Simplified purification procedure of laminin-332 and laminin-511 from human cell lines

Isis C. Sroka; Man Ling Chen; Anne E. Cress

doi:10.1016/j.bbrc.2008.08.029

Simplified purification procedure of laminin-332 and laminin-511 from human cell lines

Isis C. Sroka, Man Ling Chen, Anne E. Cress

Research output: Contribution to journal › Article › peer-review

11 Scopus citations

Abstract

Laminins are glycoproteins expressed in the basement membrane of multiple epithelial tissues. Previously described purification procedures for the human laminin variants laminin-5 (LN-332) and laminin-10 (LN-511) use tissue as starting material and have multiple steps. We demonstrate a two-step laminin immunoaffinity purification method to produce consistent quantities of intact and biologically active LN-332 and LN-511 from human keratinocyte (HaCaT) and human lung carcinoma (A549) cell lines, respectively. The purification of LN-332 and LN-551 was demonstrated by PAGE analysis, silver staining and Western blot analysis. The purification procedure includes instruction on removing a cell adhesion contaminant known as galectin-3 binding protein from purified LN-511. The biological activity of purified laminin was tested in a standard cell adhesion assay and compared to commercially available LN-111. This rapid and reproducible purification method will contribute to understanding the role of LN-332 and LN-511 in cell behavior, signaling, and gene expression.

Original language	English (US)
Pages (from-to)	410-413
Number of pages	4
Journal	Biochemical and Biophysical Research Communications
Volume	375
Issue number	3
DOIs	https://doi.org/10.1016/j.bbrc.2008.08.029
State	Published - Oct 24 2008

Keywords

Laminin-322
Laminin-511
Purification

ASJC Scopus subject areas

Biophysics
Biochemistry
Molecular Biology
Cell Biology

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10.1016/j.bbrc.2008.08.029

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title = "Simplified purification procedure of laminin-332 and laminin-511 from human cell lines",

abstract = "Laminins are glycoproteins expressed in the basement membrane of multiple epithelial tissues. Previously described purification procedures for the human laminin variants laminin-5 (LN-332) and laminin-10 (LN-511) use tissue as starting material and have multiple steps. We demonstrate a two-step laminin immunoaffinity purification method to produce consistent quantities of intact and biologically active LN-332 and LN-511 from human keratinocyte (HaCaT) and human lung carcinoma (A549) cell lines, respectively. The purification of LN-332 and LN-551 was demonstrated by PAGE analysis, silver staining and Western blot analysis. The purification procedure includes instruction on removing a cell adhesion contaminant known as galectin-3 binding protein from purified LN-511. The biological activity of purified laminin was tested in a standard cell adhesion assay and compared to commercially available LN-111. This rapid and reproducible purification method will contribute to understanding the role of LN-332 and LN-511 in cell behavior, signaling, and gene expression.",

keywords = "Laminin-322, Laminin-511, Purification",

author = "Sroka, {Isis C.} and Chen, {Man Ling} and Cress, {Anne E.}",

note = "Funding Information: The work was supported in part by CA56666, CA23074. The expert technical assistance of the mass spectrometry service staff was much appreciated. The contribution of the monoclonal antibodies and their careful characterization by several other investigators was essential for the successful completion of the work.",

year = "2008",

month = oct,

day = "24",

doi = "10.1016/j.bbrc.2008.08.029",

language = "English (US)",

volume = "375",

pages = "410--413",

journal = "Biochemical and Biophysical Research Communications",

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AU - Sroka, Isis C.

AU - Chen, Man Ling

AU - Cress, Anne E.

N1 - Funding Information: The work was supported in part by CA56666, CA23074. The expert technical assistance of the mass spectrometry service staff was much appreciated. The contribution of the monoclonal antibodies and their careful characterization by several other investigators was essential for the successful completion of the work.

PY - 2008/10/24

Y1 - 2008/10/24

N2 - Laminins are glycoproteins expressed in the basement membrane of multiple epithelial tissues. Previously described purification procedures for the human laminin variants laminin-5 (LN-332) and laminin-10 (LN-511) use tissue as starting material and have multiple steps. We demonstrate a two-step laminin immunoaffinity purification method to produce consistent quantities of intact and biologically active LN-332 and LN-511 from human keratinocyte (HaCaT) and human lung carcinoma (A549) cell lines, respectively. The purification of LN-332 and LN-551 was demonstrated by PAGE analysis, silver staining and Western blot analysis. The purification procedure includes instruction on removing a cell adhesion contaminant known as galectin-3 binding protein from purified LN-511. The biological activity of purified laminin was tested in a standard cell adhesion assay and compared to commercially available LN-111. This rapid and reproducible purification method will contribute to understanding the role of LN-332 and LN-511 in cell behavior, signaling, and gene expression.

AB - Laminins are glycoproteins expressed in the basement membrane of multiple epithelial tissues. Previously described purification procedures for the human laminin variants laminin-5 (LN-332) and laminin-10 (LN-511) use tissue as starting material and have multiple steps. We demonstrate a two-step laminin immunoaffinity purification method to produce consistent quantities of intact and biologically active LN-332 and LN-511 from human keratinocyte (HaCaT) and human lung carcinoma (A549) cell lines, respectively. The purification of LN-332 and LN-551 was demonstrated by PAGE analysis, silver staining and Western blot analysis. The purification procedure includes instruction on removing a cell adhesion contaminant known as galectin-3 binding protein from purified LN-511. The biological activity of purified laminin was tested in a standard cell adhesion assay and compared to commercially available LN-111. This rapid and reproducible purification method will contribute to understanding the role of LN-332 and LN-511 in cell behavior, signaling, and gene expression.

KW - Laminin-322

KW - Laminin-511

KW - Purification

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ER -

Simplified purification procedure of laminin-332 and laminin-511 from human cell lines

Abstract

Keywords

ASJC Scopus subject areas

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