AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit
AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit is designed to provide precise and reliable measurements of alpha-ketoglutarate (α-KG) levels in biological samples. This assay kit offers both colorimetric and fluorometric detection methods, making it a versatile tool for researchers in the fields of biochemistry, cell biology, and molecular biology.
Key Features:
- High Sensitivity: Our assay kit is optimized to detect even low levels of α-KG, allowing for accurate quantification in a wide range of samples.
- Colorimetric and Fluorometric Detection: With dual detection options, our kit provides flexibility and versatility to meet your experimental needs.
- Wide Application: The AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit can be used with various sample types, including cell lysates, tissue homogenates, and purified enzyme preparations.
- Quick and Easy: With a simple and streamlined protocol, you can obtain results in just a few easy steps, saving you valuable time in the lab.
- Robust Performance: Our kit ensures robust and reproducible results, minimizing the margin for error and enhancing the reliability of your data.
Assay Principle:
This assay kit employs a method based on the enzymatic conversion of α-KG into a product that reacts with a probe to generate a colorimetric or fluorometric signal. The intensity of the signal is directly proportional to the α-KG concentration in your sample, allowing for precise quantification.
Applications:
- Metabolic Research: Study the role of α-KG in various metabolic pathways, including the tricarboxylic acid (TCA) cycle and amino acid metabolism.
- Cellular Signaling: Investigate the involvement of α-KG in cellular signaling pathways, such as hypoxia-inducible factor (HIF) regulation and epigenetic modifications.
- Disease Biomarker: Assess α-KG levels as a potential biomarker for diseases such as cancer, metabolic disorders, and neurodegenerative diseases.
Kit Contents:
- Assay Buffer
- α-Ketoglutarate Probe
- α-Ketoglutarate Enzyme Mix
- Developer Solution
- 96-Well Assay Plate
- Detailed Protocol
Why Choose AffiASSAY®?
Our commitment to excellence and scientific rigor makes AffiASSAY® the trusted choice of researchers worldwide. We strive to provide you with innovative tools that deliver precise results and accelerate your research efforts.
AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit Protocol:
[Please note that this protocol is a general guideline, and you should always refer to the specific instructions provided in the kit manual for accurate and detailed information.]
Materials Needed:
- AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit
- Biological samples containing α-KG
- 96-well microplate
- Microplate reader capable of measuring absorbance or fluorescence intensity at specific wavelengths
- Pipettes and tips
- Microcentrifuge tubes
Protocol:
Sample Preparation:
- Prepare your biological samples containing α-KG. These samples can be cell lysates, tissue homogenates, or purified enzyme preparations.
- Ensure that your samples are stored and handled according to your specific experimental requirements.
Assay Plate Setup:
- Label the microplate wells according to your experimental design, including wells for your samples, standards, and controls.
- If using controls, prepare control samples with known α-KG concentrations.
Assay Reaction:
- Prepare the assay reaction mix by combining the Assay Buffer, α-Ketoglutarate Probe, and α-Ketoglutarate Enzyme Mix from the kit as instructed.
- Add 50-100 µL of the assay reaction mix to each well of the microplate.
Sample and Control Addition:
- Add 50-100 µL of your prepared samples and control samples to the appropriate wells on the microplate.
- Ensure that the final reaction volume in each well is consistent.
Incubation:
- Incubate the microplate at the appropriate temperature (as specified in the kit instructions) for a recommended time to allow the enzymatic reaction to occur.
Measurement:
- For colorimetric detection: Measure the absorbance of each well at a specific wavelength (provided by the kit) using a microplate reader.
- For fluorometric detection: Measure the fluorescence intensity of each well using a fluorescence microplate reader with appropriate excitation and emission wavelengths (provided by the kit).
Data Analysis:
- Calculate the α-KG concentration in your samples based on their respective absorbance or fluorescence intensity readings.
Results Interpretation:
- Interpret and analyze your results according to your research objectives.
- Note any variations or trends in α-KG levels among your samples.
Data Presentation:
- Prepare your data for presentation or publication, and include the appropriate statistical analysis if required.
Cleanup:
- Dispose of all waste materials, including used assay components, according to your laboratory's safety protocols.
- Always refer to the specific instructions provided with the AffiASSAY® Alpha-Ketoglutarate Colorimetric & Fluorometric Assay Kit for any kit-specific details or variations in the protocol.
- Additionally, ensure that you adhere to the best practices and safety guidelines in your laboratory while conducting the assay.
