Super happy that we have finally put this work out!!! 4 years in the making:.We present – DynaScreen: an imaging (FRET-FLIM) based pooled CRISPR screening platform for studying “dynamic” phenotypes. short🧵below (1/n)

If you have seen @sravmukherjee or Kees presenting this work at multiple conferences – this is the final product of our beloved project. We are looking forward to perform many more screens with this robust pipeline. Please give it a read & we are very happy to receive feedback!.

💡We successfully optimised samples for NGS with extremely low cell counts (~200 cells).💡We show why single cell measurements (via imaging) are important in comparison to bulk cell measurements (ELISAs/Western Blots) (8/n).

Other highlights of the paper –.💡We use the PA-JF646 dye augmented with some NH4Cl treatment as the photo-tagging method to selectively tag “aberrant cells”. No cloning, no stable cell lines & super bright! Just load 30 min before your experiment and you're good to go! (7/n).

Another big highlight of the paper – FAST HIPPOS (now available at – !! What an incredibly cool open-source software built by Bram van den Broek to handle image analysis, automated hit identification & data visualization and inspection! Check it out! (6/n).

With 2 pooled CRISPR screens, we have not only identified genes that shape the main cAMP signaling axis like ADRB2, GNAS, ADCY6, PDE1C we also found genes such as VPS26A, SNX13, SSTR1 and RGS14 to be regulating cAMP breakdown and basal levels. (5/n).

We’ve taken cAMP signaling as an example for this study and together with a custom designed gRNA library, we tracked real-time cAMP dynamics in response to agonist stimulation and identified genes that modulate its basal levels and response kinetics. (4/n).

With our (super) FLIM biosensors, photoactivation-based single-cell tagging and pooled CRISPR screening technology, we’ve established a scalable system to identify genes that regulate the timing, amplitude, and duration of signaling responses. (3/n).

We see a lot of high-content imaging-based genetic screens these days, but they have remained largely limited to static or binary phenotypes. (2/n).

RT @pablo_jack: Brilliant work improving the Epac based biosensors. During my pHD in Groningen I used Jalink biosensors and they always del….

🥳🥳🥳 let's go team!! 🤓🔬🧫.

🔬 We are excited to make these powerful tools available to the microscopy community and look forward to seeing how they enable deeper insights into cellular signaling and real-time dynamics in diverse research applications!!!.

These sensors, Epac-based, show improved cytosolic localization while maintaining excellent performance. They are ideal for co-expression with other FRET/FLIM sensors, offering greater flexibility in live-cell imaging experiments.

Our latest preprint from the lab is now OUT!!! 🧫🔬. Spearheaded by our PhD student Giulia Zanetti - we made cytosolic-enhanced dedicated FLIM sensors for intracellular cAMP detection! . Check it out and a small thread below 👇.

And last but not the least, this study shows that in this case, confocal TCSPC provides the more reliable data needed to study response kinetics.

The practice that recipes are kept secret and kits are offered as black-boxes solely for commercial benefit is simply not in accordance with good scientific practice which aims at a full, exact and reproducible description of experiments.

These data also strongly emphasize the crucial importance of exact knowledge on the molecular composition of reagents, buffers and media.

This study should caution researchers who perform live-cell time-lapse experiments to be aware about sample health, phototoxicity and image acquisition routines.

Our paper is out!!! (well already 3 months ago) 🔬🧫.A project spearheaded by @sravmukherjee and a very fun almost detective novel like story. Check it out!!! We also managed to put a pun in the title 😉. small thread below 👇.

RT @MicroSensing: @drmgmonaghan @FlImagin3_DN @tcddublin @MSCActions @JalinkLab @schenkelayland @raimondi_t @Femtoscopist @PicoQuant @Leica….