Functional knockdown of VCAM-1 at the posttranslational level with ER retained antibodies
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CONTRIBUTORS:
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JOURNAL:
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J Immunol Methods,
341(1-2),
30 -
40.
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YEAR:
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2009
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PUB TYPE:
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Journal Article
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SUBJECT(S):
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None
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DISCIPLINE:
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No discipline assigned
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HTTP:
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LANGUAGE:
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None
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PUB ID:
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103-447-867
(Last edited on
2009/02/18 23:43:32 US/Mountain)
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SPONSOR(S):
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ABSTRACT:
Vascular cell adhesion molecule 1 (VCAM-1) is involved in the recruitment of leukocytes to inflammatory sites. In this study we present the first functional knockdown of VCAM-1 using an ER retained antibody construct. We generated a knockdown construct encoding the VCAM-1 specific single chain variable fragment scFv6C7.1 fused to the C-terminal ER retention sequence KDEL. HEK-293:VCAM-YFP cells stably expressing a VCAM-YFP fusion protein were transiently transfected with the knockdown construct and showed down-regulation of surface VCAM-1. Knockdown efficiency of the system is time-dependent due to used transient transfection of the intrabody construct. Furthermore, intrabody mediated knockdown of HEK-293:VCAM-YFP cells also impaired cell-cell interaction with Jurkat cells that are endogenously expressing VLA-4, the physiological partner of VCAM-1. Posttranslational knockdown with ER retained antibodies seems to be a promising technique, as shown here for VCAM-1.
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