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New methods for sensitive analysis with nanoelectrospray ionization mass spectrometry
KTH, School of Chemical Science and Engineering (CHE), Chemistry, Analytical Chemistry.
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

In this thesis, new methods that address some current limitations in nanoelectrospray mass spectrometry (nESI-MS) analysis are presented. One of the major objectives is the potential gain in sensitivity that can be obtained when employing the proposed techniques.

In the first part of this thesis, a new emitter, based on the generation of electrospray from a spray orifice with variable size, is presented. Electrospray is generated from an open gap between the edges of two individually mounted, pointed tips. The fabrication and evaluation of two different types of such emitters is presented; an ESI emitter fabricated from polyethylene terephtalate (Paper I), and a high-precision silicon device (Paper II). Both emitters were surface-treated in a selective way for an improved wetting of the gap and to confine the sample solution into the gap.

In the second part of this thesis, different methods for improved sensitivity of nESI-MS analysis have been developed. In Paper III, a method for nESI-MS analysis from discrete sample volumes down to 1.5 nL is presented, using commercially available nESI needles. When analyzing attomole amounts of analyte in such a small volume of sample, an increased sensitivity was obtained, compared to when analyzing equal amounts in conventional nESI-MS analysis. To be able to analyze smaller sample volumes, needles with a narrower orifice and a higher flow resistance were needed. This triggered the development of a new method for fabrication of fused silica nESI needles (Paper IV). The fabrication is based on melting of a fused silica capillary by means of a rotating plasma, prior to pulling the capillary into a fine tip. Using the described technique, needles with sub-micrometer orifices could be fabricated. Such needles enabled the analysis of sample volumes down to 275 pL, and a further improvement of the sensitivity was obtained. In a final project (Paper V), nESI-MS was used to study the aggregation behavior of Aβ peptides, related to Alzheimer’s disease. An immunoprecipitation followed by nESI-MS was employed. This technique was also utilized to study the selectivity of the antibodies utilized.

Place, publisher, year, edition, pages
Stockholm: KTH , 2010. , vii, 70 p.
Series
Trita-CHE-Report, ISSN 1654-1081 ; 2010:41
Keyword [en]
mass spectrometry, electrospray ionization, nanoelectrospray ionization, adjustable gap, emitter, miniaturization, improved sensitivity, nanoliter/picoliter sample volumes, Alzheimer’s disease, amyloid-beta (Aβ)
National Category
Analytical Chemistry
Identifiers
URN: urn:nbn:se:kth:diva-25917ISBN: 978-91-7415-751-2 (print)OAI: oai:DiVA.org:kth-25917DiVA: diva2:360781
Public defence
2010-11-26, K1, Teknikringen 56, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
QC 20101112Available from: 2010-11-12 Created: 2010-11-04 Last updated: 2010-11-12Bibliographically approved
List of papers
1. Electrospray Ionization from a Gap with Adjustable Width
Open this publication in new window or tab >>Electrospray Ionization from a Gap with Adjustable Width
2006 (English)In: Rapid Communications in Mass Spectrometry, ISSN 0951-4198, E-ISSN 1097-0231, Vol. 20, no 21, 3176-3182 p.Article in journal (Refereed) Published
Abstract [en]

In this paper, we present a new concept for electrospray ionization mass spectrometry, where the sample is applied in a gap which is formed between the edges of two triangular-shaped tips. The size of the spray orifice can be changed by varying the gap width. The tips were fabricated from polyethylene terephthalate film with a thickness of 36 μm. To improve the wetting of the gap and sample confinement, the edges of the tips forming the gap were hydrophilized by means of silicon dioxide deposition. Electrospray was performed with gap widths between 1 and 36 μm and flow rates down to 75 nL/min. The gap width could be adjusted in situ during the mass spectrometry experiments and nozzle clogging could be managed by simply widening the gap. Using angiotensin I as analyte, the signal-to-noise ratio increased as the gap width was decreased, and a shift towards higher charge states was observed. The detection limit for angiotensin I was in the low nM range.

Keyword
article, electrospray mass spectrometry, equipment, equipment design, evaluation, instrumentation, methodology, microfluidic analysis, reproducibility, sensitivity and specificity
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-8437 (URN)10.1002/rcm.2710 (DOI)000241627000003 ()2-s2.0-33750325735 (Scopus ID)
Note
QC 20100913Available from: 2008-05-13 Created: 2008-05-13 Last updated: 2012-02-24Bibliographically approved
2. Electrospray Ionization from an Adjustable Gap between two Silicon Chips
Open this publication in new window or tab >>Electrospray Ionization from an Adjustable Gap between two Silicon Chips
Show others...
2009 (English)In: Journal of Mass Spectrometry, ISSN 1076-5174, E-ISSN 1096-9888, Vol. 44, no 2, 171-181 p.Article in journal (Refereed) Published
Abstract [en]

In this paper, a silicon chip - based electrospray emitter with a variable orifice size is presented. The device consists of two chips, with a thin beam elevating from the center of each of the chips. The chips are individually mounted to form an open gap of a narrow, uniform width between the top areas of the beams. The electrospray is generated at the endpoint of the gap, where the spray point is formed by the very sharp intersection between the crystal planes of the < 100 > silicon chips. Sample solution is applied to the rear end of the gap from a capillary via a liquid bridge, and capillary forces ensure a spontaneous imbibition of the gap. The sample solution is confined to the gap by means of a hydrophobic treatment of the surfaces surrounding the gap, as well as the geometrical boundaries formed by the edges of the gap walls. The gap width could be adjusted between 1 and 25 μm during electrospray experiments without suffering from any interruption of the electrospray process. Using a peptide sample solution, a shift toward higher charge states and increased signal-to-noise ratios was observed when the gap width was decreased. The limit of detection for the peptide insulin (chain B, oxidized) was approximately 4 nM. We also show a successful interfacing of the electrospray setup with capillary electrophoresis.

Keyword
Adjustable gap, Chip, Electrospray ionization, Emitter, Mass spectrometry, Nozzle
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-8438 (URN)10.1002/jms.1478 (DOI)000264013300002 ()18946877 (PubMedID)2-s2.0-59949093565 (Scopus ID)
Note
QC 20100913. Uppdaterad från Manuskript till Artikel (20100913)Available from: 2008-05-13 Created: 2008-05-13 Last updated: 2010-11-12Bibliographically approved
3. Electrospray ionization mass spectrometry from discrete nanoliter-sized sample volumes
Open this publication in new window or tab >>Electrospray ionization mass spectrometry from discrete nanoliter-sized sample volumes
2010 (English)In: Rapid Communications in Mass Spectrometry, ISSN 0951-4198, E-ISSN 1097-0231, Vol. 24, no 17, 2561-2568 p.Article in journal (Refereed) Published
Abstract [en]

We describe a method for nanoelectrospray ionization mass spectrometry (nESI-MS) of very small sample volumes. Nanoliter-sized sample droplets were taken up by suction into a nanoelectrospray needle from a silicon microchip prior to ESI. To avoid a rapid evaporation of the small sample volumes, all manipulation steps were performed under a cover of fluorocarbon liquid. Sample volumes down to 1.5 nL were successfully analyzed, and an absolute limit of detection of 105 attomole of insulin (chain B, oxidized) was obtained. The open access to the sample droplets on the silicon chip provides the possibility to add reagents to the sample droplets and perform chemical reactions under an extended period of time. This was demonstrated in an example where we performed a tryptic digestion of cytochrome C in a nanoliter-sized sample volume for 2.5h, followed by monitoring the outcome of the reaction with nESI-MS. The technology was also utilized for tandem mass spectrometry (MS/MS) sequencing analysis of a 2 nL solution of angiotensin I.

National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-26071 (URN)10.1002/rcm.4674 (DOI)000281355000011 ()2-s2.0-77955386519 (Scopus ID)
Note

QC 20101111

Available from: 2010-11-11 Created: 2010-11-11 Last updated: 2017-06-08Bibliographically approved
4. New Method for Fabrication of Fused Silica Emitters with Submicrometer Orifices for Nanoelectrospray Mass Spectrometry
Open this publication in new window or tab >>New Method for Fabrication of Fused Silica Emitters with Submicrometer Orifices for Nanoelectrospray Mass Spectrometry
2011 (English)In: Analytical Chemistry, ISSN 0003-2700, E-ISSN 1520-6882, Vol. 83, no 20, 7771-7777 p.Article in journal (Refereed) Published
Abstract [en]

In this paper, we describe a new method for fabrication of nanoelectrospray emitters. The needles were pulled from fused silica capillary tubing, which was melted by means of a plasma, formed by electrical discharges between two pointed platinum electrodes. A key feature of the pulling device is a rotating configuration of the electrodes, which results in an even radial heating of the capillary. The construction of the setup is straightforward, and needles with a variety of shapes can be fabricated, including orifices of submicrometer dimensions. Pulled needles with long tapered tips and an orifice of 0.5 mu m were utilized for electrospray ionization mass spectrometry (ESI-MS) of discrete sample volumes down to 275 pL. The picoliter-sized samples were transferred into the tip of the needle from a silicon microchip by aspiration. To avoid a rapid evaporation of the sample, all manipulations were performed under a cover of a fluorocarbon liquid. The limit of detection was measured to be ca. 20 attomole for insulin (chain B, oxidized).

Keyword
ELECTROSPRAY-IONIZATION SOURCE, DISCOVERY, NANOSPRAY
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-26072 (URN)10.1021/ac201308n (DOI)000295817500028 ()2-s2.0-80054701334 (Scopus ID)
Note
QC 20101111 Updated from manuscript to article in journal.Available from: 2010-11-11 Created: 2010-11-11 Last updated: 2017-12-12Bibliographically approved
5. Separation and characterization of aggregated species of amyloid-beta peptides
Open this publication in new window or tab >>Separation and characterization of aggregated species of amyloid-beta peptides
Show others...
2010 (English)In: Analytical and Bioanalytical Chemistry, ISSN 1618-2642, E-ISSN 1618-2650, Vol. 397, no 6, 2357-2366 p.Article in journal (Refereed) Published
Abstract [en]

We have investigated the use of isoelectric focusing and immunodetection for the separation of low molecular weight species of amyloid-beta (A beta) peptides from their aggregates. From solutions of A beta(1-40) or A beta(1-42) monomeric peptides, low molecular weight material appeared at a pI value of ca. 5, while the presence of aggregates was detected as bands, observed at a pI of 6-6.5. The formation of A beta aggregates (protofibrils) was verified by a sandwich ELISA, employing the protofibril conformation-selective antibody mAb158. In order to study the aggregation behavior when using a mixture of the monomers, we utilized the IEF separation combined with Western blot using two polyclonal antisera, selective for A beta(1-40) and A beta(1-42), respectively. We conclude that both monomers were incorporated in the aggregates. In a further study of the mixed aggregates, we used the protofibril conformation-selective antibody mAb158 for immunoprecipitation, followed by nanoelectrospray mass spectrometry (IP-MS). This showed that the A beta(1-42) peptide is incorporated in the aggregate in a significantly larger proportion than its relative presence in the original monomer composition. IP-MS with mAb158 was also performed, and compared to IP-MS with the A beta-selective antibody mAb1C3, where a monomeric A beta(1-16) peptide was added to the protofibril preparation. A beta(1-16) is known for its poor aggregation propensity, and acted therefore as a selectivity marker. The results obtained confirmed the protofibril conformation selectivity of mAb158.

Keyword
Alzheimer's disease, Protofibrils, Amyloid-beta (A beta), Isoelectric focusing (IEF)., Mass spectrometry (MS), Bioanalytical methods, Capillary electrophoresis / Electrophoresis
National Category
Analytical Chemistry
Identifiers
urn:nbn:se:kth:diva-26073 (URN)10.1007/s00216-010-3839-9 (DOI)000279453000038 ()2-s2.0-77955981824 (Scopus ID)
Conference
6th International Conference on Instrumental Methods of Analysis Athens, GREECE, OCT 04-08, 2009
Note
QC 20110214Available from: 2010-11-11 Created: 2010-11-11 Last updated: 2017-12-12Bibliographically approved

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