TY - JOUR
T1 - Toxin — Antitoxin Selection for Isolating Heterokaryons and Cell Hybrids
AU - Wright, W. E.
N1 - Funding Information:
This research was supported in part by the National Cancer Institute (CA40065), the American Cancer Society (CD-347), and The Council for Tobacco Research-U.S.A., Inc. I thank my former students, M. A. Clark, C. Walker, and L. Weide for their help inthis work.
PY - 1987/1
Y1 - 1987/1
N2 - This chapter discusses the methods used in toxin–antitoxin selection for isolating heterokaryons and cell hybrids. The behaviors of cell hybrids have employed genetic selection techniques, such as the HAT system, to isolate cell fusion products. Some “half-selective” strategies are devised where an established immortal cell line is fused to a normal diploid cell type which does not divide in culture, in which it is unnecessary to select against the non dividing normal diploid parental cell type. However, the inability to use genetic selection techniques with normal cells frequently represents a major limitation for many types of studies. Two alternative systems for isolating heterokaryons and cell hybrids developed are discussed in the chapter. The first one exploits the different specificities of two irreversible biochemical inhibitors. Iodoacetamide is directed primarily toward sulfhydryl groups, while diethyl pyrocarbonate reacts mainly with histidine residues. An elegant technique for introducing macromolecules into cells is introduced. Cells are first exposed to a hypenonic sucrose solution containing the molecule. Normally, the pinocytic vesicles that are formed during this period would be transported to the lysosomes, where their contents would be degraded.
AB - This chapter discusses the methods used in toxin–antitoxin selection for isolating heterokaryons and cell hybrids. The behaviors of cell hybrids have employed genetic selection techniques, such as the HAT system, to isolate cell fusion products. Some “half-selective” strategies are devised where an established immortal cell line is fused to a normal diploid cell type which does not divide in culture, in which it is unnecessary to select against the non dividing normal diploid parental cell type. However, the inability to use genetic selection techniques with normal cells frequently represents a major limitation for many types of studies. Two alternative systems for isolating heterokaryons and cell hybrids developed are discussed in the chapter. The first one exploits the different specificities of two irreversible biochemical inhibitors. Iodoacetamide is directed primarily toward sulfhydryl groups, while diethyl pyrocarbonate reacts mainly with histidine residues. An elegant technique for introducing macromolecules into cells is introduced. Cells are first exposed to a hypenonic sucrose solution containing the molecule. Normally, the pinocytic vesicles that are formed during this period would be transported to the lysosomes, where their contents would be degraded.
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U2 - 10.1016/S0076-6879(87)51021-7
DO - 10.1016/S0076-6879(87)51021-7
M3 - Article
C2 - 3501529
AN - SCOPUS:0023613418
SN - 0076-6879
VL - 151
SP - 237
EP - 252
JO - Methods in Enzymology
JF - Methods in Enzymology
IS - C
ER -