A method for incorporating macromolecules into adherent cells

P. L. McNeil, R. F. Murphy, F. Lanni, D. L. Taylor

Research output: Contribution to journalArticle

215 Citations (Scopus)

Abstract

We describe a simple method for loading exogenous macromolecules into the cytoplasm of mammalian cells adherent to tissue culture dishes. Culture medium was replaced with a thin layer of fluorescently labeled macromolecules, the cells were harvested from the substrate by scraping with a rubber policeman, transferred immediately to ice cold media, washed, and then replated for culture. We refer to the method as 'scrape-loading.' Viability of cells was 50-60% immediately after scrape-loading and was 90% for those cells remaining after 24 h of culture. About 40% of adherent, well-spread fibroblasts contained fluorescent molecules 18 h after scrape-loading of labeled dextrans, ovalbumin, or immunoglobulin-G. On average, 107 dextran molecules (70,000-mol wt) were incorporated into each fibroblast by scrape-loading in 10 mg/ml dextran. The extent of loading depended on the concentration and molecular weight of the dextrans used. A fluorescent analog of actin could also be loaded into fibroblasts where it labeled stress fibers. HeLa cells, a macrophage-like cell line, J774A.1, and human neutrophils were all successfully loaded with dextran by scraping. The method of scrape-loading should be applicable to a broad range of adherent cell types, and useful for loading of diverse kinds of macromolecules.

Original languageEnglish (US)
Pages (from-to)1556-1564
Number of pages9
JournalJournal of Cell Biology
Volume98
Issue number4
DOIs
StatePublished - Jan 1 1984

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Dextrans
Fibroblasts
Stress Fibers
Rubber
Ice
HeLa Cells
Culture Media
Actins
Cell Survival
Cytoplasm
Neutrophils
Immunoglobulin G
Molecular Weight
Macrophages
Cell Line

ASJC Scopus subject areas

  • Cell Biology

Cite this

McNeil, P. L., Murphy, R. F., Lanni, F., & Taylor, D. L. (1984). A method for incorporating macromolecules into adherent cells. Journal of Cell Biology, 98(4), 1556-1564. https://doi.org/10.1083/jcb.98.4.1556

A method for incorporating macromolecules into adherent cells. / McNeil, P. L.; Murphy, R. F.; Lanni, F.; Taylor, D. L.

In: Journal of Cell Biology, Vol. 98, No. 4, 01.01.1984, p. 1556-1564.

Research output: Contribution to journalArticle

McNeil, PL, Murphy, RF, Lanni, F & Taylor, DL 1984, 'A method for incorporating macromolecules into adherent cells', Journal of Cell Biology, vol. 98, no. 4, pp. 1556-1564. https://doi.org/10.1083/jcb.98.4.1556
McNeil, P. L. ; Murphy, R. F. ; Lanni, F. ; Taylor, D. L. / A method for incorporating macromolecules into adherent cells. In: Journal of Cell Biology. 1984 ; Vol. 98, No. 4. pp. 1556-1564.
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