TR-FRET Bridge-It® L-Tryptophan Fluorescence Assay Kit for Plasma and Serum Samples, 384-well format

L-Met 1-1-1006B

Clear selection

Description

TR-FRET Bridge-It® L-Tryptophan Fluorescence Assay Kit for Plasma and Serum Samples, 384-well 

 

Application:

  • Quantification of Tryptophan levels in plasma and serum samples

  • Measurement of total tryptophan in plasma and serum samples

  • High throughput screening for drug discovery (please contact us for bulk discount pricing)

  • Diet and nutrition, mental health, and immunology studies

 

Features:

  • Easy: Treat samples to extract tryptophan, incubate, and mix test sample or standard with the Assay solution and incubate at ~25°C. No wash steps!

  • Fast: Read fluorescent signal after 30 minutes of incubation (can be read after 8 minutes)

  • Sensitive: Assay measures tryptophan at a lower limit of sensitivity of 0.1 µM

  • (i.e. 10 pmol / well in a 96-well or 2 pmol in a 384-well black microplate)

  • Flexible: Assay is adaptable to both low- and high-throughput screening formats

  • Reproducible: Inter- and Intra-assay values are 10% or less

  • High Signal to Background Ratio: TR-FRET option of the Bridge-It Assay kit gives high signal to background ratio (8+ fold) which is useful for high throughput screening and drug discovery

 

 

Comparison to ELISA: 

Bridge-It® Assay for Plasma and Serum Samples: 4 reagents, 1 hour

ELISA for Plasma and Serum Samples: 19 reagents, 18+ hours work with overnight incubation

 

TRF vs ELISA timeline

 

Background Information on Tryptophan

L-Tryptophan

L-tryptophan (tryptophan) is one of eight essential amino acids that must be obtained from the diet. Tryptophan serves as a key building block for synthesis of proteins and as a precursor for various brain neurotransmitters including serotonin. Tryptophan is the only recognized precursor that can be converted into serotonin by the body. Serotonin promotes feelings of well being and calm and thereby helps to counterbalance the physiological effects of brain dopamine and the nor-adrenaline circuits which encourage fear, anger, tension, aggression, obsessive-compulsive actions, over-eating (especially of carbohydrates), migraine headache, depression, and sleep disturbances. Melatonin, a metabolite of serotonin, is a sleep promoting natural hormone made by the pineal gland. In addition, tryptophan is a key precursor for niacin (vitamin B3), a vitamin that is essential for normal respiration, metabolism, and synthesis of sex hormones. Because tryptophan plays such a critical role in the proper balancing of metabolism, mood and sleep patterns, insufficient dietary availability of this essential amino acid can lead to serious adverse consequences.

 

Bridge-It® L-Tryptophan Fluorescence Assay

The Bridge-It® tryptophan fluorescence assay is based on the activity of tryptophan repressor protein (TrpR), a bacterial DNA-binding protein. TrpR protein binds to its DNA-binding site in tryptophan-dependent fashion.

The central feature of this assay design is the TrpR-dependent association of two fluorochrome-labeled DNA half-fragments (one labeled with fluorescein and the other labeled with Oyster® 645 fluophore. Each fragment contains about one-half of the TrpR protein DNA-binding site. In the presence of L-tryptophan an increase in fluorescence signal can be detected as a result of the tryptophan-dependent association of the labeled DNA half-fragments. Tryptophan is readily detectable using the Bridge-It® tryptophan fluorescence assay in various types of test samples including bacterial growth medium, brain extract, yeast extract, as well as in human serum and urine. The linear range of the assay is 0.4 µM-10 µM and the minimum tryptophan detection level is ~0.1 µM. The assay is highly specific for measuring tryptophan. No significant TrpR protein binding activity was observed using the assay when L-tryptophan was replaced with each of nineteen (19) other L-amino acids (up to 100 µM) or D-tryptophan, serotonin, and the tryptophan precursor 5’HTP (up to 20 µM).

TRF Trp Standard Curve Pic

 

The TR-FRET Bridge-It® L-tryptophan fluorescence assay for plasma and serum samples is performed using the 96-well or 384-well microplate format and is, therefore, ideally suited for the rapid, simultaneous measurement of tryptophan in large numbers of test samples.

 

Pricing: The 100-well kit is a one-time trial purchase. For bulk pricing options, please contact us. For any other questions or comments, please do not hesitate to contact us. We are always happy to help!

 

To learn more about the TR-FRET Bridge-It technology, please visit our technology page. This product is protected patents and pending patents.

 


Selected References Using Mediomics’ Bridge-It L-Tryptophan Fluorescence Assay Kit

As this product is very new, currently no references for the TR-FRET assay have been generated. Below are references for the FRET Bridge-It Tryptophan Assay.

1. Röhrig UF, Majjigapu SR, Vogel P, Zoete V, Michielin O. Challenges in the Discovery of Indoleamine 2,3-Dioxygenase 1 (IDO1) Inhibitors.J. Med. Chem. 2015 May 13.

2. Chang CK, Chang Chien KM, Chang JH, Huang MH, Liang YC, Liu TH. Branched-Chain Amino Acids and Arginine Improve Performance in Two Consecutive Days of Simulated Handball Games in Male and Female Athletes: A Randomized Trial. PLOS One. 2015 March 24.

— “Plasma free tryptophan concentration was analyzed with a fluorescence assay according to manufacturer’s recommendation (Bridge-It, Mediomics, St. Louis, MO, USA).”

3. Piesiewicz A, Kedzierska U, Turkowska E, Adamska I, Majewski PM Seasonal postembryonic maturation of the diurnal rhythm of serotonin in the chicken pineal gland. Chronobiol Int. 2014 Sep 15.

— “…duplicate 10 µl aliquots were taken to measure TRP concentration with a Bridge-It L-tryptophan Fluorescence Assay Kit (Mediomics, St. Louis, MO).

4. Urata Y, et al. IL-1β Increases Expression of Tryptophan 2,3-dioxygenase and Stimulates Tryptophan Catabolism in Endometrioma Stromal Cells. AJRI, 26 Jun 2014.

— “Concentration of tryptophan in conditioned media was measured using Bridge-It L-Tryptophan Fluorescence Assay (Mediomics, St. Louis, MO, USA) according to the manufacturer’s protocol.”

5. Piesiewicz A, Kedzierska U, Adamska I, Usarek M, Zeman M, Skwarlo-Sonta K, Majewski PM. Pineal arylalkylamine N-acetyltransferase (Aanat) gene expression as a target of inflammatory mediators in the chicken. Gen Comp Endocrinol. 2012 Nov 1;179(2):143-51.

— “Pineal glands were sonicated (Bandelin electronic) in 25 μl of ice-cold sodium phosphate buffer (50 mM, pH 6.8) and duplicate 10 μl aliquots were taken to measure TRP concentration using a Bridge-It l -tryptophan Fluorescence Assay Kit (Mediomics).

6. Bacchus W, Lang M, El-Baba MD, Weber W, Stelling J, Fussenegger M. Synthetic two-way communication between mammalian cells. Nat Biotechnol. 2012 Oct;30(10):991-6.

— “L-tryptophan levels in the medium and serum were profiled using a fluorescence-based assay kit (Mediomics, St. Louis, MO, USA, cat. no. 1-1-1001B).”

7. Meininger D, Zalameda L, Liu Y, Stepan LP, Borges L, McCarter JD, Sutherland CL. Purification and kinetic characterization of human indoleamine 2,3-dioxygenases 1 and 2 (IDO1 and IDO2) and discovery of selective IDO1 inhibitors. Biochim Biophys Acta. 2011 Dec;1814(12):1947-54.

— “…were determined by measuring the production of the kynurenine product by kynurenine fluorescence or by the consumption of the Trp substrate using the Bridge-IT tryptophan fluorescence assay (Mediomics, St. Louis, MO).”

8. Piesiewicz A, Podobas E, Kedzierska U, Joachimiak E, Markowska M, Majewski P, Skwarlo-Sonta K. Season-Related Differences in the Biosynthetic Activity of the Neonatal Chicken Pineal Gland. The open Ornithology journal, 2010, 3, 134-140.

— “L-tryptophan was measured in duplicate pineal sonicate samples using a Bridge-It L-Tryptophan Fluorescence Assay Kit (Mediomics), according to the manufacturer’s protocol.”

 


 

  1. Bridge-It® is a registered trademark of Mediomics, LLC, St. Louis, Missouri, U.S.A. Mediomics has a worldwide, exclusive license for this assay platform from Saint Louis University, St. Louis, Missouri.

  2. The TR-FRET Bridge-It® tryptophan fluorescence assay is intended for laboratory research use only. This product is not approved by the U.S. Government or by the government of any other country of the world for use in disease diagnosis or treatment of humans or animals.

  3. Oyster® is a registered trademark of Denovo Biolabels, GmbH, Munster, Germany.

  4. Flownamics® Analytical Instruments, Inc., Madison, Wisconsin, is the authorized U.S. distributor for Oyster® dyes.

 

 

Catalog #: TRF-1-1-1002TRP-PS384A (100 measurements-trial kit) , TRF-1-1-1002TRP-PS384B  (384 measurements)

Protocol : 

MSDS :