TY - JOUR
T1 - [17] Preparation of Semiintact Cells for Study of Vesicular Trafficking in Vitro
AU - Pind, S.
AU - Davidson, H.
AU - Schwaninger, R.
AU - Beckers, C. J M
AU - Plutner, H.
AU - Schmid, S. L.
AU - Balch, W. E.
PY - 1993/1/1
Y1 - 1993/1/1
N2 - This chapter focuses on techniques used in the laboratory to reconstitute endoplasmic reticulum (ER)-to-Golgi transport in vitro, using adherent tissue culture cells that have been rendered semiintact by gently scraping them from the dish after hypotonic swelling. This technique is discussed in the context of other protocols that allow the investigator to perforate both adherent and nonadherent cells. The ER-to-Golgi transport assay is based on the observation that individual N-linked oligosaccharide-processing enzymes reside in distinct subcellular compartments.Potential markers include endogenous proteins, other viral glycoproteins, and proteins acquired through transfection. Moreover, because both the interior and exterior membranes of the cell are jointly accessible to a wide range of reagents and macromolecules, semiintact cells may provide a useful model system for the study of a broad range of problems in cell biology, including signal transduction, organization of the cytoskeletal matrix, and gene activation.
AB - This chapter focuses on techniques used in the laboratory to reconstitute endoplasmic reticulum (ER)-to-Golgi transport in vitro, using adherent tissue culture cells that have been rendered semiintact by gently scraping them from the dish after hypotonic swelling. This technique is discussed in the context of other protocols that allow the investigator to perforate both adherent and nonadherent cells. The ER-to-Golgi transport assay is based on the observation that individual N-linked oligosaccharide-processing enzymes reside in distinct subcellular compartments.Potential markers include endogenous proteins, other viral glycoproteins, and proteins acquired through transfection. Moreover, because both the interior and exterior membranes of the cell are jointly accessible to a wide range of reagents and macromolecules, semiintact cells may provide a useful model system for the study of a broad range of problems in cell biology, including signal transduction, organization of the cytoskeletal matrix, and gene activation.
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U2 - 10.1016/0076-6879(93)21019-5
DO - 10.1016/0076-6879(93)21019-5
M3 - Article
C2 - 8395634
AN - SCOPUS:0027199247
SN - 0076-6879
VL - 221
SP - 222
EP - 234
JO - Methods in Enzymology
JF - Methods in Enzymology
IS - C
ER -