Western blot analysis of extracts from various cell lines using CTPS1 Antibody.
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised June 2016
Protocol Id: 263
CTPS1 Antibody recognizes endogenous levels of total CTPS1 protein. This antibody does not cross-react with CTPS2 protein.Species Reactivity:
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Ala340 of human CTPS1 protein. Antibodies are purified by peptide affinity chromatography.
CTPS1 (cytidine triphosphate synthase 1 or CTP synthase 1) catalyzes the conversion of UTP to CTP through ATP-dependent amination (1). Research studies show that pyrimidine biosynthesis is upregulated in pancreatic cancer cell models of resistance to gemcitabine, a deoxycytidine analog. Increased pyrimidine biosynthesis leads to elevated levels of endogenous dCTP, which outcompetes gemcitabine from incorporating into DNA during replication and, therefore, reduces the anti-cancer effectiveness of gemcitabine. Induction of CTPS1 expression was observed in gemcitabine-resistant pancreatic cancer cell models. Gemcitabine resistance directly correlates with CTPS1 expression levels in various pancreatic cancer cell lines (2). Furthermore, CTPS1 expression was shown to increase in T cells upon T-cell receptor activation, suggesting a critical role of CTPS1 in the immune system (3).
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