TRFS-green

• CAT Number: I045574
• CAS Number: 1513848-14-0
• Molecular Formula: C20H20N2O4S2
• Molecular Weight: 416.51
• Purity: ≥95%
• Synonyms: dithiolan-4-yl N-(2-butyl-1,3-dioxobenzo[de]isoquinolin-6-yl)carbamate
• InChI: InChI=1S/C20H20N2O4S2/c1-2-3-9-22-18(23)14-6-4-5-13-16(8-7-15(17(13)14)19(22)24)21-20(25)26-12-10-27-28-11-12/h4-8,12H,2-3,9-11H2,1H3,(H,21,25)
• InChIKey: MUAYCZWBPDUZKR-UHFFFAOYSA-N
• SMILES: CCCCN1C(=O)C2=C3C(=C(C=C2)NC(=O)OC4CSSC4)C=CC=C3C1=O

TRFS-green is a highly selective off−on fluorescent probe for imaging selenoprotein thioredoxin reductase (TrxR) in living cells. TRFS-green has the maximum absorbance at around 373 nm. After it is activated by TrxR, the maximum absorbance shifts to around 440 nm[1].
TRFS-green displays a green fluorescence off−on change induced by TrxR-mediated disulfide cleavage and subsequent intramolecular cyclization to liberate the masked naphthalimide fluorophore[1].
TRFS-green gives a signal at ~480 nm when excited at 377 nm. Pretreatment of living cells with TrxR inhibitors in a dose-dependent manner can inhibit the fluorescence signal of TRFS-green in living cells, thereby facilitating the development of living cell-based screening assays for TrxR inhibitors[1].
TRFS-green shows no response to GSH or Cys, the predominant thiols in cells, or the endogenous reducing agents, such as vitamin C and NADPH[1].
Measure TrxR activity in crude cell extracts[1]: Incubate cell lysates with TRFS-green for an appropriate time (usually 1-2 hours), and measure fluorescence intensity by fluorimeter (Em= 540 nm, Ex=440 nm). It is also possible to monitor the time-dependent increase in fluorescence (Em=540 nm, Ex=440 nm) over a given period of time.
Cells treated with TRFS-green (10 μM; 2-4 h) can be observed through the green fluorescent channel of the microscope[1].
Note: In order to obtain a clear fluorescence image, it is recommended to wash the cells with PBS or fresh medium immediately before taking pictures to remove residual TRFS-green.
TRFS-green is the first fluorescent probe of TrxR with a green emission. However, the response rate of TRFS-green to the enzyme is slow, and it takes more than 2 h to reach the maximal fluorescence signal even using tris(2-carboxyethyl)phosphine (TCEP) as a reducing agent[2].

Reference

[1]. Liangwei Zhang, et al. Highly selective off-on fluorescent probe for imaging thioredoxin reductase in living cells. J Am Chem Soc. 2014 Jan 8;136(1):226-33.
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[2]. Xinming Li, et al. A fast and specific fluorescent probe for thioredoxin reductase that works via disulphide bond cleavage. Nat Commun. 2019 Jun 21;10(1):2745.
 [Content Brief]