EnzyChrom™ L-Amino Acid Assay Kit

Protocol SDS

Application

• For quantitative determination of L-Amino Acids in biological and other samples.

Key Features

• Fast and sensitive. Linear detection range: 3.3 to 500 µM (colorimetric assay) and 0.13 to 50 µM (fluorimetric assay) for 60 min reaction.
• Convenient. The procedure involves adding a single working reagent and reading after 60 minutes.
• High-throughput. Homogeneous “mix-incubate-measure” type assay. Can be readily automated to process thousands of samples per day.

Method

• OD570nm or FL530/585nm

Samples

• Serum, culture media, tissue homogenates, cell lysates, urine, food samples, etc

Species

• All

Procedure

• 60 min

Size

• 100 tests

Detection Limit

• 3.3 µM (colorimetric assay), 0.13 µM (fluorimetric assay)

Shelf Life

• 6 months

More Details

L-AMINO-ACIDS are the building blocks of proteins in biology. Almost all of the common 20 amino acids exist as the L-enantiomer. BioAssay Systems’ L-amino acid assay uses an enzyme-catalyzed oxidation of L-amino acids to convert a dye into a colored and fluorescent form. The absorbance at 570 nm or fluorescence intensity at λ_ex/em = 530/585 nm is directly proportional to the L-amino acid concentration in the sample.

Can urine samples be used with this kit?

Urine samples should be diluted at least 5-fold with dH₂O and an internal standard should be used.

Samples will need three separate reactions:
1) sample plus standard
2) sample alone
3) sample blank.

For the internal standard prepare 200 µL 1250 µM standard by mixing 125 µL 2 mM Standard and 75 µL dH2O.

For the sample plus standard well, add 5 µL 1250 µM standard and 20 µL sample.

For the sample and sample blank wells, add 5 µL dH₂O and 20 µL sample.

Prepare enough Working Reagent (WR) for all assay wells by mixing, for each well, 85 µL Assay Buffer, 1 µL LAA Enzyme, 1 µL HRP Enzyme, 1 µL Dye Reagent. Prepare a WR without the LAA Enzyme for the sample blank well(s).

Add 80 µL of the WR to each well. Tap plate briefly to mix.

Incubate at room temperature for 60 min. Use a plate reader to read OD_570nm.

The sample L-amino acid concentration is computed as follows:

where R_SAMPLE, R_BLANK, and R_STANDARD are the OD or fluorescence values of the Sample, Sample Blank, and the Sample plus Standard respectively. n is the sample dilution factor. Note: The volume of the internal standard is 4× lower than the sample volume; thus, the internal standard concentration should be divided by 4.

What samples have you tested?

This kit has been tested on serum, culture media, mammalian cell lysates, urine, and solid food samples (e.g. ground beef).

Can I store unused reagents for future use?

Yes, unused reagents can be stored according to the assay protocol.

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If you or your labs do not have the equipment or scientists necessary to run this assay, BioAssay Systems can perform the service for you.

– Fast turnaround
– Quality data
– Low cost

Please email or call 1-510-782-9988 x 2 to discuss your projects.