Time-resolved Förster Resonance Energy Transfer Assays for Measurement of Endogenous Phosphorylated STAT Proteins in Human Cells

Author(s):  
Jaime Padros ◽  
Geneviève Chatel ◽  
Mireille Caron
Biochemistry ◽  
2005 ◽  
Vol 44 (11) ◽  
pp. 4312-4321 ◽  
Author(s):  
Olivier Dalmas ◽  
Marie-Ange Do Cao ◽  
Miguel R. Lugo ◽  
Frances J. Sharom ◽  
Attilio Di Pietro ◽  
...  

2019 ◽  
Vol 24 (3) ◽  
pp. 386-397 ◽  
Author(s):  
Pierre Baillargeon ◽  
Virneliz Fernandez-Vega ◽  
Banu Priya Sridharan ◽  
Steven Brown ◽  
Patrick R. Griffin ◽  
...  

The Scripps Research Molecular Screening Center (SRMSC) was founded in 2004 and comprises more than $22 million of specialized automation. As part of the Translational Research Institute (TRI), it comprises early drug discovery labs and medicinal chemistry. Together with Scripps Research at the La Jolla, California, campus, this represents one of the most competitive academic industrial screening centers worldwide. The SRMSC uses automated platforms, one a screening cell and the other a cherry-picking platform. Matched technologies are available throughout Scripps to allow scientists to develop assays and prepare them for automated screening. The library comprises more than 1 million drug-like compounds, including a proprietary collection of >665,000 molecules. Internal chemistry has included ~40,000 unique compounds that are not found elsewhere. These collections are screened against a myriad of disease targets, including cell-based and biochemical assays that are provided by Scripps faculty or from global investigators. Scripps has proven competence in all detection formats, including high-content analysis, fluorescence, bioluminescence resonance energy transfer (BRET), time-resolved fluorescence resonance energy transfer (TR-FRET), fluorescence polarization (FP), luminescence, absorbance, AlphaScreen, and Ca++ signaling. These technologies are applied to NIH-derived collaborations as well as biotech and pharma initiatives. The SRMSC and TRI are recognized for discovering multiple leads, including Ozanimod.


2019 ◽  
Vol 122 (3) ◽  
pp. 397-404
Author(s):  
Alexandre Ho-Pun-Cheung ◽  
Hervé Bazin ◽  
Florence Boissière-Michot ◽  
Caroline Mollevi ◽  
Joëlle Simony-Lafontaine ◽  
...  

Abstract Background Triple-negative breast cancer (TNBC) has a worse prognosis compared with other breast cancer subtypes, and biomarkers to identify patients at high risk of recurrence are needed. Here, we investigated the expression of human epidermal receptor (HER) family members in TNBC and evaluated their potential as biomarkers of recurrence. Methods We developed Time Resolved-Förster Resonance Energy Transfer (TR-FRET) assays to quantify HER1, HER2 and HER3 in formalin-fixed paraffin-embedded (FFPE) tumour tissues. After assessing the performance and precision of our assays, we quantified HER protein expression in 51 TNBC specimens, and investigated the association of their expression with relapse-free survival. Results The assays were quantitative, accurate, and robust. In TNBC specimens, HER1 levels ranged from ≈4000 to more than 2 million receptors per cell, whereas HER2 levels varied from ≈1000 to 60,000 receptors per cell. HER3 expression was very low (less than 5500 receptors per cell in all samples). Moderate HER2 expression was significantly associated with higher risk of recurrence (HR = 3.93; P = 0.003). Conclusions Our TR-FRET assays accurately quantify HER1, HER2 and HER3 in FFPE breast tumour specimens. Moderate HER2 expression may represent a novel prognostic marker in patients with TNBC.


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