The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal

Louise Bjerremann Jensen; Anna Maria Torp; Sven Bode Andersen; Per Stahl Skov; Lars K. Poulsen; Edward F. Knol; Els van Hoffen

doi:10.1016/j.jim.2008.02.012

The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal

Louise Bjerremann Jensen, Anna Maria Torp, Sven Bode Andersen, Per Stahl Skov, Lars K. Poulsen, Edward F. Knol, Els van Hoffen

Department of Clinical Medicine

20 Citations (Scopus)

Abstract

The application of recombinant (His)(6)-tagged proteins in cell culture assays is associated with problems due to lipopolysaccharide (LPS) contamination. LPS stimulates cells of the immune system, thereby masking antigen-specific activation of T cells. Due to the affinity of LPS for histidine it is associated with difficulties to remove LPS from recombinant (His)(6)-tagged proteins. Here we describe that the Triton X-114 phase separation method can be used to remove LPS from (His)(6)-tagged proteins and that the recombinant proteins retain their biological activity.

Original language	English
Journal	Journal of Immunological Methods
Volume	335
Issue number	1-2
Pages (from-to)	116-20
Number of pages	5
ISSN	0022-1759
DOIs	https://doi.org/10.1016/j.jim.2008.02.012
Publication status	Published - 2008

Keywords

Allergens
Antigens, Plant
Basophils
Cells, Cultured
Cloning, Molecular
Dose-Response Relationship, Drug
Glycoproteins
Histamine Release
Histidine
Humans
Interleukin-10
Leukocytes, Mononuclear
Lipopolysaccharides
Plant Proteins
Polyethylene Glycols
Protein Binding
Recombinant Proteins
T-Lymphocytes
Time Factors
Tumor Necrosis Factor-alpha

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10.1016/j.jim.2008.02.012

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title = "The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal",

abstract = "The application of recombinant (His)(6)-tagged proteins in cell culture assays is associated with problems due to lipopolysaccharide (LPS) contamination. LPS stimulates cells of the immune system, thereby masking antigen-specific activation of T cells. Due to the affinity of LPS for histidine it is associated with difficulties to remove LPS from recombinant (His)(6)-tagged proteins. Here we describe that the Triton X-114 phase separation method can be used to remove LPS from (His)(6)-tagged proteins and that the recombinant proteins retain their biological activity.",

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author = "Jensen, {Louise Bjerremann} and Torp, {Anna Maria} and Andersen, {Sven Bode} and Skov, {Per Stahl} and Poulsen, {Lars K.} and Knol, {Edward F.} and {van Hoffen}, Els",

year = "2008",

doi = "10.1016/j.jim.2008.02.012",

language = "English",

volume = "335",

pages = "116--20",

journal = "Journal of Immunological Methods",

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publisher = "Elsevier",

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T1 - The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal

AU - Jensen, Louise Bjerremann

AU - Torp, Anna Maria

AU - Andersen, Sven Bode

AU - Skov, Per Stahl

AU - Poulsen, Lars K.

AU - Knol, Edward F.

AU - van Hoffen, Els

PY - 2008

Y1 - 2008

N2 - The application of recombinant (His)(6)-tagged proteins in cell culture assays is associated with problems due to lipopolysaccharide (LPS) contamination. LPS stimulates cells of the immune system, thereby masking antigen-specific activation of T cells. Due to the affinity of LPS for histidine it is associated with difficulties to remove LPS from recombinant (His)(6)-tagged proteins. Here we describe that the Triton X-114 phase separation method can be used to remove LPS from (His)(6)-tagged proteins and that the recombinant proteins retain their biological activity.

AB - The application of recombinant (His)(6)-tagged proteins in cell culture assays is associated with problems due to lipopolysaccharide (LPS) contamination. LPS stimulates cells of the immune system, thereby masking antigen-specific activation of T cells. Due to the affinity of LPS for histidine it is associated with difficulties to remove LPS from recombinant (His)(6)-tagged proteins. Here we describe that the Triton X-114 phase separation method can be used to remove LPS from (His)(6)-tagged proteins and that the recombinant proteins retain their biological activity.

KW - Allergens

KW - Antigens, Plant

KW - Basophils

KW - Cells, Cultured

KW - Cloning, Molecular

KW - Dose-Response Relationship, Drug

KW - Glycoproteins

KW - Histamine Release

KW - Histidine

KW - Humans

KW - Interleukin-10

KW - Leukocytes, Mononuclear

KW - Lipopolysaccharides

KW - Plant Proteins

KW - Polyethylene Glycols

KW - Protein Binding

KW - Recombinant Proteins

KW - T-Lymphocytes

KW - Time Factors

KW - Tumor Necrosis Factor-alpha

U2 - 10.1016/j.jim.2008.02.012

DO - 10.1016/j.jim.2008.02.012

M3 - Journal article

C2 - 18377922

SN - 0022-1759

VL - 335

SP - 116

EP - 120

JO - Journal of Immunological Methods

JF - Journal of Immunological Methods

IS - 1-2

ER -

The biological activity of a recombinantly expressed (His)(6)-tagged peanut allergen (rAra h 1) is unaffected by endotoxin removal

Abstract

Keywords

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