c) or four (d) hexoses. The masses su ppo rt the sequenceproposed by Kuchler et (M, = 15 239), as opposedto that proposed by Shipolini et a/.20 M , = 14552).The agreement with Ref. 19 is not perfect, but the differ-ence could be caused by the partial oxidation of on e ofthe methionine residues in the molecule. The glycoformsobserved are consistent with the carbohydrate contentdetermined by chemical meth ods,2 0*2 lthough thefucosylated forms suggested by Weber et a/. were notobserved.

it is the result of history rather than an objectivenaming process. Early studies of the effect of laser irra-diation on molecules adsorbed on metal substrates ledto the naming of the process by which these moleculeswere removed from the surface as laser desorption.Here, desorption was the opposite of adsorption. Theprocess that leads to the matrix-assisted laser desorp-tion effect is of a different type. Microscopic observa-tion of the surface has demonstrated that a volume ofmaterial is removed in each laser shot. Presumably theformation of protein molecule ions is a direct conse-quence of the removal of this bulk material, not aremoval of a protein adsorbed on a stable layer ofmatrix molecules. Ablation, i.e. the removal of materialfrom a surface without reference to the mechanism ofthat removal or the final physical state of the removedmaterial, is probably a more accurate description of theprocess. Sublimation (the process of changing from asolid to a vapor without an intervening liquid state)may be the best term for the phenomenon, but its use ismodel dependent. There is little hope of changing thename of the phenomenon at this point, as matrix-assisted laser desorption has become a fixture in the lit-erature.MATRICES AND EXPERIMENTALPARAMETERS

A NOTE ON NOMENCLATUREThe term matrix-assisted laser desorption appears tohave been coined by the group in Munster in 1986,when it app ea rs in the title of a conference abstract.22 A1985 paper that outlines the concept of the process doesnot use this expression. Th e papers from D r Tanakasgro up at Shimadzu did n ot use this term at all.**Theyrefer to their technique as the ultra-fine metal plusliquid matrix method. Popular perception has tiedthese two different methods together, so the termmatrix-assisted laser desorption is now linked to bothgroups.The use of the word desorption is rather deceptive;

The properties of the matrix material used in theHillenkampKaras method are cri t ical . Only a fewcompounds are useful as matrices for proteins and poly-peptides. Despite the importance of the matrix com-pound, very little is known about what makes amaterial a good matrix. S urprisingly little research hasgone into discovering the factors that affect the func-tioning of these matrices. The few useful materials thathave been discovered have been exploited analytically.Reviewing the matrices that have been discovered,some common qualities can be identified. These condi-tions are the solubility of the matrix in the appropriatesolvents, the absorption spectrum of the matrix and thereactivity of the matrix.


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ACCOUNTS 657ablation process or was caused by the ionization step.The result does indicate that most of the ferulic acidmolecules ablated from the surface remain intact.After an appropriate matrix has been selected, thenext most im port ant experimental factors governing theproduction of ions are the characteristics of the laserirradiation. The amount of energy deposited by thelaser is critical for achieving good results. Experimentsw ith a lin ear tim e- of -fligh t i n s t r ~ m e n t ~ * ~ ~emonstratedthat a mass resolution of 400 was possible for proteinions and that the protein ion peaks broadened rapidlyas the intensity of the laser irradiation was increased.The relationship between the flux of protein ions pro-duced by a single pulse and the intensity of the pulse ishighly non-linear ( see Fig. 3). Below a thresho ld value,no protein ions are produced. Above the threshold, theion production increases rapidly to an apparent plateau.Although this threshold behavior is very importantfor both theoretical and practical considerations, thereis little agreement in the literature as to its value. Thisdiscrepancy may be caused by the difficulty in measur-ing the true intensity of the beam. The parameters usedto characterize a laser beams intensity are its irra-diance (J s - cm - ) or its fluence (J cm ). T o calcu-late the irradiance properly, it is necessary to know thetemporal profile of the beam, its energy and its spatialprofile at the sample. Calculating the fluence requiresonly the total energy and the spatial profile at thesample. Unfortunately, the only one of these parametersthat is easily measured is the total energy. The otherparameters are difficult to measure accurately, resultingin the use of approximations that do not account forshot-to-shot variability in the laser or the real spatialprofile of the focused beam. The calculation of thefluence is less sensitive to laser fluctuations than theirradiance, but it may not be the appropriate param eterto use when comparing effects produced by lasers ofwidely different pu lse lengths.There is no agreement in the literature about thethreshold irradiance. The original HillenkampKarasvalue was 100 MW c m- using a nicotinic acid matrix.Studies done a t R ockefeller University suggested 1 M Wcm- or the nicotinic acid matrix and similar valuesfor cinnamic acid derivative mat rice^.^.'^ The group a tUppsala found a value near 100 MW cm - for nicotinicacid and 100 M W cm-or sinapic an d ferulic acids,29although a value of - MW cm-2 was quoted for the

Throthold bradincePIatoau

/Protein

Io nCurrent

Laser lrradianceFigure 3. The behavior of protein ion intensity as a function oflaser irradiance near the ion p roduc tion threshold.

same matrices in anoth er reference. In a series ofcareful experiments on the shape of the ion yield versusirradiance curve, Ens et aLZ8 ound a value of 2 MWc m -2 for ferulic acid. Spen der et aL3 stated that theirthreshold value for sinapic acid was in the range 1-10M W c m - . The substantial disagreement am ong carefulinvestigators suggests that more work should be doneto determine the correct value for this importantparameter.A parameter that has proven to be less critical is thewavelength of the laser light. Many different pulsed UVlasers have been used to pro duce protein ions. Th e mainconstraint on the laser wavelength is the absorptionproperties of the matrix; the matrix must absorb thelight. Secondarily, the analyte should be transparent tothe light so that unw anted photochemical reactions arenot produced by the irradiation. For proteins , ab so rption in the amino acid side-chains becomes importantbelow 270 nm. Proteins that contain covalentlyattached chromophores, e.g. the heme group in cyto-chrome c, may require longer wavelengths to avoidabsorption.THE MASS SPECTROMETERTh e ion source used for matrix-assisted laser desorptionis conceptually very simple. Th e matrix-protein depositis made on a metal probe. The deposit is then irradiatedby the focused laser and the ions produced are extractedinto the mass spectrometer. This ion source is necessar-ily pulsed; ions are produced for a short time inresponse to the laser pulse. The minimum width report-ed for the extracted ion pulse is -50 ns for insulinions.32 N o information is available for the true ion pro-duction time width.

Most of the experimental studies that have been per-formed using a matrix-assisted laser desorption ionsource have used a time-of-flight mass spectrometer.The pulsed nature of the ion emission process meansthat no additional pulsing is necessary to establish astart time for the experiment. The time-of-flight massspectrometer is also ideally suited to analyzing a shortburst of ion s; all of the masses are separated anddetected without any scanning. Results have beenreported using a sector instrument followed by a time-of-f l ight ana1y~er. j~esults have also been shown for aqua drup ole ion-trap mass spectrometer with a m atrix-assisted laser de so rp tio n ~ o u c e . ~ ~he mass range ofthese latter studies has been limited, but some resultswere obtained.Detecting the large ions produced by matrix-assistedlaser desorption is more difficult than might be imag-ined. These ions have very large mass-to-charge ratios.Therefore, they are accelerated to low velocities by theion source. In order to be detected, these ions must beconverted to electrons when they collide with the detec-tors first electrode. The velocity of the high-mass ions isbelow the commonly accepted velocity threshold forcollision-induced electron conversion, so the detectionefficiency for these ions is probably very Themost commonly used conversions surfaces areberyllium-copper (in discrete dynode electronmultipliers) and lead oxide-coated glass (in multi-


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658 R. C. BEAVlS

channel plates). The development of high-efficiencydetectors for these large, slow-moving ions shouldgreatly improve the intensity of ion signals.Unlike the situation encountered by plasma desorp-tion mass spectrometry in its early development period,matrix-assisted laser desorption has led to the develop-ment of four commercial instruments, so far. The ear-liest of these is a reflectron instrument that wasmarketed by Shimadzu to take advantage of theTanaka method. The second instrument was a lineartime-of-flight mass spectrometer sold by Vestec, usingthe design developed at Rockefeller University. Twoother machines have recently become available fromFinnigan-M AT. One is a linear time-of-flight mass spec-trometer and the other is a laser microprobe ion sourcewith a reflectron analyzer (this latter machine wasdeveloped in cooperation with Professor Hillenkampsgroup in Miinster). The current commercial interest inthe mass spectrometry of proteins and time-of-flightmass spectrometers has stirred the interest of investiga-tors in both industry and academia.

FUTURE DIRECTIONSThere are two general branches of future research:applications and basic understanding. The work onapplications has already begun with the distribution ofcommercial mass spectrometers. The current standardtechnique available to a biomedical researcher for deter-mining the molecular mass of a protein is sodiumdodecyl sulfate polyacrylamide gel electrophoresis.Mass spectrometry, using matrix-assisted laser desorp-tion and electrospray ionization, promises to replaceelectrophoretic measurements with higher accuracy andresolution. The ability to determine a n accu rate molecu-lar mass for proteins will change the manner in whichbiomedical research is performed.

A basic understanding of the mechanism of matrix-assisted laser desorption and the role of the matrix maytake longer. Th e problem is a difficult one. The ablatio nevent is a fleeting one, which occurs in conditions thatare very far from equilibrium. Understanding the eventwill take a substantial effort by many technically sophis-ticated groups. A problem that has produced a greatdeal of research and speculation is improving the massresolution obtained from mass spectrometers that usematrix-assisted laser desorption. The use of reflectronanalyzers may prove to be the answer (these instru-ments correct the time aberrations caused by trans-lational energy distributions). However, the results sofar have not been as good as expected. Research intothe more fundamental problems may have to wait untilthe resolution problem has been solved.Another line of related research is the work beingcarried ou t by Peter W illiams gro up at Arizona StateU n i ~ e r s i t y . ~ ~ . ~ hey have reported the use of a laserablation technique using a water-ice substra te (on acryostat) with polynucieotide analyte molecules. Thepolynucleotides are dissolved in liquid water, which isused to coat a cryostat tip. The ice-analyte mixture isirradiated with a focused visible wavelength laser. Theice is transparent to the laser, but it is absorbed by themetal substrate. The result of this irradiation is the pro-duction of intact polynucleotide molecules that havebeen observed with a linear time-of-flight mass spec-trometer. The results so far have been preliminary, butthey are so encouraging that there should be muchmore to come.AcknowledgementsI thank Brian Chait, Franz Hillenkamp. Micheal Karas, Ute Bahr, BoSundqvist, Per HAkansson, Yvon LeBeyec, Franz Rollgen, WernerEns, Ken Standing, Peter Williams, Randy Nelson, Peter Roepstorfand Bo b Cotter for their contributions to matrix-assisted laserdesorption and their willingness to discuss the subject at length.

REFERENCES1. K. Tanaka, Y. Ido. S. Akita, Y. Yoshida and T. Yoshida, in Pro-ceedings of the Second Japan-China Joint Sympsoium onMass Spectrometry (Osak a,Japan) (1987).2. M. aras and F. Hillenkamp, in Ion Formation from OrganicSolids ( IFOS IV), ed . by A. Benninghoven, p. 103. Wiley,New York (1989).3. M. Karas and F. Hillenkamp, Adv. Mass Spectrom., 11A, 354(1988).4. K. Tanaka, H. Waki, Y. Ido, S. Akita, Y. Yoshida and T.Yoshida, Rapid Commun. Mass Spectrom.2, 151 (1 988).5. M. Karas and F. Hillenkamp.Ana1. Chem.60, 299 (1988).6. R . C. Beavis and B. T. Chait, Rapid Commun. Mass Spectrom.3, 233 (1989).7. M. Salehpour, I.Perera, J. Kjellberg, A. Hedin, M. A. Islamian,

P.Hakansson and B. U. R. Sundqvist, Rapid Commun. MassSpectrom.3. 259 (1989).8. B. Spengler and R. J. Cotter, Anal. Chem. 62, 793 (1 990).9. R . 0. Macfarlane and D. F. Torgesson, Science 191, 970(1976).10. G. P. Jonsson, A. B. Hedin, P. L. Hakansson, B. U. R. Sun-dqvist, 8. G. S. Save, P. F. Nielsen, P. Roepstorff, K.-E.Johansson, I. Kamensky and M. S. L. Lindberg, Anal. Chem.58,1084 (1986).11. A. Benninghoven and W. Sichtermann, Anal. Chem. 50, 1180(1978).

12. M. Barber, R. S. Bordoli, R. D. Sedgwick and A. N. Tyler,Nature (London ) 293, 270 (1981).13. M. Barber, R. S. Bordoli, R. C. Sedgwick and A. N. Tyler, Anal.Chem. 64,645A (1982).14. M. Karas, D. Bachmann and F. Hillenkamp, Anal. Chem. 57,2935 (1985).15. M. Karas, D. Bachmann. U. Bahr and F. Hillenkamp, Int . J .Mass Spectrom . on Processes 78, 53 (1987).16. M. Karas, U. Bahr and F. Hillenkamp, Int. J . Mass Spectrom.Ion Processes 92, 231 (1989).17. R . C. Beavis and B. T. Chait, Anal. Chem. 62, 1836 (1990).18. B. Alberts, D.Bray, J. Lewis, M. Raff. K. Roberts and J. D.Watson, The Molecular Biology of the Cell, 2nd edn, pp. 446-449. Garland, New York (1989).19. K. Kuchler, M. Gmachl, M. J. Sippl and G. Kreil, Eur. J.Biochem. 184,249 (1989).20. R . A. Shipolini, G. L. Callewaert, R. C. Cottrell and C. A.Vernon, Eur. J . Biochem.48,465 (1974).21. A. Weber, L. M2rz and F. Altmann, Com p. Biochern. Physiol.838. 321 (1 986).22. M. Karas, D. Bachmann and F. Hillenkamp, in Advances inMass Spectrometry, Vol. 106. ed. y J. F. J. Todd, Wiley, Chi-Chester, p. 1986 (1986).23. R. C. Beavis and B. T. Chait. Rapid Commun. Mass Spectrom.3, 432 (1989).


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ACCOUNTS 65924. M. Karas, F. Hillenkamp, R. C. Beavis and B. T. Chait, Anal.Chem. 63,1193A (1991).25. S. J. Doktycz, P. J. Savickas and D. A. Krueger, RapidCommun. Mass Specrrom.5 , 145 (1 991).26. T. Huth-Fehre and C. H. Becker, Rapid Commun. Mass Spec-trom. 5 , 378 (1991).27. R. C. Beavis and B. T. Chait. Proc. Narl. Acad. Sci. , USA , 87,6873 (1 990).28. W. E. Ens, Y. Mao, F. M&er and K . G. Standing, RapidCommun. Mass Specrrom. 5, 117 (1 991).29. 1. K. Perera, J. M. Candy, P. Hakansson, A. E. Oakley, G.Brinkmalm and B. U. R. Sundqvist, Rapid Commun. MassSpecrrom.4, 527 (1990).30. I. K. Perera, E. Uzcategui, P. Hakansson, G. Brinkmalm, G.Pettersson, G. Johansson and B. U. R. Sundqvist, RapidCommun. Mass Specrrom.4, 285 (1 990) .31. B. Spengler, D. Kirsch, R. Kaufmann, Rapid Commun. MassSpecrrom.5, 198 (1 991).

BIOGRAPHYRonald Beavis received his PhD in Physics from theUniversity of Manitoba in 1987 where he worked underKen Standing on the analysis of biopolymers by sec-ondary ion mass spectrometry. He then spent eighteenmonths as a NATO Research Fellow at the TechnicalUniversity of Munich, in the laboratory of EdwardSchlag, working on the role of chemical reactions thatoccur during IR laser desorption. He then spent twoyears at Rockefeller University working with BrianChait on problems related to matrix-assisted laserdesorption. Currently, he is an Assistant Professor ofPhysics at the Memorial University of Newfoundland,where he is continuing his examination of the matrix-assisted laser desorption process.

32. R. C. Beavis and B. T. Chait, in Methods and Mechanisms forProducing Large Ions from Large Molecules, ed. by W. Ens &K. G. Standing, NATO AS1 Science Series. p. 227. PlenumPress, New York (1991).33. F. H. Strobel, T. Solouki, M. A. White and D. H. Russel,J.A m .Soc. Mass Spectrom.2, 91 (1991).34. R. L. Hettich and M. V. Buchanan, J. A m . SOC.Mass Spec-rrom. 2, 22 (1 990).35. G. B. Baptista, A. Brunelle, P. Chaurand, S. Della-Negra, J.Depauw and Y. Le Beyec, Rapid Commun. Mass Spectrom. 5,

632 (1991).36. R. W. Nelson, M. J. Rainbow, D. E. Lohr and P. Williams,Science 246,1585 (1 989).37. R. W. Nelson, R. M. Thomas and P. Williams, Rapid Comm un.Mass Spectrom.4, 348 (1990).

BS313 T1 Maldi Supp Reading - UV MALDI-Evolution and Principle

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