Specific detection and semi-quantitative analysis of TRPC4 protein expression by antibodies.
In mouse tissues two variants of the transient receptor potential (canonical) (TRPC) 4 protein are expressed: the "full-length" TRPC4 protein and a slightly smaller variant, called TRPC4Delta(761-864), which lacks 84 amino acid residues. Although the presence of mRNA encoding the TRPC4 protein in mammalian cells and the detection of the heterologously expressed TRPC4 protein by Western blot analysis have been reported, the unequivocal detection of endogenous TRPC4 proteins has proven difficult. In the present study we compared polyclonal antibodies for the detection of TRPC4 proteins in mouse tissues and monitored their specificity and reliability by analysing corresponding tissues from TRPC4-deficient mice. In addition we introduced a procedure that allows us to estimate the amount of TRPC4 protein expressed in a single cell. Using this technique it appears that the amount of TRPC4 protein expressed stably in HEK 293 cells is at least fourfold higher than the amount of TRPC4 protein expressed endogenously in the bovine adrenocortical cell line SBAC.
Flockerzi
Veit
V
Experimentelle und Klinische Pharmakologie und Toxikologie, Medizinische Fakultät, Universität des Saarlandes, 66421 Homburg, Deutschland. veit.flockerzi@uniklinik-saarland.de
Jung
Christine
C
Aberle
Thomas
T
Meissner
Marcel
M
Freichel
Marc
M
Philipp
Stephan E
SE
Nastainczyk
Wolfgang
W
Maurer
Patrick
P
Zimmermann
Richard
R
D000818:Animals; D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002474:Cell-Free System; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D051379:Mice; D018482:Muscle, Skeletal / Q000378:metabolism; D050052:TRPC Cation Channels / Q000032:analysis* / Q000235:genetics / Q000276:immunology
D050052:TRPC Cation Channels; C489571:TRPC4 ion channel