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Blomberg, Lars G.
Alternative names
Publications (10 of 91) Show all publications
Bohlin, M., Johannesson, I., Carlsson, G., Heegaard, N. H. & Blomberg, L. G. (2012). Estimation of the amount of β2-glycoprotein I adsorbed at the inner surface of fused silica capillaries after acidic, neutral and alkaline pretreatment. Electrophoresis, 33(12), 1695-1702
Open this publication in new window or tab >>Estimation of the amount of β2-glycoprotein I adsorbed at the inner surface of fused silica capillaries after acidic, neutral and alkaline pretreatment
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2012 (English)In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 33, no 12, p. 1695-1702Article in journal (Refereed) Published
Abstract [en]

Sample adsorption to the inner surface of fused silica capillaries is a common problem in

CE when analyzingmacromolecules and is harmful to the analysis. We previously utilized

the pH hysteresis effect of fused silica to facilitate electrophoresis of the strongly adsorbing

protein β2gpI in plain-fused silica capillaries at neutral pH. In the present paper, the

effect of different pretreatments of the capillary on the adsorption of the β2-glycoprotein

I has been investigated using electroosmosis markers, SDS mobilization, and imaging

based on indirect immunofluorescence microscopy for direct visualization. The amount

of β2gpI adsorbed on the surface was probed using all these independent techniques after

electrophoresis at neutral pH on capillaries pretreated with HCl, background electrolyte

(BGE), and NaOH. BGE pretreatment was included as a positive control. We found that

80% or more of the starting material was adsorbed to the inner surface of the silica

capillaries during electrophoresis after pretreatment with only BGE or with NaOH, but

after acidic pretreatment the loss was consistently less than 20%. NaOH most efficiently

removes adsorbed protein between runs. A theoretical calculation of the pH change of

the BGE showed that electrolysis affects the pH more than the deprotonation of silanols

during electrophoresis. We conclude that acidic pretreatment of fused silica capillaries

diminishes adsorption of β2gpI by decreasing charge-dependent wall adsorption.

 

Place, publisher, year, edition, pages
Weinheim, Germany: John Wiley & Sons, 2012
Keywords
Acidic pretreatment / Capillary electrophoresis / pH-hysteresis effect / Protein adsorption
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-30939 (URN)10.1002/elps.201100592 (DOI)000305792500003 ()22674218 (PubMedID)
Available from: 2014-01-20 Created: 2014-01-20 Last updated: 2019-09-19Bibliographically approved
Bohlin, M. E., Blomberg, L. G. & Heegaard, N. H. (2011). Effects of ionic strength, temperature and conformation on affinity interactions of β2-glycoprotein I monitored by capillary electrophoresis. Electrophoresis, 32, 728-737
Open this publication in new window or tab >>Effects of ionic strength, temperature and conformation on affinity interactions of β2-glycoprotein I monitored by capillary electrophoresis
2011 (English)In: Electrophoresis, ISSN 0173-0835, E-ISSN 1522-2683, Vol. 32, p. 728-737Article in journal (Refereed) Published
Abstract [en]

We have used CE to evaluate the interaction between β2-glycoprotein I (β2gpI) and heparin. β2gpI is a human plasma protein involved in the blood coagulation cascade. It is of interest to functionally characterize the interactions of β2gpI because the exact function is not entirely known and because circulating autoantibodies against β2gpI are associated with an increased risk of thrombotic events.

 

The effect of the ionic strength, temperature, and conformation of the protein on the interaction between β2gpI and heparin has been studied. The CE procedure for this study is simple, fast and automatic. β2gpI and heparin were allowed to interact during electrophoresis at different ionic strength buffers and at different capillary temperatures. To mimic perturbation of the conformation of β2gpI, different denaturing agents (SDS, ACN and urea) were added to the background electrolyte. While simple 1:1 binding isotherms were obtained at 22 °C the data strongly suggests that at physiological temperature the binding stoichiometry is not 1:1 and/or that cooperative interactions begin to play a role. We found that (i) the KD values differed by a factor of 60 at the ionic strengths studied (ii) β2gpI was resistant to denaturation with SDS and ACN, but was partially denatured by urea and (iii) the KD for the β2gpI-heparin interaction in the presence of urea was 10 times higher than the KD determined at the same conditions without urea added. Therefore, we conclude that the interaction between β2gpI and heparin is dependent on electrostatic interactions and on the conformation of β2gpI. 

Keywords
affinity capillary electrophoresis; β2-glycoprotein I; conformation; heparin; pH hysteresis effect
National Category
Analytical Chemistry
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-8275 (URN)10.1002/elps.201000538 (DOI)000288602000012 ()
Available from: 2011-09-19 Created: 2011-09-19 Last updated: 2018-01-15Bibliographically approved
Bohlin, M., Blomberg, L. G., Olsson, O. & Heegaard, N. H. . (2011). Structure-activity studies of human beta2-glycoprotein I using capillary electrophoresis. Paper presented at Joint congress 2011: Microscale Bioseparations San Diego. Paper presented at Joint congress 2011: Microscale Bioseparations San Diego.
Open this publication in new window or tab >>Structure-activity studies of human beta2-glycoprotein I using capillary electrophoresis
2011 (English)Conference paper, Published paper (Refereed)
Abstract

We have investigated various modes of CE to evaluate the interaction between beta2-glycoprotein I (b2gpI) and a number of anionic ligands to contribute to the elucidation of the structure-function relationship of b2gpI. b2gpI is a plasma protein which is involved in the blood coagulation cascade under normal, physiological conditions, however, its precise function is undefined. It is also involved in pathological conditions such as the so-called anti-phospholipid syndrome, where anti-b2gpI autoantibodies induce a prothrombotic state. Therefore, functional characterization of b2gpI under near physiological conditions is of interest.

To avoid charge-dependent analyte adsorption to the inner surface of the capillary wall, we have utilized the pH hysteresis effect, where an acidic pretreatment of the capillary made it possible to perform subsequent CE analyses of b2gpI at neutral pH.

The interaction between b2gpI and the anionic ligand heparin was studied with migration shift ACE, where the ionic strength, temperature and conformation of b2gpI were easily varied. The interaction between b2gpI and phosphatidylcholine/phosphatidylserine liposomes are subject to an ongoing investigation by means of migration shift ACE, frontal analysis CE, partial filling CE and pre-equilibration partial filling ACE.

We conclude that differential, but relatively low binding affinities that are highly dependent on electrostatic interactions and on a preserved conformation of the protein, characterize its interactions with ligands that in vivo will be present in multiple copies on e.g. cell surfaces. The CE procedure for this study is simple, fast and automatic and quantitative binding affinity parameters are conveniently obtained using small amounts of biological materials.

National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-10742 (URN)
Conference
Joint congress 2011: Microscale Bioseparations San Diego
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2013-06-12Bibliographically approved
Bohlin, M. & Blomberg, L. G. (2010). Affinity studies of beta-2-glycoprotein I using capillary electrophoresis. Paper presented at Microscale Bioseparations 2010, Prag, Tjeckien. Paper presented at Microscale Bioseparations 2010, Prag, Tjeckien.
Open this publication in new window or tab >>Affinity studies of beta-2-glycoprotein I using capillary electrophoresis
2010 (English)In: / [ed] Lars G. Blomberg, Niels H. H. Heegaard, 2010Conference paper, Published paper (Refereed)
Abstract

Beta2-glycoprotein I (b2gpI), also known as apolipoprotein H, is a plasma protein which is involved in the blood coagulation cascade. It binds negatively charged substances such as heparin, DNA, and anionic phospholipids. A number of functions of b2gpI have been proposed, however, the precise function is still not entirely known. Circulating autoantibodies against b2gpI are associated with increased risk of thrombotic events, such as thrombosis and reoccurring fetal loss. It is therefore of interest to functionally characterize b2gpI including the influence of anti-b2gpI autoantibodies on the ligand binding behavior of the protein. The characterization of interactions between biological molecules may be accomplished by capillary electrophoresis under non-denaturing conditions, without the need for immobilization. To avoid charge dependent analyte adsorption to the negative charges of the capillary wall we found the pH hysteresis effect of silica very useful. An acidic pretreatment of the capillary made it possible to perform a subsequent analysis at neutral pH. We were able to perform binding studies between b2gpI and heparin and monosaccharides at different ionic strengths and temperatures in a simple way. We could also study the effect of mildly denaturing conditions on the binding to the different ligands simply by adding sodium dodecyl sulfate (SDS), urea and ACN to the background electrolyte.

The approach is simple, fast and automatic. The ionic strength, temperature and other parameters such as denaturing agents could easily be changed to characterize the binding between b2gpI and different ligands.

National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-10136 (URN)
Conference
Microscale Bioseparations 2010, Prag, Tjeckien
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2013-06-12Bibliographically approved
Bohlin, M. & Blomberg, L. G. (2010). Affinity studies of beta2-glycoprotein I using capillary electrophoresis. Paper presented at Analysdagarna 2010, Uppsala. Paper presented at Analysdagarna 2010, Uppsala.
Open this publication in new window or tab >>Affinity studies of beta2-glycoprotein I using capillary electrophoresis
2010 (English)In: / [ed] Lars G. Blomberg, Niels Heegaard, 2010Conference paper, Published paper (Refereed)
Abstract

beta2-glycoprotein I (b2gpI), also known as apolipoprotein H, is a plasma protein which is involved in the blood coagulation cascade. It binds negatively charged substances such as heparin, DNA, and anionic phospholipids. A number of functions of b2gpI have been proposed, however, the precise function is still not entirely known. Circulating autoantibodies against b2gpI are associated with increased risk of thrombotic events, such as thrombosis and reoccurring fetal loss. It is therefore of interest to functionally characterize b2gpI including the influence of anti-b2gpI autoantibodies on the ligand binding behavior of the protein. The characterization of interactions between biological molecules may be accomplished by capillary electrophoresis under non-denaturing conditions, without the need for immobilization. To avoid charge dependent analyte adsorption to the negative charges of the capillary wall we found the pH hysteresis effect of silica very useful. An acidic pretreatment of the capillary made it possible to perform a subsequent analysis at neutral pH. We were able to perform binding studies between b2gpI and heparin at different ionic strengths and temperatures in a simple way. We could also study the effect of mildly denaturing conditions on the binding to the ligand simply by adding sodium dodecyl sulfate (SDS), urea and ACN to the background electrolyte. The approach is simple, fast and automatic

National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-10137 (URN)
Conference
Analysdagarna 2010, Uppsala
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2013-06-12Bibliographically approved
Blomberg, L. G., Onnerud, H., Bassiouny, F. & Abdel-Rehim, M. (2010). Chromatographic behaviour of bupivacaine and five of its major metabolites in human plasma, utilizing solid-phase extraction and capillary gas chromatography. Journal of Chromatographic Science, 48(4), 294-298
Open this publication in new window or tab >>Chromatographic behaviour of bupivacaine and five of its major metabolites in human plasma, utilizing solid-phase extraction and capillary gas chromatography
2010 (English)In: Journal of Chromatographic Science, ISSN 0021-9665, E-ISSN 1945-239X, Vol. 48, no 4, p. 294-298Article in journal (Refereed) Published
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-9886 (URN)000276055100008 ()20412652 (PubMedID)
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2017-12-07Bibliographically approved
Blomberg, L. G., Altun, Z., Skoglund, C. & Abdel-Rehim, M. (2010). Monolithic methacrylate packed 96-tips for high throughput bioanalysis. Journal of Chromatography A, 1217(16), 2581-2588
Open this publication in new window or tab >>Monolithic methacrylate packed 96-tips for high throughput bioanalysis
2010 (English)In: Journal of Chromatography A, ISSN 0021-9673, Vol. 1217, no 16, p. 2581-2588Article in journal (Refereed) Published
Place, publisher, year, edition, pages
Elsevier, 2010
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-9885 (URN)10.1016/j.chroma.2009.10.072 (DOI)
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2015-06-24Bibliographically approved
Blomberg, L. G. (2010). Professional supervision of doctoral education in science. In: A. Ott och C.E. Olivestam (Ed.), Strömstad akademi, Vision och verklighet (pp. 91-112). Strömstad: Strömstad akademi
Open this publication in new window or tab >>Professional supervision of doctoral education in science
2010 (English)In: Strömstad akademi, Vision och verklighet / [ed] A. Ott och C.E. Olivestam, Strömstad: Strömstad akademi , 2010, p. 91-112Chapter in book (Other academic)
Place, publisher, year, edition, pages
Strömstad: Strömstad akademi, 2010
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-9888 (URN)9789186607005 (ISBN)
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2012-02-09Bibliographically approved
Blomberg, L. G. & Abdel-Rehim, M. (2010). Recent advances in microextraction by packed sorbent for bioanalysis. Journal of Chromatography A, 1217(16), 2569-2580
Open this publication in new window or tab >>Recent advances in microextraction by packed sorbent for bioanalysis
2010 (English)In: Journal of Chromatography A, ISSN 0021-9673, E-ISSN 1873-3778, Vol. 1217, no 16, p. 2569-2580Article in journal (Refereed)
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-9884 (URN)
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2017-12-07Bibliographically approved
Blomberg, L. G. & Abdel-rehim, M. (2009). Current advances in microextraction by packed sorbent (MEPS) for bioanalysis applications. LCGC Europé, 22(1), 8-19
Open this publication in new window or tab >>Current advances in microextraction by packed sorbent (MEPS) for bioanalysis applications
2009 (English)In: LCGC Europé, ISSN 1471-6577, Vol. 22, no 1, p. 8-19Article in journal (Refereed) Published
Place, publisher, year, edition, pages
Woodland Hills: Advanstar Communications, 2009
National Category
Chemical Sciences
Research subject
Chemistry
Identifiers
urn:nbn:se:kau:diva-9869 (URN)
Available from: 2012-02-08 Created: 2012-02-08 Last updated: 2015-07-16Bibliographically approved
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