Description
Creatine Assay Kit (Colorimetric/Fluorometric) (BA0095) (BA0095)
The Creatine Assay Kit (Colorimetric/Fluorometric) (SKU: BA0095) provides a simple, direct and automation-ready procedure for measuring creatine. Creatine is present in vertebrates and helps to supply energy to muscle; approximately half of creatine originates from food, and creatine supplementation has been investigated for the treatment of muscular, neuromuscular, neurological and neurodegenerative diseases. This assay is based on enzymatic reactions leading to formation of a pink coloured product. The optical density at 570 nm, or fluorescence intensity at excitation/emission 530/590 nm, is directly proportional to the creatine concentration in the sample.
| Product Name: | Creatine Assay Kit (Colorimetric/Fluorometric) (BA0095) |
| SKU: | BA0095 |
| Detection Method: | Colorimetric or Fluorometric |
| Detection Range: | Colorimetric 4 to 1000 uM; fluorometric 0.5 to 50 uM creatine |
| Sample Type: | Serum, plasma, urine, saliva and other biological samples |
| Species Reactivity: | All |
| Assay Time: | 30 minutes at room temperature |
| Kit Size: | 100 Assays |
| Equipment Required: | Microplate reader |
| Storage: | -20C |
| Shelf Life: | 12 months after receipt |
| Shipping: | Gel Pack |
Quantitative colorimetric or fluorometric determination of creatine. Enzymatic reactions form a pink product read at 570 nm or by fluorescence at excitation/emission 530/590 nm; the signal is proportional to creatine concentration.
- High sensitivity and wide linear range; uses 10 uL sample
- Colorimetric linear detection range 4 to 1000 uM
- Fluorometric linear detection range 0.5 to 50 uM creatine
- Homogeneous, simple mix-and-measure procedure completed within 30 minutes
- Direct assays of creatine in biological samples such as serum, plasma, urine and saliva
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 | Sample preparation. Avoid SH-group-containing reagents (e.g. mercaptoethanol, DTT) and EDTA. Extract solid samples by homogenisation in distilled water and filter or centrifuge; assay liquid samples such as serum, plasma and urine directly. |
| 2 | Standards and samples (colorimetric). Equilibrate all components to room temperature and briefly centrifuge tubes. Prepare a 1000 uM creatine standard premix by mixing 15 uL of the 20 mM standard and 285 uL distilled water, then dilute as shown in the dilution table. Transfer 10 uL standards into separate wells of a clear flat-bottom 96-well plate. Transfer 10 uL of each sample into two separate wells, one serving as a sample blank and one as a sample well. |
| 3 | Enzyme reaction. For each standard and sample well, prepare working reagent by mixing 90 uL Assay Buffer, 1 uL Enzyme A, 1 uL Enzyme B and 1 uL Dye Reagent; add 90 uL to the four standards and the sample wells. Prepare blank control reagent by mixing 90 uL Assay Buffer, 1 uL Enzyme B and 1 uL Dye Reagent (no Enzyme A) and add 90 uL only to the sample blank wells. Tap to mix and incubate 30 minutes at room temperature. |
| 4 | Read OD570nm. |
| 5 | Fluorometric procedure. The procedure is the same as the colorimetric assay, except the detection range is up to 50 uM creatine, a black flat-bottom 96-well plate is used and creatine standards of 0, 15, 30 and 50 uM are prepared. After 30 minutes at room temperature, read fluorescence at excitation 530 nm and emission 590 nm. |
Subtract the standard values from the blank value (#4) and plot dOD or dF against standard concentrations to determine the slope. [Creatine] = (R-sample - R-blank) / slope x n (uM), where R is the optical density or fluorescence reading and n is the sample dilution factor. Conversions: 1000 uM creatine equals 13.1 mg/dL or 131 ppm. If the calculated concentration exceeds 1000 uM (colorimetric) or 50 uM (fluorometric), dilute the sample in distilled water, repeat and multiply by the dilution factor.
| Component | Quantity | Storage |
| Assay Buffer | 20 mL | -20C |
| Enzyme A | 120 uL | -20C |
| Enzyme B | 220 uL | -20C |
| Standard (20 mM creatine) | 400 uL | -20C |
| Dye Reagent | 220 uL | -20C |
