A Single Fluorescent Protein-Based Indicator with a Time-Resolved Fluorescence Readout for Precise pH Measurements in the Alkaline Range
The real-time monitoring of the intracellular pH in live cells with high precision represents an important methodological challenge. Most of the genetically encoded fluorescent indicators are hindered by the inability to determine an absolute pH value and/or a narrow dynamic range of the signal. The team of scientists from the Laboratory of genetically encoded molecular tools IBCh RAS studied the pH sensitivity of a green-fluorescence-protein (GFP)-based emitter (EGFP-Y145L/S205V) with the alkaline-shifted chromophore’s pKa and demonstrate that, in the pH range of 7.5–9.0, its fluorescence lifetime changes by a factor of ~3.5 in a quasi-linear manner in mammalian cells. Considering the relatively strong lifetime response in a narrow pH range, authors proposed the mitochondria, which are known to have a weakly alkaline milieu, as a target for live-cell pH measurements. Using FLIM they succeeded in determining the absolute pH value of the mitochondria and recorded the ETC-uncoupler-stimulated pH shift with a precision of 0.1 unit. Thus the scientists showed that a single GFP with alkaline-shifted pKa can act as a high-precision indicator that can be used in a specific pH range. The work is published in the International Journal of Molecular Sciences. Learn more
- Relationship between phenolic compounds and antioxidant activity in berries and leaves of raspberry genotypes and their genotyping using SSR markers
In recent years, the enrichment of fruits and berries with biologically active substances, in particular, antioxidants has received increased development in breeding (the creation of functional food). Researchers from the Forest Biotechnology Group of the FIBCh, together with colleagues from other institutes, studied the content of total phenols, flavonoids and anthocyanins, their relationship with the antioxidant activity of berries and leaves of various raspberry cultivars of Russian and foreign origin, as well as new breeding lines.
- A Single Fluorescent Protein-Based Indicator with a Time-Resolved Fluorescence Readout for Precise pH Measurements in the Alkaline Range
The real-time monitoring of the intracellular pH in live cells with high precision represents an important methodological challenge. Although genetically encoded fluorescent indicators can be considered as a probe of choice for such measurements, they are hindered mostly by the inability to determine an absolute pH value and/or a narrow dynamic range of the signal, making them inefficient for recording the small pH changes that typically occur within cellular organelles.
- Longitudinal profiling of human peripheral blood B cell clonal repertoire: memory B cell persistence and signs of negative and positive selection at the BCR repertoire level
Persistence of B cell clones and fine-tuning of their BCRs allow them to respond effectively to repeated immune challenges. Researchers from the Genomics of adaptive immunity department of IBCH in collaboration with colleagues from IITP and Skoltech, performed long-term study of structure and dynamics of BCR repertoire of memory B cells and antibody-secreting cells from peripheral blood of healthy donors.