Fluorescence responsiveness of unicellular marine algae Dunaliella to stressors under laboratory conditions.

Nanoparticles (NPs) proved to have numerous applications in various fields, including biomedicine and environmental sciences. In this work, we designed and created an apparatus for fabrication of metal NPs directly in liquids initiated by femtosecond laser pulses. The laser parameters leading to ~10 μJ/pulse energy and 0.1 GW peak power resulted in predominantly spherical particles with the sizes varying from <10 nm to ~100 nm in diameter. NPs generated from Cobalt and Zinc targets were smaller in order of magnitude compared to that of Nickel. The fabricated NPs were characterized by scanning electron microscopy, while spectroscopic properties were investigated by absorption spectroscopy and spectrally resolved fluorescence imaging. We also tested the possible interaction of the created NPs with living algae for their potential use for environmental research. Employing such ultrashort laser opens route to provide on-demand production of NP's in-situ at even factory environment.

Preparation of metal nanoparticles by femtosecond laser ablation

Pollution by heavy metals represents a significant environmental burden. We employed confocal microscopy with spectral detection and fluorescence lifetime imaging (FLIM) to evaluate the effect of nanoparticles (NPs) from various metals (Zinc, Nickel, Cobalt, Copper) on endogenous fluorescence of Fontinalis antipyretica moss. Short term (3-5 day) exposure to NPs, designed and fabricated by direct synthesis using femtosecond laser ablation in water, was studied. The green flavonoid and/or lignin endogenous fluorescence peaking between 500-560 nm was found increased by Zn and significantly reduced by Cu. The overall red chlorophyll fluorescence intensity with maximum at 680 nm remained largely unchanged after exposure to Ni and Zn, but was decreased in the presence of Co and completely abolished by Cu. All NPs but Zn induced changes in the fluorescence lifetimes, demonstrating increased sensitivity of this parameter to environmental pollution. Gathered data indicate fast responsiveness of the endogenous fluorescence in the Fontinalis antipyretica moss to the presence of heavy metals that can thus potentially serve as a biosensing tool for monitoring environmental pollution in the moss natural environment..

/pdf/biosensing-the-presence-of-metal-nanoparticles-by-spectrally-2cvy608cqg.pdf

Biosensing the presence of metal nanoparticles by spectrally- and time-resolved endogenous fluorescence in water moss Fontinalis antipyretica.

To better understand pH-dependence of endogenous fluorescence of algae, we employed spectroscopy and microscopy methods, including advanced time-resolved fluorescence imaging microscopy (FLIM), using green algae Chlorella sp. as a model system. Absorption spectra confirmed two peaks, at 400-420 nm and 670 nm. Emission was maximal at 680 nm, with smaller peaks between 520 and 540 nm. Acidification led to a gradual decrease in the red fluorescence intensity with the maximum at 680 nm when excited by 450 nm laser. FLIM measurements, performed using 475 nm picoseconds excitation, uncovered that this effect is accompanied by a shortening of the tau1 fluorescence lifetime. Under severe acidification, we also noted an increase in the green fluorescence with a maximum between 520-540 nm and a shift toward 690-700 nm of the red fluorescence, accompanied by prolongation of the tau2 fluorescence lifetime. Gathered data increase our knowledge on the responsiveness of algae to acidification and indicate that endogenous fluorescence derived from chlorophylls can potentially serve as a biosensing tool for monitoring pH change in its natural environment.

https://iopscience.iop.org/article/10.1088/2050-6120/ab77f4/pdf

Time-resolved endogenous chlorophyll fluorescence sensitivity to pH: study on Chlorella sp. algae.

The last decade has witnessed a rapid growth of nanoscale-oriented biosensors that becomes one of the most promising and rapidly growing areas of modern research. Despite significant advancements in conception of such biosensors, most are based at evaluation of molecular, or protein interactions. It is however becoming increasingly evident that functionality of a living system does not reside in genome or in individual proteins, as no real biological functionality is expressed at these levels. Instead, to comprehend the true functioning of a biological system, it is essential to understand the integrative physiological behavior of the complex molecular interactions in their natural environment and precise spatio-temporal topology. In this contribution we therefore present a new concept for creation of biosensors, bio-inspired from true functioning of living cells, while monitoring their endogenous fluorescence, or autofluorescence.

A bio-inspired design of live cell biosensors

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A bio-inspired design of live cell biosensors