Product namePyruvate dehydrogenase (PDH) Protein Quantity Dipstick Assay Kit
See all Pyruvate dehydrogenase kits
Sample typeCell culture extracts, Tissue
Assay typeSandwich (quantitative)
Species reactivityReacts with: Mouse, Rat, Cow, Human
ab109883 (MSP31) is used to quantify the activity of the PDH enzyme complex (pyruvate dehydrogenase) from human, bovine, mouse, and rat samples.
PDH is present in all tissues; it plays a central role in metabolism as it is a key regulatory enzyme that functions at the junction between glycolysis and the tricarboxylic acid cycle. The PDH complex is composed of multiple copies of three catalytic component enzymes; pyruvate dehydrogenase or E1 (EC 220.127.116.11), dihydrolipoamide transacetylase or E2 (EC 18.104.22.168) and dihydrolipoamide dehydrogenase or E3 (EC 22.214.171.124). Inherent to its regulatory function, a number of other proteins regulate PDH activity. One of these, dihydrolipoamide dehydrogenase-binding protein (E3Bp) is necessary for the interaction of the E2 and E3 components.
This dipstick assay kit utilizes two monoclonal antibodies. One recognizes the E2 enzyme (monoclonal antibody bound to the dipstick) and the other antibody (gold-conjugated) recognizes both the E2 and E3Bp components of the PDH complex. The signal intensity is measured by a dipstick reader or analyzed by other imaging systems such as a flatbed scanner. The kit is compatible with a variety of sample types (tissue or cell culture) from a number of different species (human, bovine, mouse, and rat).
All components are stable in their provided containers at room temperature out of direct sunlight.
After diluting the 10X Blocking Buffer to 2X, store at 4°C.
For long-term storage, all buffers can be stored at 4°C.
Tested applicationsSuitable for: Sandwich ELISAmore details
Storage instructionsStore at +4°C. Please refer to protocols.
Components 90 tests 30 tests Buffer B (10X Blocking solution) 3 x 0.4ml 1 x 0.4ml Dipsticks 1 x 90 units 1 x 30 units Extraction Buffer (ab260490) 3 x 15ml 1 x 15ml Gold-conjugated antibody (dried in microplate wells) 1 x 90 tests 1 x 30 tests Wash buffer 3 x 2ml 1 x 2ml
Our Abpromise guarantee covers the use of ab109883 in the following tested applications.
The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
|Sandwich ELISA||Use at an assay dependent dilution.|
An example using ab109883 to quantify PDH levels using various concentration of rat tissue protein extract. Based on the above standard curve values, 18 µg (control - undiluted), 9 µg (sample A - diluted 1:2) and 1.8 µg (sample B - diluted 1:10) of extract were loaded to each dipstick. Two dipsticks per sample were run (typical intra-assay precision is 90-98%). Using GraphPad software, the signal intensity from the standard curve was interpolated and the quantity of PDH in samples A and B was determined. Based on the above analysis, the unknown samples, A and B, had 48% and 9% of control PDH levels, respectively.
An example using ab109883 to quantify PDH levels using various concentration of rat tissue protein extract. Shown is a 1:2 dilution series using a positive control sample. Approximately 7 to 8 dipsticks are suitable for covering the entire working range and the blank for background levels. In this example the dilution series starts with 20 µg of rat heart tissue extract. A one-site hyperbola line was generated for best-fit analysis using GraphPad.
Dipstick assays use the well-established lateral flow concept, whereby capture antibodies are striped onto nitrocellulose membrane and a Whatman paper wicking pad draws the sample through the antibody bands. Detector antibodies, conjugated to gold, are dried in the wells of a 96-well plate. Sample is added to the well, the dipstick inserted, and within minutes the line for each target is revealed as the protein-detector antibody-gold complex binds with the capture antibodies. Multiplexing dipstick assays have multiple target protein lines. A positive control goat anti-mouse antibody line is included on all assays to ensure that adequate wicking of the sample occurred.
ab109883 has not yet been referenced specifically in any publications.