Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography.

G. Aboagye-Mathiesen, F. Tóth, C. Juhl, N. Nørskov-Lauritsen, P. M. Petersen, P. Ebbesen

Research output: Article

6 Citations (Scopus)

Abstract

Human placental trophoblast challenged with Sendai virus induced IFNs mainly of the beta-type (75%) and relatively low levels of the alpha-type (25%). A two-step high performance liquid chromatographic procedure ("two-dimensional HPLC") has been developed for the complete purification of the placental trophoblast interferon beta (tro-IFN-beta) from serum-containing culture supernatant. The method involved a combination of high performance liquid affinity chromatography (HPLAC) on Cibacron Blue 3GA immobilized on an activated pressure stable macroporous synthetic polymer, 2-hydroxyethyl methacrylate vinyl sulphone (HEMA-BIO 1000 VS), as the first dimension and reversed-phase high performance liquid chromatography (RP-HPLC) on Separon SGX C-18 as the second. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunoblot experiments showed that the tro-IFN-beta was present as a 24 kDa protein. Densitometric scanning analysis of Coomassie-stained gel revealed the purity of the final preparation to be greater than 99%. The purified tro-IFN-beta had a specific activity of 1.03 x 10(8) IU/mg of protein and the overall recovery was 81% of the total IFN-beta activity in the crude preparation and 61% of the total IFN activity.

Original languageEnglish
Pages (from-to)35-51
Number of pages17
JournalPreparative Biochemistry
Volume21
Issue number1
Publication statusPublished - 1991

Fingerprint

Interferon-beta
Trophoblasts
High performance liquid chromatography
Interferons
Purification
High Pressure Liquid Chromatography
Affinity chromatography
Liquid chromatography
Electrophoresis
Viruses
Sendai virus
Sodium Dodecyl Sulfate
Polymers
Proteins
Reverse-Phase Chromatography
Gels
Affinity Chromatography
Scanning
Polyacrylamide Gel Electrophoresis
Recovery

ASJC Scopus subject areas

  • Biochemistry
  • Genetics

Cite this

Aboagye-Mathiesen, G., Tóth, F., Juhl, C., Nørskov-Lauritsen, N., Petersen, P. M., & Ebbesen, P. (1991). Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography. Preparative Biochemistry, 21(1), 35-51.

Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography. / Aboagye-Mathiesen, G.; Tóth, F.; Juhl, C.; Nørskov-Lauritsen, N.; Petersen, P. M.; Ebbesen, P.

In: Preparative Biochemistry, Vol. 21, No. 1, 1991, p. 35-51.

Research output: Article

Aboagye-Mathiesen, G, Tóth, F, Juhl, C, Nørskov-Lauritsen, N, Petersen, PM & Ebbesen, P 1991, 'Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography.', Preparative Biochemistry, vol. 21, no. 1, pp. 35-51.
Aboagye-Mathiesen, G. ; Tóth, F. ; Juhl, C. ; Nørskov-Lauritsen, N. ; Petersen, P. M. ; Ebbesen, P. / Purification of human placental trophoblast interferon by two-dimensional high performance liquid chromatography. In: Preparative Biochemistry. 1991 ; Vol. 21, No. 1. pp. 35-51.
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