Abstract
The stability of nicotinic acetylcholine receptors (AChR) solubilized from mammalian skeletal muscle in nonionic detergent was investigated under various conditions of pH, chaotropic ions, and unfolding reagents in order to allow its purification in high yield by immunoadsorption to monoclonal antibodies. Preservation of the antigenicity and/or binding sites for α-bungarotoxin was used as an indicator of the receptor protein's integrity. Both were preserved in the pH range 6.5-8.0, but when exposed for 1 h at 4°C to a pH outside this range, >50% activity was lost. Of the chaotropic ions studied (NaSCN, NaI, NaNO3, NaCl), only NaCl was tolerated. Most of the AChR's toxin-binding activity was preserved after exposure to 2 M NaCl, which was suitable for dissociating AChR when a monoclonal antibody with relatively low binding affinity was selected as the immunoadsorbent. Yields of purified AChR were optimal (30%) when a low amount of monoclonal antibody was coupled to cyanogen bromide-activated agarose (1 mg protein/ml gel).
| Original language | English (US) |
|---|---|
| Pages (from-to) | 59-64 |
| Number of pages | 6 |
| Journal | Journal of Neurochemistry |
| Volume | 42 |
| Issue number | 1 |
| State | Published - 1984 |
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ASJC Scopus subject areas
- Biochemistry
- Cellular and Molecular Neuroscience
Cite this
The stability of solubilized mammalian muscle acetylcholine receptors during purification by monoclonal immunoadsorption. / Momoi, M. Y.; Lennon, Vanda A.
In: Journal of Neurochemistry, Vol. 42, No. 1, 1984, p. 59-64.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - The stability of solubilized mammalian muscle acetylcholine receptors during purification by monoclonal immunoadsorption
AU - Momoi, M. Y.
AU - Lennon, Vanda A
PY - 1984
Y1 - 1984
N2 - The stability of nicotinic acetylcholine receptors (AChR) solubilized from mammalian skeletal muscle in nonionic detergent was investigated under various conditions of pH, chaotropic ions, and unfolding reagents in order to allow its purification in high yield by immunoadsorption to monoclonal antibodies. Preservation of the antigenicity and/or binding sites for α-bungarotoxin was used as an indicator of the receptor protein's integrity. Both were preserved in the pH range 6.5-8.0, but when exposed for 1 h at 4°C to a pH outside this range, >50% activity was lost. Of the chaotropic ions studied (NaSCN, NaI, NaNO3, NaCl), only NaCl was tolerated. Most of the AChR's toxin-binding activity was preserved after exposure to 2 M NaCl, which was suitable for dissociating AChR when a monoclonal antibody with relatively low binding affinity was selected as the immunoadsorbent. Yields of purified AChR were optimal (30%) when a low amount of monoclonal antibody was coupled to cyanogen bromide-activated agarose (1 mg protein/ml gel).
AB - The stability of nicotinic acetylcholine receptors (AChR) solubilized from mammalian skeletal muscle in nonionic detergent was investigated under various conditions of pH, chaotropic ions, and unfolding reagents in order to allow its purification in high yield by immunoadsorption to monoclonal antibodies. Preservation of the antigenicity and/or binding sites for α-bungarotoxin was used as an indicator of the receptor protein's integrity. Both were preserved in the pH range 6.5-8.0, but when exposed for 1 h at 4°C to a pH outside this range, >50% activity was lost. Of the chaotropic ions studied (NaSCN, NaI, NaNO3, NaCl), only NaCl was tolerated. Most of the AChR's toxin-binding activity was preserved after exposure to 2 M NaCl, which was suitable for dissociating AChR when a monoclonal antibody with relatively low binding affinity was selected as the immunoadsorbent. Yields of purified AChR were optimal (30%) when a low amount of monoclonal antibody was coupled to cyanogen bromide-activated agarose (1 mg protein/ml gel).
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UR - http://www.scopus.com/inward/citedby.url?scp=0021368298&partnerID=8YFLogxK
M3 - Article
C2 - 6358416
AN - SCOPUS:0021368298
VL - 42
SP - 59
EP - 64
JO - Journal of Neurochemistry
JF - Journal of Neurochemistry
SN - 0022-3042
IS - 1
ER -