Pulsed-field gel electrophoresis for the separation of large protein molecules exemplified by the isoforms of apolipoprotein (a)

G. Csako, B. Nagy, R. Costello, J. C. Castelli, A. M. Hruszkewycz

Research output: Contribution to journalArticle

3 Citations (Scopus)

Abstract

The performance of pulsed-field gel electrophoresis (PFGE) was assessed for the separation of protein molecules. The allelic isoforms of apolipoprotein (a) (apo[a]) served as a model for this study because apo(a) is an unusually large protein, consisting of a variable number of repeating units. PFGE and, for comparison, conventional electrophoresis of human sera were carried out under reducing conditions in agarose gel. After blotting proteins onto nitrocellulose membrane, a combination of monospecific rabbit anti-apo(a) antibody and alkaline phosphatase-conjugated protein A detected apo(a) isoforms in all sera tested. The various apo(a) isoforms were effectively resolved within two repeating units ('kringles') by both PFGE and conventional electrophoresis, but the type of agarose gel used greatly affected the speed of electrophoretic separation. In a series of 89 human sera, 59 double-band and 30 single-band patterns were seen using either electrophoretic system. However, one specimen produced double bands with PFGE, while only a single band could be detected by conventional electrophoresis, and with another specimen the opposite occurred. A total of 22 different apo(a) isoforms were identified among these patterns. It is concluded that the increasingly available PFGE technology is a practical alternative to conventional agarose electrophoresis for the separation of large protein molecules.

Original languageEnglish
Pages (from-to)926-929
Number of pages4
JournalElectrophoresis
Volume15
Issue number7
Publication statusPublished - 1994

Fingerprint

Apoprotein(a)
Pulsed Field Gel Electrophoresis
Electrophoresis
Protein Isoforms
Gels
Molecules
Proteins
Sepharose
Serum
Kringles
Collodion
Agar Gel Electrophoresis
Staphylococcal Protein A
Alkaline Phosphatase
Rabbits
Technology
Membranes
Antibodies

ASJC Scopus subject areas

  • Clinical Biochemistry

Cite this

Pulsed-field gel electrophoresis for the separation of large protein molecules exemplified by the isoforms of apolipoprotein (a). / Csako, G.; Nagy, B.; Costello, R.; Castelli, J. C.; Hruszkewycz, A. M.

In: Electrophoresis, Vol. 15, No. 7, 1994, p. 926-929.

Research output: Contribution to journalArticle

Csako, G. ; Nagy, B. ; Costello, R. ; Castelli, J. C. ; Hruszkewycz, A. M. / Pulsed-field gel electrophoresis for the separation of large protein molecules exemplified by the isoforms of apolipoprotein (a). In: Electrophoresis. 1994 ; Vol. 15, No. 7. pp. 926-929.
@article{0541a8157dcc4c49a54e98194f415ef1,
title = "Pulsed-field gel electrophoresis for the separation of large protein molecules exemplified by the isoforms of apolipoprotein (a)",
abstract = "The performance of pulsed-field gel electrophoresis (PFGE) was assessed for the separation of protein molecules. The allelic isoforms of apolipoprotein (a) (apo[a]) served as a model for this study because apo(a) is an unusually large protein, consisting of a variable number of repeating units. PFGE and, for comparison, conventional electrophoresis of human sera were carried out under reducing conditions in agarose gel. After blotting proteins onto nitrocellulose membrane, a combination of monospecific rabbit anti-apo(a) antibody and alkaline phosphatase-conjugated protein A detected apo(a) isoforms in all sera tested. The various apo(a) isoforms were effectively resolved within two repeating units ('kringles') by both PFGE and conventional electrophoresis, but the type of agarose gel used greatly affected the speed of electrophoretic separation. In a series of 89 human sera, 59 double-band and 30 single-band patterns were seen using either electrophoretic system. However, one specimen produced double bands with PFGE, while only a single band could be detected by conventional electrophoresis, and with another specimen the opposite occurred. A total of 22 different apo(a) isoforms were identified among these patterns. It is concluded that the increasingly available PFGE technology is a practical alternative to conventional agarose electrophoresis for the separation of large protein molecules.",
author = "G. Csako and B. Nagy and R. Costello and Castelli, {J. C.} and Hruszkewycz, {A. M.}",
year = "1994",
language = "English",
volume = "15",
pages = "926--929",
journal = "Electrophoresis",
issn = "0173-0835",
publisher = "Wiley-VCH Verlag",
number = "7",

}

TY - JOUR

T1 - Pulsed-field gel electrophoresis for the separation of large protein molecules exemplified by the isoforms of apolipoprotein (a)

AU - Csako, G.

AU - Nagy, B.

AU - Costello, R.

AU - Castelli, J. C.

AU - Hruszkewycz, A. M.

PY - 1994

Y1 - 1994

N2 - The performance of pulsed-field gel electrophoresis (PFGE) was assessed for the separation of protein molecules. The allelic isoforms of apolipoprotein (a) (apo[a]) served as a model for this study because apo(a) is an unusually large protein, consisting of a variable number of repeating units. PFGE and, for comparison, conventional electrophoresis of human sera were carried out under reducing conditions in agarose gel. After blotting proteins onto nitrocellulose membrane, a combination of monospecific rabbit anti-apo(a) antibody and alkaline phosphatase-conjugated protein A detected apo(a) isoforms in all sera tested. The various apo(a) isoforms were effectively resolved within two repeating units ('kringles') by both PFGE and conventional electrophoresis, but the type of agarose gel used greatly affected the speed of electrophoretic separation. In a series of 89 human sera, 59 double-band and 30 single-band patterns were seen using either electrophoretic system. However, one specimen produced double bands with PFGE, while only a single band could be detected by conventional electrophoresis, and with another specimen the opposite occurred. A total of 22 different apo(a) isoforms were identified among these patterns. It is concluded that the increasingly available PFGE technology is a practical alternative to conventional agarose electrophoresis for the separation of large protein molecules.

AB - The performance of pulsed-field gel electrophoresis (PFGE) was assessed for the separation of protein molecules. The allelic isoforms of apolipoprotein (a) (apo[a]) served as a model for this study because apo(a) is an unusually large protein, consisting of a variable number of repeating units. PFGE and, for comparison, conventional electrophoresis of human sera were carried out under reducing conditions in agarose gel. After blotting proteins onto nitrocellulose membrane, a combination of monospecific rabbit anti-apo(a) antibody and alkaline phosphatase-conjugated protein A detected apo(a) isoforms in all sera tested. The various apo(a) isoforms were effectively resolved within two repeating units ('kringles') by both PFGE and conventional electrophoresis, but the type of agarose gel used greatly affected the speed of electrophoretic separation. In a series of 89 human sera, 59 double-band and 30 single-band patterns were seen using either electrophoretic system. However, one specimen produced double bands with PFGE, while only a single band could be detected by conventional electrophoresis, and with another specimen the opposite occurred. A total of 22 different apo(a) isoforms were identified among these patterns. It is concluded that the increasingly available PFGE technology is a practical alternative to conventional agarose electrophoresis for the separation of large protein molecules.

UR - http://www.scopus.com/inward/record.url?scp=0028147841&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0028147841&partnerID=8YFLogxK

M3 - Article

C2 - 7813396

AN - SCOPUS:0028147841

VL - 15

SP - 926

EP - 929

JO - Electrophoresis

JF - Electrophoresis

SN - 0173-0835

IS - 7

ER -