Description
Oxaloacetate Assay Kit (Colorimetric/Fluorometric) (BA0139) (BA0139)
The Oxaloacetate Assay Kit (Colorimetric/Fluorometric) (SKU: BA0139) provides a simple, direct and automation-ready procedure for measuring oxaloacetate concentration in biological samples. Oxaloacetate (OAA) is an intermediate in the citric acid cycle and participates in gluconeogenesis, being formed by the oxidation of malate, the deamidation of aspartate or the condensation of carbon dioxide with pyruvate or phosphoenolpyruvate. In this assay, oxaloacetate is converted into pyruvate which is then oxidised with the conversion of the dye into a coloured and fluorescent form. The colour intensity of the oxidised dye at 570 nm or fluorescence intensity at 530/585 nm is directly proportional to the oxaloacetate concentration in the sample. The linear detection range is 7 to 400 uM oxaloacetate for colorimetric assays and 1 to 40 uM for fluorometric assays.
| Product Name: | Oxaloacetate Assay Kit (Colorimetric/Fluorometric) (BA0139) |
| SKU: | BA0139 |
| Detection Method: | Colorimetric/Fluorometric |
| Detection Range: | 7 to 400 uM (colorimetric); 1 to 40 uM (fluorometric) |
| Sample Type: | Plasma, serum, tissue and culture media |
| Species Reactivity: | All |
| Assay Time: | 15 minutes |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | -20 degrees C |
| Shelf Life: | 6 months after receipt |
| Shipping: | Gel Pack |
This kit measures oxaloacetate by converting it into pyruvate, which is then oxidised with the conversion of the dye into a coloured and fluorescent form. The colour intensity of the oxidised dye at 570 nm or fluorescence intensity at 530/585 nm is directly proportional to the oxaloacetate concentration in the sample.
- Sensitive and accurate with a linear detection range of 7 to 400 uM oxaloacetate for colorimetric assays and 1 to 40 uM for fluorometric assays
- Simple, direct and automation-ready procedure
- Provides both colorimetric and fluorometric detection formats with a short 15 minute incubation
- Direct measurement of oxaloacetate in plasma, serum, tissue and culture media
Note: The below protocol is a sample protocol. Protocols are specific to each batch/lot. For the correct instructions please follow the protocol included in your kit.
| Step | Procedure |
| 1 | Dissolve the Oxaloacetate Standard with 100 uL distilled water to make a 10 mM stock (used within 2 weeks and stored at -20 degrees C), prepare a 400 uM Premix by diluting 20 uL of the 10 mM standard with 480 uL distilled water, then dilute the standards as described in the dilution table and transfer 20 uL of each standard to separate wells of a clear flat-bottom 96-well plate. |
| 2 | Add 20 uL of each sample to two separate wells (each sample requires a sample blank); samples requiring an internal standard need three reactions (sample plus standard, sample alone and sample blank). |
| 3 | Prepare enough Working Reagent for all standards and samples by combining, per reaction, 85 uL Developer, 1 uL ODC Enzyme and 1 uL Dye Reagent; for the sample blanks prepare a Working Reagent without the ODC Enzyme. |
| 4 | Add 80 uL of the appropriate Working Reagent to each standard and sample well, mix well and incubate protected from light for 15 minutes at room temperature. |
| 5 | Read OD570nm, or for the fluorometric procedure dilute standards 1:10 in distilled water, use a black 96-well plate and read fluorescence at 530/585 nm after a 15 minute incubation at room temperature. |
Subtract the blank value from the standard values and plot the delta OD or delta F against standard concentrations to determine the slope, then calculate [Oxaloacetate] = (RSAMPLE - RBLANK) / [Slope (uM-1)] x n (uM). If an internal standard was used, [Oxaloacetate] = (RSAMPLE - RBLANK) / (RSTANDARD - RSAMPLE) x 20 (uM). Conversions: 100 uM oxaloacetate equals 13.1 mg/L, 0.00131% or 13.1 ppm.
| Component | Quantity | Storage |
| Developer | 10 mL | -20 degrees C |
| ODC Enzyme | 120 uL | -20 degrees C |
| Dye Reagent | 120 uL | -20 degrees C |
| Oxaloacetate Standard (Dried) | 1 tube | -20 degrees C |
